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T cells have a central role in allograft rejection. The specificity of the alloimmune response is determined by the interaction of the major histocompatibility complex (MHC) molecule on antigen-presenting cells with the T-cell receptor (TCR) on T cells, consisting of ‘signal 1'. However, in order to fully activate a naïve T cell, a second antigen-independent co-signal must be delivered by co-stimulatory molecules (signal 2). Therefore, co-stimulation has a key role in determining the outcome of the T-cell encounter with the alloantigen, with important therapeutic applications in transplantation.1, 2

r, in order to fully activate a naïve T cell, a second antigen-independent co-signal must be delivered by co-stimulatory molecules (signal 2). Therefore, co-stimulation has a key role in determining the outcome of the T-cell encounter with the alloantigen, with important therapeutic applications in transplantation.1, 2 The best characterized co-stimulatory pathway is the B7:CD28 pathway. In both, mice and humans, CD28 is constitutively expressed on all naïve CD4+ and CD8+ T cells,3 and it can interact with two ligands, B7.1 (CD80) and B7.2 (CD86), expressed on antigen-presenting cells (Figure 1a). In the presence of TCR signaling, B7:CD28 interaction leads to full activation and expansion of T cells, whereas blocking this pathway results in anergy and/or apoptosis of responding T cells (Figure 1b).4 In the transplant setting, initial studies in experimental animal models with blockade of B7 ligands revealed promising results. However, the continuous expansion of co-stimulation knowledge led to some unexpected findings.5 Additional co-stimulatory molecules were discovered that shared ligands with each other, and some of these receptors demonstrated capability of inhibiting rather than activating T cells. For example, cytotoxic T-lymphocyte antigen-4 (CTLA4) was found to be structurally related to CD28 and bind to the same ligands on antigen-presenting cells (B7.1 and B7.2) as CD28; however, its interaction with B7 ligands led to inhibition of T-cell activation (Figure 1c). Moreover, other positive co-stimulatory pathways were discovered with non-redundant and compensatory roles in T-cell activation.5 Currently, it is clear that the integration of multiple positive and negative co-stimulatory signals ultimately determines the outcome of the T-cell response.5 In this review, we will focus on the B7:CD28 co-stimulatory pathway and discuss the evolution of this therapeutic target from bench to bedside, and then back to the bench, in an attempt to understand and explain recent clinical outcomes in kidney transplantation.

timately determines the outcome of the T-cell response.5 In this review, we will focus on the B7:CD28 co-stimulatory pathway and discuss the evolution of this therapeutic target from bench to bedside, and then back to the bench, in an attempt to understand and explain recent clinical outcomes in kidney transplantation. B7:CD28 BLOCKADE: FROM BENCH TO BEDSIDE After an alloantigen encounter, B7:CD28 signaling was shown to help in fully activating T cells by increasing transcription and mRNA stability of interleukin-2 (IL-2),6 elevating the expression of antiapoptotic molecules such as Bcl-XL,7 and decreasing the threshold of T-cell receptor activation.8 After failed attempts to develop an effective CD28 blocking antibody,9 researchers were able to successfully target CD28 ligands with antibodies against B7.1 and B7.2.10, 11 In the early 1990s, a recombinant fusion protein, CTLA4-Ig, was developed by fusing the extracellular domain of human CTLA4 with an Ig heavy chain tail.12 This antibody had a higher affinity to the B7 ligands than CD28, and was shown to be a powerful inhibitor of T-cell activation in vitro.12 Subsequent testing of CTLA4-Ig revealed its capability of protecting the allograft against acute rejection in MHC-mismatched cardiac transplantation, as well as in islet cell transplantation in murine models.13, 14 Despite its potent effect, CTLA4-Ig alone was incapable of inducing tolerance in non-human primates,15, 16 requiring additional immunosuppression to promote graft survival.17

the allograft against acute rejection in MHC-mismatched cardiac transplantation, as well as in islet cell transplantation in murine models.13, 14 Despite its potent effect, CTLA4-Ig alone was incapable of inducing tolerance in non-human primates,15, 16 requiring additional immunosuppression to promote graft survival.17 Further mechanistic studies revealed that CTLA4-Ig was 100-fold less potent in inhibiting B7.2 co-stimulation compared with B7.1 co-stimulation,18 possibly explaining its lower-than-expected potency in vivo. Therefore, a modification of this antibody was undertaken, with substitution of two amino acids within the B7.2 binding domain, creating a second generation of CTLA4-Ig (LEA29Y), which was shown to have higher affinity to both B7.1 and B7.2, translating into a 10-fold increase in biological potency.18 LEA29Y, later named belatacept (Bristol–Myers Squibb, New York, NY), was tested in non-human primates and showed superior prolongation in renal allograft survival as monotherapy, when compared with the first-generation CTLA4-Ig, and led to a marked improvement in survival when used in combination with other immunosuppressive regimens such as mycophenolate mofetil and steroids, or an anti-IL-2 receptor antibody.18

mates and showed superior prolongation in renal allograft survival as monotherapy, when compared with the first-generation CTLA4-Ig, and led to a marked improvement in survival when used in combination with other immunosuppressive regimens such as mycophenolate mofetil and steroids, or an anti-IL-2 receptor antibody.18 On the basis of these encouraging results, belatacept moved to a phase II clinical trial to evaluate the efficacy of this drug in kidney transplant recipients in comparison with cyclosporine (CsA).19 During this trial, recipients were assigned to receive either an intensive or a less-intensive regimen of belatacept, compared with CsA. Induction therapy consisted of basiliximab (IL-2 receptor monoclonal antibody), and maintenance therapy included steroids and mycophenolate mofetil. The results of this trial suggested that belatacept was non-inferior to CsA and it could potentially have a beneficial effect on glomerular filtration rate at 1 year after transplantation, presumably by the absence of calcineurin inhibitor-induced nephrotoxicity.19 Subsequently, a phase III clinical trial (BENEFIT) was published, in which an unexpected higher rate of acute rejection was observed in the belatacept group, especially in the intensive arm receiving more frequent doses (22 vs 7% on CsA arm).20 Moreover, these rejections were more severe than the ones with CsA, with most of them with the Banff grades of IIA or higher. Nevertheless, the belatacept groups had a similar graft survival at 1 year and demonstrated superior renal function when compared with CsA.20 Overall, the unexpected higher rate of rejection, especially in the more intensive regimen, was intriguing and suggested some unexpected consequences of the intensive B7:CD28 blockade that required further investigation.

similar graft survival at 1 year and demonstrated superior renal function when compared with CsA.20 Overall, the unexpected higher rate of rejection, especially in the more intensive regimen, was intriguing and suggested some unexpected consequences of the intensive B7:CD28 blockade that required further investigation. TARGETING B7:CD28 PATHWAY: BACK TO THE BENCH Regulatory T cells Although targeting B7:CD28 pathway to improve graft survival was becoming a reality, basic knowledge of this pathway and of the alloimmune response kept expanding. The discovery of regulatory T cells as a sub-population of T cells with inhibitory function and capability of controlling the alloimmune response generated great enthusiasm in the transplant community, as inducing this sub-population of T cells could potentially lead to tolerance development.21 Indeed, several groups published initial exciting results with cell-based therapy with regulatory T cells (Tregs) in tolerance induction.22, 23 Importantly, the development and homeostasis of Tregs was shown to be directly dependent on B7:CD28 co-stimulation, and deficiency on this pathway significantly decreased the amount of regulatory T cells in rodents (Figure 2).24 This was a concern, as blocking this pathway could potentially affect Treg generation. Recently, in a single MHC class II-mismatched model of murine cardiac transplantation, in which allografts survive long term because of the emergence of Tregs that inhibit alloreactive T cells, the deficiency of either B7 or CD28 in recipients paradoxically led to an accelerated rejection.25 This effect was related to the significantly lower number of Tregs in the deficient mice, tipping the balance toward more T-effector/memory cells rather than Tregs.

he emergence of Tregs that inhibit alloreactive T cells, the deficiency of either B7 or CD28 in recipients paradoxically led to an accelerated rejection.25 This effect was related to the significantly lower number of Tregs in the deficient mice, tipping the balance toward more T-effector/memory cells rather than Tregs. Therefore, B7:CD28 signal is important not only for the activation of pathogenic effector T cells but also for the generation of regulatory T cells, being the balance of effector T cells and Tregs that ultimately determines the fate of an allograft.26 In an attempt to understand the observations above in rodents, we could hypothesize that in a fully allogeneic mismatched model, the pool of alloreactive T cells is much greater in size than on a single mismatched model; therefore, blockade of B7:CD28 is especially efficient in suppressing the activation and decreasing the alloimmune response in the former (Figure 3a). However, if the pool of alloreactive T cells is smaller, such as in the single mismatch model described above, B7:CD28 blockade could have a more deleterious effect in Tregs than on effector T cells, tipping the balance toward the pathogenic side and precipitating rejection (Figure 3b). The pool size of the alloreactive T cells could potentially correlate in humans with different scenarios, such as in deceased donor recipients and in sensitized patients vs first living-related transplants in a non-sensitized recipient (Figure 3).

ance toward the pathogenic side and precipitating rejection (Figure 3b). The pool size of the alloreactive T cells could potentially correlate in humans with different scenarios, such as in deceased donor recipients and in sensitized patients vs first living-related transplants in a non-sensitized recipient (Figure 3). Although an initial small study did not reveal a significant decrease in peripheral regulatory T cells of patients treated with belatacept in comparison with CsA,27 this finding requires further evaluation, as the degree of mismatch, the sensitization of the recipient, the dose, and timing of belatacept must be taken into account, given that some recipients with well-matched grafts might suffer from the deleterious effects of B7:CD28 blockade on Treg generation. B7:CTLA4 pathway The failure of the CTLA4-Ig to uniformly induce tolerance may also relate, at least in part, to its blocking capacity of B7:CTLA4 interaction, which has been shown to be a critical inhibitory co-stimulatory signal.28 In fact, CTLA4-deficient mice develop a severe systemic autoimmune disorder leading to death at several weeks of age,28, 29 demonstrating a key role of CTLA4 in maintaining self-tolerance. In addition, blockade of CTLA4 with an anti-CTLA4 antibody has been shown to precipitate rejection and prevent induction of allograft tolerance in the transplant setting,30 reinforcing the important role of CTLA4 signaling in inhibiting the alloimmune response.

trating a key role of CTLA4 in maintaining self-tolerance. In addition, blockade of CTLA4 with an anti-CTLA4 antibody has been shown to precipitate rejection and prevent induction of allograft tolerance in the transplant setting,30 reinforcing the important role of CTLA4 signaling in inhibiting the alloimmune response. CTLA4 is constitutively expressed on Tregs and has a 20-fold higher affinity than CD28 for both B7.1 and B7.2 ligands.31 In addition to the direct effect of Tregs on effector T cells by the secretion of inhibitory cytokines (e.g., IL-10) and apoptotic effects through granzyme B, the function of Tregs was also shown to be dependent on CTLA4 expression32 (Figure 2b). CTLA4 on Tregs can interact with B7.1/B7.2 ligands on antigen-presenting cells and downregulate the expression of these ligands while upregulating the expression of indoleamine 2,3-dioxygenase, a potent inhibitory molecule.33 An agonistic agent to CTLA4 could potentially promote tolerance and improve graft survival; however, attempts of developing this agent have so far been unsuccessful. Overall, CTLA4 is an important inhibitory signaling pathway, and its blockade by CTLA4-Ig could affect the regulation of the alloimmune response.

ecule.33 An agonistic agent to CTLA4 could potentially promote tolerance and improve graft survival; however, attempts of developing this agent have so far been unsuccessful. Overall, CTLA4 is an important inhibitory signaling pathway, and its blockade by CTLA4-Ig could affect the regulation of the alloimmune response. Th17 cells After antigen encounter, naïve T helper (Th) cells might differentiate into different subtypes according to signals delivered by antigen-presenting cells and the cytokines present in the microenvironment.34 The subtypes of Th cells are characterized by diverse cytokine productions, with Th1 cells producing predominantly interferon-γ, whereas Th2 cells secreting IL-4. Donor-specific T cell responses after transplantation are typically dominated by interferon-γ-producing T cells (Th1).35 More recently, T cells that produce IL-17 were discovered and showed an association with allograft rejection.36 The concern about Th17 cells is that they have been reported to be resistant to current available immunosuppression, and especially resistant to co-stimulation blockade.37, 38 In fact, CD28 co-stimulation reduced the frequency of Th17 cells, whereas CTLA4-Ig facilitated both murine and human Th17 differentiation in vitro.39 In addition, CTLA4:B7 interaction was also demonstrated to inhibit Th17 cell differentiation and suppress the development of Th17-mediated autoimmunity.40 Collectively, these findings suggest that B7:CD28 blockade might favor Th17 cell differentiation with potential concern for allograft outcome; however, the true role of IL-17 in the alloimmune response in humans still needs to be clarified.

l differentiation and suppress the development of Th17-mediated autoimmunity.40 Collectively, these findings suggest that B7:CD28 blockade might favor Th17 cell differentiation with potential concern for allograft outcome; however, the true role of IL-17 in the alloimmune response in humans still needs to be clarified. Memory T-cell resistance Memory T cells are lymphocytes that have been previously activated and possess a unique capacity to generate rapid effector functions upon rechallenge with antigen. This capacity is related to their lower threshold for activation, less dependence on co-stimulation, and enhanced trafficking/adhesion mechanisms, being especially important in the response to infectious organisms.41 Humans develop alloreactive memory T cells after exposure to blood transfusions, pregnancies, or prior transplantation. More recently, it has been proposed that alloreactive memory T cells can also be generated by exposure to pathogens and environmental antigens, because of the resemblance of allogeneic MHC and microbial Ag/self-MHC complex (cross-reactive response).42, 43 Finally, T-cell-depleting induction therapies used in transplantation have been shown to promote homeostatic proliferation of non-depleted T cells and these proliferating cells carry a memory phenotype.44, 45

because of the resemblance of allogeneic MHC and microbial Ag/self-MHC complex (cross-reactive response).42, 43 Finally, T-cell-depleting induction therapies used in transplantation have been shown to promote homeostatic proliferation of non-depleted T cells and these proliferating cells carry a memory phenotype.44, 45 The presence of memory T cells has important clinical relevance, as higher frequency of alloreactive T cells before transplantation correlate with an increased risk of rejection.46, 47 Furthermore, these memory T cells are more resistant to B7:CD28 co-stimulation blockade48 and, consequently, targeting solely this pathway might be ineffective in inducing tolerance. Indeed, the previous observations of effective results of CTLA4-Ig in naïve recipients with naïve T-cell repertoire in a laboratory-controlled environment (rodents) and failure of the same agent in promoting tolerance in recipients with memory cells (non-human primates) suggest a key role of these cells in tolerance resistance to co-stimulation blockade. Confirming this, the combination of CTLA4-Ig with a selective memory T cell agent (CD2-specific fusion protein alefacept) was shown to improve allograft survival in non-human primates,49 opening potential new avenues in targeting memory cells. Nevertheless, targeting these cells carry its own risks as they have a key role in immunity against infectious diseases.

with a selective memory T cell agent (CD2-specific fusion protein alefacept) was shown to improve allograft survival in non-human primates,49 opening potential new avenues in targeting memory cells. Nevertheless, targeting these cells carry its own risks as they have a key role in immunity against infectious diseases. FUTURE OF TARGETING CO-STIMULATORY PATHWAYS IN THE CLINIC Co-stimulation has a central role in T-cell activation, and targeting this pathway has become a reality in transplantation, consisting of a true translational research. However, the complex interplay between different co-stimulatory pathways and the function of these pathways in different cell types raised a number of challenges, and it is now clear that targeting a single pathway will likely be ineffective for the induction of transplantation tolerance. To improve long-term graft survival, avoiding calcineurin inhibitor nephrotoxicity and long-term cardiovascular and metabolic side-effects are important goals. A recent open-label phase II trial has demonstrated that switching from a calcineurin inhibitor-based therapy to a belatacept-based regimen at 6 months after transplant is feasible and well tolerated, demonstrating potential improvements in renal function at 12 months.50 This latter switch could be beneficial as it will require a less-intensive dose of belatacept in face of the smaller pool of alloreactive T cells later after transplant, and might be associated with a lower rejection rate.

asible and well tolerated, demonstrating potential improvements in renal function at 12 months.50 This latter switch could be beneficial as it will require a less-intensive dose of belatacept in face of the smaller pool of alloreactive T cells later after transplant, and might be associated with a lower rejection rate. Another experimental approach with remarkable results is the combination of CTLA4-Ig with a T-cell-depleting agent such as thymoglobulin. In a stringent transplant model in rodents, this combination tipped the balance of Tregs/Teff in favor of Tregs, promoting regulation and favoring graft survival.51 Moreover, the development of newer synergistic agents targeting negative co-stimulatory pathways such as PD-1:PD-L1 could enhance immune regulation and promote tolerance.52 Finally, the role of B cells in chronic rejection has been increasingly recognized,53 and the generation of selective agents that are capable of decreasing alloantibody production and generating regulatory B cells will likely lead to considerable improvements in graft outcomes. Overall, the future of co-stimulation targeting in kidney transplantation will most likely involve the combination of agents with different mechanisms of action, with the goal of inhibiting pathogenic lymphocytes and promoting regulatory ones, limiting single-drug toxicity, and possibly achieving the Holy Grail of transplant tolerance. This study was supported by a research grant from the American Society of Transplantation to LVR and by National Institutes of Health (NIH) grants RO1 AI51559 and PO1AI56299 to MHS.

Overall, the future of co-stimulation targeting in kidney transplantation will most likely involve the combination of agents with different mechanisms of action, with the goal of inhibiting pathogenic lymphocytes and promoting regulatory ones, limiting single-drug toxicity, and possibly achieving the Holy Grail of transplant tolerance. This study was supported by a research grant from the American Society of Transplantation to LVR and by National Institutes of Health (NIH) grants RO1 AI51559 and PO1AI56299 to MHS. TO CITE THIS ARTICLE: Riella LV and Sayegh MH. T-cell co-stimulatory blockade in kidney transplantation: back to the bench. Kidney Int Sup 2011; 1: 25–30. All the authors declared no competing interests. Figure 1 Role of co-stimulation in T-cell activation. (a) Upon antigen encounter, B7.1/B7.2 ligands expressed on antigen-presenting cells interact with CD28 receptors on T cells, leading to full T-cell activation. (b) In the absence of B7:CD28 interaction, T cells become anergic and/or apoptotic in the context of T-cell receptor (TCR) stimulation. This can be seen, for example, with blockade of the B7 ligands by CTLA4-Ig. (c) Co-stimulatory pathways can also be inhibitory as in the case of B7:CTLA4, which is able suppress T-cell activation. It is the balance between positive and negative co-stimulatory pathways that will ultimately determine T-cell outcome. APC, antigen-presenting cells; CTLA4, cytotoxic T-lymphocyte antigen-4.

TLA4-Ig. (c) Co-stimulatory pathways can also be inhibitory as in the case of B7:CTLA4, which is able suppress T-cell activation. It is the balance between positive and negative co-stimulatory pathways that will ultimately determine T-cell outcome. APC, antigen-presenting cells; CTLA4, cytotoxic T-lymphocyte antigen-4. Figure 2 Regulatory T cells and co-stimulatory pathways. (a) B7:CD28 pathway was demonstrated to have a major role in the generation and maintenance of regulatory T cells (Tregs), leading to some concerns on the effect of long-term B7:CD28 blockade on Tregs in transplantation. (b) Another concern arose from the discovery that Tregs use the inhibitory pathway B7:CTLA4 to suppress dendritic cell (DC) function via induction of indoleamine 2,3-dioxygenase (IDO) and inhibition of the maturation of DCs (not shown). However, Tregs also exert their suppressive function through other mechanisms, such as secretion of inhibitory cytokines (e.g., interleukin (IL)-10, transforming growth factor (TGF)-β) and granzyme B (GrB), possibly representing parallel pathways of immune regulation. APC, antigen-presenting cells; CTLA4, cytotoxic T-lymphocyte antigen-4; Teff, effector T cell.

function through other mechanisms, such as secretion of inhibitory cytokines (e.g., interleukin (IL)-10, transforming growth factor (TGF)-β) and granzyme B (GrB), possibly representing parallel pathways of immune regulation. APC, antigen-presenting cells; CTLA4, cytotoxic T-lymphocyte antigen-4; Teff, effector T cell. Figure 3 The pool size of alloreactive T cells and the effect of co-stimulation blockade. (a) In the setting of a large pool size of alloreactive T cells, blockade of B7:CD28 pathway by CTLA4-Ig leads to a predominant effect on effector T cells, tipping the balance toward regulatory T cells (Tregs)/tolerance. (b) When the number of alloreactive T cells is smaller, blockade of B7:CD28 leads to a dominant inhibitory effect on the generation of regulatory T cell, leading to more effector T cells than Tregs, and precipitating rejection in the transplant setting; CTLA4, cytotoxic T-lymphocyte antigen-4.

T cells are under continuous education to be able to discern between self- and foreign antigens. After leaving the thymus, naïve T cells can differentiate into many different subsets, depending on the environment milieu in which they are activated.1 Upon activation by professional antigen-presenting cells, naïve T cells can differentiate into activated T cells, driving primary responses against the antigen. After clearance of the antigen, most of the T cells undergo apoptosis, whereas the remaining T cells will be recruited into the memory T-cell pool. Once the antigen re-enters into the host, those memory immune T cells are able to mount a more efficient and faster response against it. In the transplantation setting, the memory cells can be generated by several ways after previous encounters with human leukocyte antigen molecules in sensitized patients (after pregnancy, blood transfusions), and in non-sensitized patients by cross-reactivity (molecular mimicry and bystander proliferation),2, 3 and homeostatic proliferation after lymphopenia.4

cells can be generated by several ways after previous encounters with human leukocyte antigen molecules in sensitized patients (after pregnancy, blood transfusions), and in non-sensitized patients by cross-reactivity (molecular mimicry and bystander proliferation),2, 3 and homeostatic proliferation after lymphopenia.4 In mice and humans, several populations of memory T cells have been described with different effector functions, tissue localizations, and phenotypic characteristics.5 The central memory T cells (TCM) are confined to lymphoid tissues, and express CD62L and CD45RO. They have a silent phenotype and therefore could fit in a tolerance profile; however, after reactivation, they could develop into effector memory T cells (TEM), losing partially CD62L expression with very strong effector properties and the capacity to drive an effector response even without the requirement for co-stimulatory molecules. There are different markers that help to define these subpopulations: CD27, CD28, CD95, CD127, and chemokine receptor-5 and chemokine receptor-7, reviewed elsewhere.5 One important caveat is that most of the transplant animal models that assess mechanisms of tolerance and alloimmune responses are under pathogen-free conditions in which the memory arm could be impaired or absent. In spite of this argument, there are a number of models in which the establishment of tolerance is not achieved in the presence of memory T cells,6 although treatments blocking co-stimulatory pathways were able to prolong graft survival.7

der pathogen-free conditions in which the memory arm could be impaired or absent. In spite of this argument, there are a number of models in which the establishment of tolerance is not achieved in the presence of memory T cells,6 although treatments blocking co-stimulatory pathways were able to prolong graft survival.7 The involvement of memory T cells in alloimmune responses is widely accepted and could have a role in the development of a rejection episode and, more specifically, in human renal transplantation. Nevertheless, such an issue has been poorly investigated. The present study addressed the changes in the number of circulating TCM and TEM in renal transplant recipients undergoing an acute rejection (AR) episode, as well as the effect that different immunosuppressant drugs have on them. RESULTS AND DISCUSSION CD4+ TEM cells and their association with acute rejection Within the increasing number of potential subsets involved in the development of allograft rejection, the role of memory T cells in AR pathogenesis was demonstrated as early as in the 70s in murine models.8 These cells are able to drive an AR episode earlier and faster than naïve T cells.9 After 20 years, two new subsets of memory T cells with different properties were described (TCM and TEM).10 Although both populations of CD8+ TEM and TCM were able to mount an effective alloimmune response against the graft in a skin graft model, the CD8+ TEM rejected allografts better than TCM in the absence of secondary lymphoid tissues,11 pointing to the TEM subset as a main candidate to drive secondary alloresponses.

.10 Although both populations of CD8+ TEM and TCM were able to mount an effective alloimmune response against the graft in a skin graft model, the CD8+ TEM rejected allografts better than TCM in the absence of secondary lymphoid tissues,11 pointing to the TEM subset as a main candidate to drive secondary alloresponses. In human heart and renal transplantation, the degree of AR was correlated with the degree of memory CD8+ T-cell infiltration.12, 13 However, in liver transplantation, an increased infiltration of naïve T cells was found.14 Further evidence of the role of memory T-cell activation in AR was demonstrated by the correlation of interferon-γ production measured by enzyme-linked immunosorbent spot with AR and poor graft outcome.15, 16

infiltration.12, 13 However, in liver transplantation, an increased infiltration of naïve T cells was found.14 Further evidence of the role of memory T-cell activation in AR was demonstrated by the correlation of interferon-γ production measured by enzyme-linked immunosorbent spot with AR and poor graft outcome.15, 16 During the last decades, several groups have investigated AR biomarkers, although because of good short-term results with the use of new therapeutic strategies, the interest in those biomarkers decreased. Despite this, 5–10% of kidney transplant recipients still suffer from an AR episode. Within the candidate biomarkers, memory T subsets have gained interest within the transplant research community. In liver transplantation, a recent study showed a shift from naïve CD8+ cells to the different memory subsets before AR episodes.17 In human heart transplantation, an increase in naïve and a decrease in TCM CD4+ cells before heart transplantation in patients developing AR has been demonstrated.18 Our group has studied the different subsets of memory T cells in 21 renal transplant patients during the first 2 months after transplantation. Three out of the 21 patients developed an AR episode and showed a decrease in the percentage of CD4+ TEM and CD8+ TEM during the first 2 months after transplantation (Figure 1a), although not significantly, probably because of the less number of patients. We were able to measure the number of circulating T-cell subsets in an independent group of eight patients with deterioration of renal function at the time of biopsy because of suspicion of AR. In such a moment, the frequencies of peripheral blood CD4+ TEM cells were increased, with a trend toward decrease, or remain at the same level at 1 month after biopsy (Figure 1b). Circulating CD8+ TEM cells did not change at biopsy and at 1 month later (not shown). These findings point to a possible role of CD4+ TEM immediately after transplantation as inducers of the immune response responsible for the AR and as AR biomarkers.

se, or remain at the same level at 1 month after biopsy (Figure 1b). Circulating CD8+ TEM cells did not change at biopsy and at 1 month later (not shown). These findings point to a possible role of CD4+ TEM immediately after transplantation as inducers of the immune response responsible for the AR and as AR biomarkers. Induction therapy with thymoglobulin, but not with basiliximab, induces increased levels of circulating CD4+ TEM cells One of the major limitations to the long-term success of transplantation is the need for immunosuppression during lifetime, and its deleterious effect on the graft and the recipient. Understanding the impact of different immunosuppressive regimens on memory T cells would be a very valuable tool in order to direct the therapy against the harmful subsets involved in transplantation. In an interesting in vitro model, after sorting CD8+ TEM cells cocultured with different immunosuppressant medications currently used in clinic, only the calcineurin inhibitor treatment was able to suppress their function, as compared with mammalian target of rapamycin inhibitors, steroids, or mycophenolic acid.19 This in vitro finding may have important consequences regarding immunosuppressant therapy in early post-transplant stages, as the TEM subset may have a role in driving an AR episode.

eatment was able to suppress their function, as compared with mammalian target of rapamycin inhibitors, steroids, or mycophenolic acid.19 This in vitro finding may have important consequences regarding immunosuppressant therapy in early post-transplant stages, as the TEM subset may have a role in driving an AR episode. In transplant models, several studies have recently been published using pre-sensitized animals in which the inhibition of the memory arm after co-stimulation blockade induction with anti-CD134L (OX-40L), anti-CD122 (beta-chain of interleukin-2 receptor), anti-CD154, and anti-LFA-1 was effective to prolong heart graft survival but unable to achieve tolerance.7

recently been published using pre-sensitized animals in which the inhibition of the memory arm after co-stimulation blockade induction with anti-CD134L (OX-40L), anti-CD122 (beta-chain of interleukin-2 receptor), anti-CD154, and anti-LFA-1 was effective to prolong heart graft survival but unable to achieve tolerance.7 With regard to induction therapy regimens, a fast recovery of blood TEM cells was observed after thymoglobulin treatment, whereas TCM levels were restored only after 3 months post-treatment.20 In renal transplant patients, Campath-1H induction evokes a severe lymphopenia, in which TCM cells are more resistant to depletion, and progressive restoration of TEM is found with time. Even one rejection episode was associated with a higher proportion of circulating CD4+ TEM cells.21 In our cohort population, we observed an increased proportion of CD4+ TEM cells at 6 months after transplantation in those patients who received thymoglobulin induction, whereas no differences were observed in patients without induction or in those on basiliximab treatment (Figure 2). No significant differences in CD8+ TEM cell frequencies were observed at 6 months after transplantation in patients undergoing thymoglobulin (30.7±15.0) or basiliximab induction (29.7±14), as compared with those patients who did not received induction therapy (33.5±12.0). Such a finding could be related to the phenomenon of homeostatic proliferation after the lymphopenia induced by thymoglobulin.4 However, basiliximab did not induce lymphopenia after induction treatment.

basiliximab induction (29.7±14), as compared with those patients who did not received induction therapy (33.5±12.0). Such a finding could be related to the phenomenon of homeostatic proliferation after the lymphopenia induced by thymoglobulin.4 However, basiliximab did not induce lymphopenia after induction treatment. Kinetics of peripheral blood memory T cells in the long-term follow-up Although memory T cells may be mainly involved in AR, with new induction therapies and depleting treatments, these populations could have an important role in long-term graft loss. There is evidence showing infiltration of memory populations in biopsies of renal transplant patients diagnosed with chronic allograft nephropathy.22 This is indirectly in agreement with previous data from our group showing decreased number of TCM CD3+ cells in patients with long-term failed grafts and reintroduced in the waiting list.23 In the present study, we observed that even several months after diagnosis, those patients suffering with an AR episode maintained increased percentages of CD4+ TEM and CD8+ TEM cells (Figure 3), although it did not affect graft survival. This is in contrast with the apparent decrease of both TEM subsets in the short-term follow-up of those patients suffering AR (Figure 1a and b). Nevertheless, the possible effect of memory T cells on long-term prognosis and chronic rejection in renal transplantation still awaits investigation.

ot affect graft survival. This is in contrast with the apparent decrease of both TEM subsets in the short-term follow-up of those patients suffering AR (Figure 1a and b). Nevertheless, the possible effect of memory T cells on long-term prognosis and chronic rejection in renal transplantation still awaits investigation. CONCLUDING REMARKS Despite the well-accepted role of memory T cells as initiators of the potent secondary immune response, they have not been particularly addressed as clinical biomarkers in the setting of renal transplantation. In the present study, we evaluated the quantitative changes in the two main memory T-cell subsets, TCM and TEM, in the peripheral blood of renal transplant recipients. Our data point to a possible role of CD4+ TEM immediately after transplantation as inducers of the immune response that would finally activate the effector CD8+ TEM responsible for AR. There is practically no data regarding the association between memory T cells and chronic rejection in human renal transplantation. The different immunosuppresive regimens seem to have no quantitative effect on memory T cells. However, induction therapy with thymoglobulin, but not with basiliximab, induced increased numbers of circulating CD4+ TEM cells.

ng the association between memory T cells and chronic rejection in human renal transplantation. The different immunosuppresive regimens seem to have no quantitative effect on memory T cells. However, induction therapy with thymoglobulin, but not with basiliximab, induced increased numbers of circulating CD4+ TEM cells. PATIENTS AND METHODS Patients A cohort of 90 patients who underwent consecutive renal transplant in the Hospital Universitario Marqués de Valdecilla from September 2007 was recruited, and informed consent given for participation in this study. All the clinical data were recorded at the Nephrology Department. The patients were divided into two groups depending on the time of follow-up: the first group consisted of 69 patients with long-term follow-up (6 months, 1 and 2 years after transplantation), whereas the second one consisted of 21 patients with a short-term follow-up during 2 months. Within long-term follow-up group, 15 patients suffered an AR episode (21.7%), whereas within the short-term follow-up group 14.3% had an AR episode. Immunological, demographic, and clinical parameters are summarized in Table 1. No significant differences in these parameters between the non-AR and the AR group were observed in any of the cohorts studied.

5 patients suffered an AR episode (21.7%), whereas within the short-term follow-up group 14.3% had an AR episode. Immunological, demographic, and clinical parameters are summarized in Table 1. No significant differences in these parameters between the non-AR and the AR group were observed in any of the cohorts studied. Phenotype analysis Four-color flow cytometry analyses were performed on peripheral whole blood collected in heparin anticoagulant tubes, within 2 h after collection. The blood cells were stained with the following monoclonal antibodies (BD Biosciences, San Jose, CA): anti-CD62L-fluorescein isothiocyanate, anti-CD45RO-phycoerythrin, anti-CD4-peridinin chlorophyll protein, anti-CD8-peridinin chlorophyll protein, and anti-CD3-allophycocyanine. TEM cells were defined by their phenotype CD45RO+CD62L+, whereas the TCM cells were defined by CD45RO+CD62 L−. Cells were incubated for 30 min in the dark at room temperature. Subsequently, the erythrocytes were lysed after 10-min incubation with FACS lysing solution (BD Biosciences) and washed with phosphate-buffered saline. After centrifugation, the cells were resuspended in 0.2 ml of phosphate-buffered saline before acquisition of 50 000 events in lymphocyte gate by FACScalibur flow cytometer. All the data were analyzed using the CellQuest Pro software (BD Biosciences).

solution (BD Biosciences) and washed with phosphate-buffered saline. After centrifugation, the cells were resuspended in 0.2 ml of phosphate-buffered saline before acquisition of 50 000 events in lymphocyte gate by FACScalibur flow cytometer. All the data were analyzed using the CellQuest Pro software (BD Biosciences). Statistical analysis The data from both long- and short-term groups were nonparametrically distributed (Kolmogorov–Smirnov test). Differences in the percentage of CD4+CD45RO+CD62 L− TEM cells between the group suffering an AR episode and those who were rejection-free were analyzed by using the Mann–Whitney U test. The clinical and demographical parameters were analyzed using Student's t-test and Mann–Whitney U test as appropriate. P-values <0.05 were considered significant. This work was partially supported by grants from the Fondo de Investigaciones Sanitarias-ISCIII (PI070683, PI080157, REDINREN 06/16 ISCIII) and from the Fundación Mutua-Madrileña (ACI 07/01). DSS is a recipient of a Lopez-Albo grant from the Fundación Marqués de Valdecilla-IFIMAV. IB and MG are technicians supported by grants from the Fundación Marqués de Valdecilla-IFIMAV. TO CITE THIS ARTICLE: Segundo DS, Fernández-Fresnedo G, Gago M et al. Changes in the number of circulating TCM and TEM subsets in renal transplantation: relationship with acute rejection and induction therapy. Kidney Int Sup 2011; 1: 31–35. All the authors declared no competing interests.

This work was partially supported by grants from the Fondo de Investigaciones Sanitarias-ISCIII (PI070683, PI080157, REDINREN 06/16 ISCIII) and from the Fundación Mutua-Madrileña (ACI 07/01). DSS is a recipient of a Lopez-Albo grant from the Fundación Marqués de Valdecilla-IFIMAV. IB and MG are technicians supported by grants from the Fundación Marqués de Valdecilla-IFIMAV. TO CITE THIS ARTICLE: Segundo DS, Fernández-Fresnedo G, Gago M et al. Changes in the number of circulating TCM and TEM subsets in renal transplantation: relationship with acute rejection and induction therapy. Kidney Int Sup 2011; 1: 31–35. All the authors declared no competing interests. Figure 1 Changes in the frequencies of circulating TEM cells in early post-transplantation. (a) Frequencies of CD4+ TEM (left) and CD8+ TEM (right) cells in peripheral blood of renal transplant patients during the first 2 months after transplantation. Patients were grouped according to the development of acute rejection (A) or not (N). Each dot represents one patient and horizontal bars represent the mean value in each group. No significant differences were observed. (b) Changes in the frequencies of peripheral blood CD4+ TEM cells in a group of eight patients at the time of diagnosis of acute rejection by biopsy and 1 month after biopsy.

or not (N). Each dot represents one patient and horizontal bars represent the mean value in each group. No significant differences were observed. (b) Changes in the frequencies of peripheral blood CD4+ TEM cells in a group of eight patients at the time of diagnosis of acute rejection by biopsy and 1 month after biopsy. Figure 2 Frequencies of CD4+ TEM cells in peripheral blood of long-term follow-up renal transplant recipients at 6 months after transplantation according to the induction therapy received. Significant increased frequencies were observed in the thymoglobulin (TG)-treated patients as compared with basiliximab (anti-CD25)-treated patients or those not receiving induction therapy. Each dot represents one patient and horizontal bars represent the mean value in each group. Empty dots represent patients who did not receive induction therapy. Filled dots represent patients who received induction therapy with anti-CD25. Triangles represent patients who received induction therapy with TG.

ion therapy. Each dot represents one patient and horizontal bars represent the mean value in each group. Empty dots represent patients who did not receive induction therapy. Filled dots represent patients who received induction therapy with anti-CD25. Triangles represent patients who received induction therapy with TG. Figure 3 Changes in the frequencies of circulating TEM cells in late post-transplantation. Frequencies of CD4+ TEM (left) and CD8+ TEM (right) cells in peripheral blood of renal transplant patients during the first 2 years after transplantation. Patients were grouped according to the development of acute rejection (A) or not (N). Each dot represents one patient and horizontal bars represent the mean value in each group. Data were collected at 6 months, 1 year, and 2 years after transplantation. P-value is indicated when statistically significant difference was found. Triangles in the left figure represent those patients who did not suffer acute rejection (N) whereas empty dots represent patients who developed acute rejection (A). In the right figure, filled dots represent patients who did not develop acute rejection (N) whereas empty dots represent patients who developed acute rejection (A). ESRD, end-stage renal disease.

ose patients who did not suffer acute rejection (N) whereas empty dots represent patients who developed acute rejection (A). In the right figure, filled dots represent patients who did not develop acute rejection (N) whereas empty dots represent patients who developed acute rejection (A). ESRD, end-stage renal disease. Table 1 Demographic, immunological, and clinical data from short-term and long-term follow-up renal transplant patients Short-term follow-up Long-term follow-up N 21 69 Donor age (years, mean±s.d.) 58±17.3 50.4±15.7 Recipient age (years, mean±s.d.) 63.3±7.2 52.8±12 Time on dialysis (days, mean±s.d.) 287±271 25.9±18.6 Number of transplants (1/2/3) (21/0/0) (49/15/5) Acute rejection (yes/no, %) 3/18 (14.3) 15/54 (21.7) Banff classification (BL/Ia/Ib/IIa/IIb/H) 2/0/0/1/0/0 3/5/1/3/1/2 Kidney disease (% of GN) 33 36 Mismatches (A/B/DR, mean) 1.1/1.0/1.0 0.91/1.27/1.0 Induction therapy (no/thymoglobulin/basiliximab) Not determined 46/6/17 Serum creatinine (mg/dl, mean±s.d.)a 2.27±0.67 6 months: 1.52±0.44 1 year: 1.47±0.38 2 years: 1.51±0.46 MDRD (ml/min per 1.73 m2)a 37.1±25.6 6 months: 51±14.8 1 year: 52±12.5 2 years: 51.3±14.9 Abbreviations: BL, borderline; GN, glomerulonephritis; MDRD, modification of diet in renal disease; s.d., standard deviation. a Data in the short-term follow-up group are at 2 months after transplantation. In the long-term follow-up, data are indicated at (6 months, 1 year, 2 years).

Protocol biopsy studies have contributed to the understanding of the natural history of renal allograft lesions. These studies have shown that histological damage precedes the appearance of proteinuria or renal functional deterioration in different conditions such as subclinical rejection (SCR), interstitial fibrosis/tubular atrophy (IF/TA), chronic humoral rejection, recurrence of the primary disease, de novo glomerulonephritis, or polyoma virus infection. Protocol biopsy studies have shown that early histological lesions constitute an independent predictor of graft outcome. Of all these conditions, SCR has captured the interest of the transplant community for a long period of time.1 SUBCLINICAL REJECTION AND GRAFT OUTCOME The term SCR was coined in a study of serial protocol biopsies in 1995 when immunosuppressive treatment consisted of cyclosporine, azathioprine, and prednisone.2 In this pioneering study, the prevalence of SCR at different time points during the first year was over 50%. Accordingly, different studies evaluated whether SCR was associated with outcome. In serial protocol biopsy studies, the presence of SCR in a first biopsy was associated with progression of IF/TA, impairment of glomerular adaptation, and progression of glomerulosclerosis in the second one,3, 4 suggesting that early inflammation favors progression of chronic lesions.

SCR was associated with outcome. In serial protocol biopsy studies, the presence of SCR in a first biopsy was associated with progression of IF/TA, impairment of glomerular adaptation, and progression of glomerulosclerosis in the second one,3, 4 suggesting that early inflammation favors progression of chronic lesions. During the 90s, an association between the presence of IF/TA and graft survival was consistently described in protocol biopsy studies. Similarly, in 2005, an association between SCR in 2-week protocol biopsies and graft survival was also documented.5 More recently, it has been further described that the association of inflammation and IF/TA in protocol biopsies implies a poorer outcome than IF/TA or SCR alone.6, 7, 8 Altogether, these observations suggest an association between SCR and poor graft outcome, and raise the question whether treatment of SCR may improve outcome.

ently, it has been further described that the association of inflammation and IF/TA in protocol biopsies implies a poorer outcome than IF/TA or SCR alone.6, 7, 8 Altogether, these observations suggest an association between SCR and poor graft outcome, and raise the question whether treatment of SCR may improve outcome. PREVALENCE OF SUBCLINICAL REJECTION AND IMMUNOSUPPRESSION The potential benefit of SCR treatment was explored in a randomized trial in patients receiving cyclosporine, azathioprine, and prednisone. In the study group, patients were biopsied at 1, 2, and 3 months and treated with steroid boluses, whereas the control group was not biopsied and accordingly not treated. IF/TA at 6 months and serum creatinine at 2 years were lower in the treatment group, suggesting that treatment of SCR may improve graft outcome.9 However, introduction of tacrolimus and mycophenolate reduced the prevalence of SCR to ∼10%,10, 11 showing that SCR can be prevented with more efficient immunosuppression. In a study of serial protocol biopsies, it was observed that reduction of SCR with tacrolimus and mycophenolate was also associated with a lower prevalence of IF/TA at 1 year, in comparison with cyclosporine-based regimens.3 Moreover, in a randomized study comparing a calcineurin-based regimen, either associated with mycophenolate mofetil or associated with sirolimus, the sirolimus groups displayed a lower prevalence of acute rejection and SCR during the first year and a lower prevalence of IF/TA at 5 years.12 Altogether, these data suggest that SCR prevention with more efficient immunosuppression may improve graft outcome.

ed with mycophenolate mofetil or associated with sirolimus, the sirolimus groups displayed a lower prevalence of acute rejection and SCR during the first year and a lower prevalence of IF/TA at 5 years.12 Altogether, these data suggest that SCR prevention with more efficient immunosuppression may improve graft outcome. RISK FACTORS FOR SUBCLINICAL REJECTION The close association between immunosuppressive treatment and prevalence of SCR has favored the assumption that it represents, at the histological level, the balance between alloimmune response and efficiency of immunosuppressive treatment. From the epidemiological point of view, it has been described that the degree of sensitization and clinical episodes of acute rejection preceding the protocol biopsy constitute a risk factor for SCR, further reinforcing the notion that SCR represents the intensity of the immune response.1 However, non-immune factors are also associated with SCR, such as donor age.5 Kidneys from older donors are more susceptible to insult and have reduced capacity for tissue regeneration.13 Thus, it is tempting to speculate that SCR may not only reflect alloimmune response but also may reflect the inflammatory response associated with tissue injury and repair.

ociated with SCR, such as donor age.5 Kidneys from older donors are more susceptible to insult and have reduced capacity for tissue regeneration.13 Thus, it is tempting to speculate that SCR may not only reflect alloimmune response but also may reflect the inflammatory response associated with tissue injury and repair. INNATE IMMUNE ALTERATIONS, TISSUE DAMAGE, AND TISSUE REPAIR Innate immunity constitutes the first line of defense against infection and has a major role in tissue repair.14 Few highly conserved structures on microorganisms, that is, pathogen-associated molecular patterns, are recognized by pattern recognition receptors that are expressed on effector cells of the innate immune system, such as macrophages, dendritic cells, or B cells. This pattern recognition receptors can be divided into three types: endocytic, signaling, and secreted. Probably, the best-known secreted pattern recognition receptors molecule is mannose-binding lectin (MBL) that activates complement by the lectin pathway and favors inflammation and phagocytosis at the site of infection. Cellular necrosis or apoptosis that follows tissue injury is characterized by the secretion of danger signals known as alarmins, which are sensed by the innate immune system. Recognition of alarmins triggers inflammation and favors phagocytosis of necrotic and apoptotic cells. Once necrotic and apoptotic cells have been cleared, inflammation fades and tissue regeneration leads to injury healing.15, 16

cretion of danger signals known as alarmins, which are sensed by the innate immune system. Recognition of alarmins triggers inflammation and favors phagocytosis of necrotic and apoptotic cells. Once necrotic and apoptotic cells have been cleared, inflammation fades and tissue regeneration leads to injury healing.15, 16 As the response to infection and tissue damage is similar, pathogen-associated molecular patterns and alarmins are also termed as danger-associated molecular patterns. Alterations of the innate immune response are associated with inefficient tissue healing, chronic inflammation, and autoimmune disease due to exposure of intracellular antigens and inefficient necrotic and apoptotic cell phagocytosis. All these alterations may also contribute to renal damage after transplantation.

rns. Alterations of the innate immune response are associated with inefficient tissue healing, chronic inflammation, and autoimmune disease due to exposure of intracellular antigens and inefficient necrotic and apoptotic cell phagocytosis. All these alterations may also contribute to renal damage after transplantation. MBL LEVELS AND DISEASE IN GENERAL POPULATION Innate immune alterations are associated with the prevalence and outcome of different diseases. Different polymorphisms of the MBL gene have been described and all of them are associated with decreased MBL levels and impaired MBL function. However, the relationship between MBL and disease is rather complex. Whereas low serum MBL levels are associated with an increased prevalence of infection, autoimmunity, diabetes, and cardiovascular disease, high serum MBL levels are associated with a poorer outcome of certain autoimmune diseases.17 An example is that low serum MBL level is associated with higher risk for diabetes,18 whereas in patients with diabetes, high serum MBL level is associated with more severe renal damage.19 Furthermore, in acute tissue injury after renal ischemia reperfusion injury, high serum MBL level is associated with more severe functional deterioration.20

rum MBL level is associated with higher risk for diabetes,18 whereas in patients with diabetes, high serum MBL level is associated with more severe renal damage.19 Furthermore, in acute tissue injury after renal ischemia reperfusion injury, high serum MBL level is associated with more severe functional deterioration.20 MBL LEVELS AND RENAL TRANSPLANT OUTCOME In renal transplantation, an association between MBL polymorphisms or low serum MBL levels and increased susceptibility to cytomegalovirus, bacterial, or fungal infections has been reported in some,21, 22, 23 but not in all, studies.24, 25 Despite the higher incidence of infection in low serum MBL patients, no association between MBL levels and mortality has been described.26, 27 On the contrary, an association between high MBL serum levels and decreased death-censored renal allograft survival has been observed. The incidence of acute rejection was not associated with serum MBL levels, but acute rejection was more often the cause for graft failure in patients with high serum MBL levels, suggesting that high MBL levels may be associated with severe forms of acute rejection.26 Similarly, in kidney–pancreas transplantation, allograft survival was lower in patients with high serum MBL levels.28 In contrast, in heart transplantation, low MBL levels were associated with an increased prevalence of acute rejection and transplant-associated coronary artery disease.29 Taking all these data together, it is difficult to interpret the apparent discrepancies between kidney and heart transplantation. A difference between kidney and heart transplants is that protocol biopsies are usually used to monitor heart histology and, only in few centers, to monitor renal allografts. Furthermore, transplant vasculopathy is actively monitored in the heart by means of coronariography or intravascular ultrasounds. Until now, no studies have analyzed the association between histological damage in protocol renal allograft biopsies and serum MBL levels.

, only in few centers, to monitor renal allografts. Furthermore, transplant vasculopathy is actively monitored in the heart by means of coronariography or intravascular ultrasounds. Until now, no studies have analyzed the association between histological damage in protocol renal allograft biopsies and serum MBL levels. In a recent study, we measured serum MBL levels after transplantation in a cohort of consecutive renal transplants and classified MBL as low or high according to tertile distribution. The first tertile was considered as the low MBL group and the two higher tertiles were considered as the high MBL group. Despite the fact that donor and recipient characteristics were similar between groups, and that the prevalence of delayed graft function or acute rejection was not different between groups, we observed that patients with low MBL levels displayed more severe low-grade inflammation before transplantation as measured by serum levels of soluble tumor necrosis factor receptor 2, suffered a higher incidence of bacterial or fungal infections, and showed an increased prevalence of new-onset diabetes after transplantation.23

that patients with low MBL levels displayed more severe low-grade inflammation before transplantation as measured by serum levels of soluble tumor necrosis factor receptor 2, suffered a higher incidence of bacterial or fungal infections, and showed an increased prevalence of new-onset diabetes after transplantation.23 In the above-mentioned study, a 3-month surveillance biopsy was done in patients with a stable serum creatinine <200 μmol/l and without significant proteinuria (<1 g/day) after obtaining informed consent. There were 60 out of 125 recruited patients with an adequate surveillance biopsy. Histological examination of these biopsies according to the Banff criteria led to the diagnosis of SCR in 7 of 18 (38.9%) low MBL patients and in 3 of 42 (7.1%) high MBL patients (P=0.0054). Induction and maintenance immunosuppression was not different in patients with low and high MBL levels (Table 1). This observation suggests that innate immune alterations such as MBL deficiency may contribute to the degree of inflammation of the renal allograft (Figure 1). Thus, it is tempting to speculate that patients with low MBL levels may have an impaired capacity for tissue repair leading to more severe and prolonged inflammation.

bservation suggests that innate immune alterations such as MBL deficiency may contribute to the degree of inflammation of the renal allograft (Figure 1). Thus, it is tempting to speculate that patients with low MBL levels may have an impaired capacity for tissue repair leading to more severe and prolonged inflammation. CONCLUSION Innate immune alterations modulate different comorbidities in renal transplant patients and may influence graft outcome. Despite this association, mechanisms linking innate immunity and graft damage have not been clearly elucidated. In one hand, it has been suggested that patients with high MBL levels may suffer from more severe episodes of rejection, whereas in other studies low MBL levels have been associated with a higher prevalence of subclinical or clinical rejection. Thus, the relationship between graft outcome and innate immune alterations deserves further studies. This study was supported by Fondo Investigaciones Sanitarias grants from the Spanish Minister of Health (PI04/0086, PI04/0177, and PI10/02496) and a Spanish Society of Nephrology grant. MI was supported by a Catalan Society of Transplantation grant. TO CITE THIS ARTICLE: Ibernon M, Moreso F, Serón D. Subclinical rejection in renal transplants is associated with low serum mannose-binding lectin levels. Kidney Int Sup 2011; 1: 36–39. All the authors declared no competing interests. Figure 1 Relationship between low serum MBL levels and clinical events after renal transplantation. MBL, mannose-binding lectin.

TO CITE THIS ARTICLE: Ibernon M, Moreso F, Serón D. Subclinical rejection in renal transplants is associated with low serum mannose-binding lectin levels. Kidney Int Sup 2011; 1: 36–39. All the authors declared no competing interests. Figure 1 Relationship between low serum MBL levels and clinical events after renal transplantation. MBL, mannose-binding lectin. Table 1 Induction and maintenance immunosuppression in patients with low or high MBL levels Low MBL High MBL P-value Induction None 1 4 Anti-IL2 antibodies 12 29 Thymoglobulin 5 9 ns Maintenance immunosuppression CNI+MMF+P 5 14 mTORi+MMF+P 8 12 CNI+mTORi+P 3 8 Belatacept+MMF+P 2 8 ns Abbreviations: Anti-IL2, anti-interleukin 2; CNI, calcineurin inhibitors; MBL, mannose-binding lectin; MMF, mycphenolate mofetil; mTORi, mammalian target of rapamycin inhibitors; ns, not significant; P, prednisone.

Solid organ transplantation is the preferred treatment for end-stage kidney failure, as it substantially improves patient survival and quality of life over continued dialysis therapy. However, in order to avoid rejection, graft recipients have to undergo lifelong immunosupression (IS). This poses a difficult challenge for patients and clinicians as a delicate equilibrium needs to be reached. On one hand, the transplant needs to be protected with sufficient IS, whereas on the other hand the patient needs to maintain an immune system that is healthy enough to fight-off infections and cancer. Moreover, immunosupressive agents have metabolic side effects (hypertension, hyperlipidemia, and hyperglycemia) that contribute to the high level of morbidity in patients post-transplantation. As a consequence, the 10-year overall survival rate of a transplanted kidney is just above 50% in Europe,1 which puts a yearly increased strain on the number of people waiting for further transplants. Causes of transplant failure are diverse, uncontrolled activity of the inflammatory, and immune systems being one of the main contributors.2, 3, 4

ar overall survival rate of a transplanted kidney is just above 50% in Europe,1 which puts a yearly increased strain on the number of people waiting for further transplants. Causes of transplant failure are diverse, uncontrolled activity of the inflammatory, and immune systems being one of the main contributors.2, 3, 4 Ideally, the best clinical situation for a transplant recipient would be the development of donor-specific immunological tolerance. This would allow long-term patient and graft survival without the need for IS. Thus, there is a pressing need to establish the correct level of IS in each patient, such as to individualize their therapy. In this process, the lack of analytical parameters able to indicate the correct treatment for each patient has prompted a wide search for biomarkers of clinical use in transplantation. In the early-phase post-transplantation, the ability to predict acute rejection would allow us to adjust IS accordingly, whereas in the late-phase post-transplantation, our inability to detect whether a patient has developed a degree of tolerance to their graft is absolute. This has led us to use the term ‘the hidden phenotype'. Currently, there are no clinically validated tools to dictate whether to increase or decrease the level of IS that each patient needs to be maintained with. In this review, we will discuss the progress made to date and the future avenues in the development of biomarkers of immunological tolerance in kidney transplantation.

ently, there are no clinically validated tools to dictate whether to increase or decrease the level of IS that each patient needs to be maintained with. In this review, we will discuss the progress made to date and the future avenues in the development of biomarkers of immunological tolerance in kidney transplantation. THE PAST: THE INDICES OF TOLERANCE STUDY Despite promising studies reporting the induction of tolerance in experimental models of solid organ transplantation, few of these approaches have been translated to clinical transplantation. The Indices of Tolerance (IoT) study was initiated as a necessary shift in the paradigm to approaches of inducing transplant tolerance. If we could identify the specific immunological characteristics of tolerance in human subjects, then we could use more targeted and informed approaches to tolerance-induction therapies, with the potential to manipulate the immune compartments responsible for generating donor-specific immune regulation. In 2010, we reported on the findings of the IoT multicenter study, which was conducted over a period of 6 years, and culminated in the description of several immunological characteristics uniquely associated with the tolerant state in renal transplant patients.5 As an investigator-led study, based within an academic institute, we were faced with the logistic challenges of conducting a clinical study from a primarily bench-based laboratory perspective. Overcoming this was a steep learning curve for the scientists and clinicians involved alike. In fact, the first challenge we faced on initiating the study was the recruitment of tolerant renal transplant subjects. As described later, the risk of renal allograft loss due to IS cessation, based on clinical experience, is thought to be high, and as a consequence, operational tolerance is estimated to be a rare event. Nevertheless, we were able to locate 11 tolerant renal transplant recipients throughout Europe. A number of pathways had led to this outcome, from medical non-compliance to withdrawal due to malignancies. We also encountered some exceptional cases, such as a renal allograft recipient who was unable to gain access to his IS for several weeks, because of severe flooding, which left him stranded on his roof. This latter example seems serendipitous, but may also defy the traditional dogma that the establishment of renal transplant tolerance is a rare occurrence and needs to occur over a period of time.

who was unable to gain access to his IS for several weeks, because of severe flooding, which left him stranded on his roof. This latter example seems serendipitous, but may also defy the traditional dogma that the establishment of renal transplant tolerance is a rare occurrence and needs to occur over a period of time. As discussed later, currently, this cannot be tested by any ethically sound approach. By comparing parameters of the immune system in the identified operationally tolerant recipients (stable renal transplant recipients who have ceased all immunosuppressive drugs for more than a year) with control groups (patients with stable renal function maintained on immunosuppressive drugs, patients with biopsy-proven chronic rejection, and healthy individuals), we identified a biomarker signature of tolerance. As indicated later, we adopted both cellular and molecular analytical approaches in our attempt to identify tolerant patients. One of the hypotheses tested was to find an absent functional or active anti-donor immune response. Analysis of cellular immune components revealed elevated numbers of B and NK cells in peripheral blood, diminished numbers of recently activated CD4+ T cells, and donor-specific hyporesponsiveness of CD4+ T cells. Further, we identified a discrete set of genes with altered expression, and a high ratio of FoxP3/α-1.2-mannosidase gene expression in peripheral blood. These findings were then validated on an independent cohort of tolerant renal transplant patients as part of a collaboration with the Immune Tolerance Network (ITN) in the USA, which subsequently led to a parallel publication showing highly comparable findings.6 The cross-platform approach we adopted toward biomarker identification highlighted a particularly prominent role for B cells within transplantation tolerance, which have very recently re-emerged with newly defined roles within inflammation and immunity.7, 8, 9

a parallel publication showing highly comparable findings.6 The cross-platform approach we adopted toward biomarker identification highlighted a particularly prominent role for B cells within transplantation tolerance, which have very recently re-emerged with newly defined roles within inflammation and immunity.7, 8, 9 THE PRESENT: THE VALIDATION OF THE BIOMARKERS OF TOLERANCE Although a set of biomarkers associated with tolerance had been defined at the completion of the IoT project, more questions were then raised: First, would newly identified tolerant patients show the same biomarkers? Second, would the biomarker fingerprint of tolerance also be detectable within groups of renal transplant patients displaying stable allograft function while undertaking immunosuppressive regimens? If so, could the fingerprint be used to predict which patients are more likely to develop tolerance and, as a consequence, would it be possible to successfully minimize their IS under supervision? Finally, could the fingerprint also define patients at the other spectrum of tolerance; that is, patients undergoing chronic rejection? In which case, a set of biomarkers could potentially be used as a monitoring tool for kidney function. To address these questions we initiated our current project: Genetic Analysis and Monitoring of Biomarkers of Immunological Tolerance.10

the other spectrum of tolerance; that is, patients undergoing chronic rejection? In which case, a set of biomarkers could potentially be used as a monitoring tool for kidney function. To address these questions we initiated our current project: Genetic Analysis and Monitoring of Biomarkers of Immunological Tolerance.10 We approached this project with the advantage of having learned many lessons from the IoT. We have put into place a network of several participating centers within the United Kingdom and Europe, together with a continuing firm collaboration with the ITN. We have recruited a technically skilled team that includes not only scientists and clinicians but also statisticians, data managers, and project administrators. We also now have the benefit of new developments in high-throughput techniques, which, because of the NIHR and the UK government investments in translational medical research, have allowed us to establish the infrastructure, facilities, and capacity to perform cutting-edge research. In addition, we are also participants of the International Solid Organ Transplant Registry, a renal transplant registry aiming to establish a multicenter database registering patients who underwent solid organ transplantation, in whom operational tolerance has developed.11 International Solid Organ Transplant Registry represents a meeting place for patients, families, and physicians, thus facilitating the enrolment of interested patients into appropriate studies of transplant tolerance.

ing patients who underwent solid organ transplantation, in whom operational tolerance has developed.11 International Solid Organ Transplant Registry represents a meeting place for patients, families, and physicians, thus facilitating the enrolment of interested patients into appropriate studies of transplant tolerance. GENETIC ANALYSIS AND MONITORING OF BIOMARKERS OF IMMUNOLOGICAL TOLERANCE The aim of the Genetic Analysis And Monitoring Of Biomarkers Of Immunological Tolerance study is essentially to assess the biomarkers of tolerance in a larger cohort of kidney transplant patients. This primary aim is approached from several angles.

GENETIC ANALYSIS AND MONITORING OF BIOMARKERS OF IMMUNOLOGICAL TOLERANCE The aim of the Genetic Analysis And Monitoring Of Biomarkers Of Immunological Tolerance study is essentially to assess the biomarkers of tolerance in a larger cohort of kidney transplant patients. This primary aim is approached from several angles. Tolerant patients We aim to analyze an independent set of tolerant renal transplant recipients in an observational longitudinal study. Working with samples from newly recruited operationally tolerant patients identified by the collaborating centers around Europe, we are performing the assays described in the IoT fingerprint5 to assess their probability of being tolerant. This will provide further important information on the validation of the previously identified biomarkers. In addition, because of the acquisition of new equipment and expertise within the research team, a selection of new flow cytometry parameters and a combination of new genes have also been included into the analyses. Finally, we are performing further screening of the biomarkers to be conducted over a period of 3 years in previously identified tolerant patients, testing the important aspect of the stability of the fingerprint. To keep assessing the specificity of the signature, we will also recruit healthy volunteers, patients under different degrees of IS because of a transplant, and patients taking immunsosuppression treatment because of other immunological causes apart from transplantation, such as autoimmune diseases.

of the fingerprint. To keep assessing the specificity of the signature, we will also recruit healthy volunteers, patients under different degrees of IS because of a transplant, and patients taking immunsosuppression treatment because of other immunological causes apart from transplantation, such as autoimmune diseases. Long-term stable kidney function This part of the study will compare the biomarker signature among patients with stable graft function under conventional IS with patients undergoing chronic rejection. We expect the signature to be present in a small percentage (5–10%, based on the IoT analyses) of patients with stable kidney function. Identifying patients under IS, who may be labeled ‘tolerant', is a crucial step toward translating these biomarkers into the clinic. IS withdrawal We will assess the frequency of patients showing the tolerance signature in an observational, prospective study of kidney transplant patients who are undergoing steroid withdrawal within 1 year post-transplantation. This protocol will also identify whether patients under IS withdrawal acquire the signature over time, allowing us to test the usefulness of the fingerprint as a biomarker for successful drug weaning.

ospective study of kidney transplant patients who are undergoing steroid withdrawal within 1 year post-transplantation. This protocol will also identify whether patients under IS withdrawal acquire the signature over time, allowing us to test the usefulness of the fingerprint as a biomarker for successful drug weaning. Basic research In parallel to these translational projects, we will investigate the role of B cells in tolerance in more detail as a follow-up of the B-cell function enrichment found in the genetic signature. Indeed, since the IoT and ITN studies, the role of B cell in transplantation tolerance has been further confirmed by others.12 The goal is to examine whether any unique phenotypic or functional characteristics can be identified within the subsets of these expanded cells during operational tolerance. An understanding of the mechanistic basis of the tolerant state would not only allow us to better identify it, but also permit the development of molecular or cellular targets for tolerance induction therapy in renal transplant patients.

dentified within the subsets of these expanded cells during operational tolerance. An understanding of the mechanistic basis of the tolerant state would not only allow us to better identify it, but also permit the development of molecular or cellular targets for tolerance induction therapy in renal transplant patients. OTHER MULTINATIONAL EFFORTS IN DETECTING TRANSPLANT TOLERANCE Detecting and/or inducing tolerance requires large multicenter networks that facilitate patient recruitment and ensure the necessary technical and scientific expertise. Several institutions are running projects with similar aims (Table 1). Immediately after the IoT study commenced, the RISET consortium was formed with similar aims: the development of reliable tests to predict tolerance, with a view to assess patients enrolled in RISET pilot clinical studies. Further, RISET aimed to stimulate debate regarding ethical aspects of tolerance induction protocols and establish educational programs in transplantation tolerance, not only for scientists and clinicians but also for patients and their families.18 The RISET consortium is currently compiling the final report to summarize their activities. The first study to emerge in the Framework 7th research program from the EU commission is THE ONE study. This study led by Dr Geissler in Regensburg is going to test a number of cellular therapies to induce tolerance in kidney transplantation in several European centers.

the final report to summarize their activities. The first study to emerge in the Framework 7th research program from the EU commission is THE ONE study. This study led by Dr Geissler in Regensburg is going to test a number of cellular therapies to induce tolerance in kidney transplantation in several European centers. The ITN is presently funding several clinical trials in solid organ transplantation, as a continuum of the biomarkers of tolerance study. One of these studies, ARTIST (An Observational Study to Assess the Prevalence of a Tolerance Signature in Renal Transplant Recipients),13 aims to examine a large cohort of renal transplant recipients in order to prospectively determine the frequency and stability of the tolerance signature in patients maintained on IS treatment. The ITN has also made a worldwide call to locate operationally tolerant patients in an effort to create an additional Registry of Tolerant Kidney Transplant Recipients. Finally, the Gradual Withdrawal of Immunosuppression in Patients Receiving a Liver Transplant (AWISH)17 study aims to investigate whether liver transplant recipients can be weaned from immunosuppressive drugs under medical supervision. Another active group within the tolerance biomarker field, based at Stanford University in the United States, is headed by Professor Minnie Sarwal. Their work focuses on candidate gene expression studies in both adult and pediatric liver and kidney transplant patients.15 In collaboration with Professor Jean Paul Soullilou's team, the group described a transcriptional profile, specific for operational tolerant patients in 2007.19 In Barcelona, the group headed by Dr Sanchez-Fueyo is focused on the characterization of biomarkers predictive of tolerance in the setting of liver transplantation. Their findings to date have led to the description of the first protocol in clinical transplantation, in which a tolerance signature has been used to monitor disease and inform decisions on drug withdrawal.20

focused on the characterization of biomarkers predictive of tolerance in the setting of liver transplantation. Their findings to date have led to the description of the first protocol in clinical transplantation, in which a tolerance signature has been used to monitor disease and inform decisions on drug withdrawal.20 METHODOLOGICAL CHALLENGES IN THE STUDY OF BIOMARKERS OF TOLERANCE The major methodological challenge associated with the study of tolerance in kidney transplantation is the ‘hidden' nature of the phenotype. The tolerance state is, as yet, unknown. The patients voluntarily or directed are weaned off IS for a long period of time, and this is not accompanied by rejection; therefore, we assume are tolerant. It is important to point out that any molecular changes may have already occurred because of the tolerance state; however, it may also be partially due to the absence of immunosuppression. For this reason, biomarker studies of tolerance to date have been purely cross-sectional, where patients at all stages of the post-transplant period are allocated to defined clinical groups. As a consequence, it is unknown when the tolerant signature arises and how it develops from the time of transplant. For example, it is unknown whether the signature detected by these studies is detectable if the tolerant patients are still under IS. This raises the problem of the appropriate control comparison group for tolerant patients. Enrolling healthy volunteers can serve as a control for the absence of immunosuppressive drugs, whereas stable patients or chronic rejectors are the ideal clinical comparison groups. It is expected that long-term immune monitoring studies will shed light into this question, making use of a more appropriate longitudinal design, and providing the most valuable type of data for this purpose; that is, the molecular differences between tolerant and stable/chronic rejection patients before successful weaning. Unfortunately, longitudinal studies currently rely on the independent decision of the patients to withdraw their medication, which is a rare and unsafe event. The rarity of the event puts limits to the statistical power of tolerance studies, whereas the high risk limits experimental allocation of patients to a ‘tolerant' condition, as it is not currently ethically appropriate to encourage a patient to undergo weaning.

hdraw their medication, which is a rare and unsafe event. The rarity of the event puts limits to the statistical power of tolerance studies, whereas the high risk limits experimental allocation of patients to a ‘tolerant' condition, as it is not currently ethically appropriate to encourage a patient to undergo weaning. Another interesting, and probably more feasible, design is to detect and follow up patients who are partially weaned off immunosupression for clinical reasons. Comparing the molecular changes in patients who succeed the process with no rejection episodes and those who suffer rejection as a consequence would help to further disentangle the tolerance signature from the drug's signature.

t and follow up patients who are partially weaned off immunosupression for clinical reasons. Comparing the molecular changes in patients who succeed the process with no rejection episodes and those who suffer rejection as a consequence would help to further disentangle the tolerance signature from the drug's signature. To confront the problem of statistical power, most studies establish multicenter and often international collaborations, which necessarily introduce noise due to stratification effects in genetic variation, and differences across countries and centers in clinical protocols. A clear definition of the phenotypes, that is, Stable vs. Tolerant vs. Chronic Rejector, and a careful choice of inclusion and exclusion criteria can help minimize the noise in the data. Thus, although a predictive model is better validated when replicated in miscellaneous populations,21 the original development should be carried out using a restrictive phenotype, for example, ‘A tolerant patient is that who has ceased all medication for at least 1 year, has not suffered any episodes of acute rejection or displayed indications of deteriorating/chronic kidney dysfunction, and whose kidney function has changed <15% in the previous year' is a better candidate than ‘A tolerant patient who has ceased all medication and has not suffered allograft failure'. Generalization to a broader definition of tolerance can be tested subsequently through external validation in an independent patient sample, and the test can be updated when necessary for its application in significantly different populations.22 In addition, the effects of known confounders can be tested explicitly by adding clinical information to the multivariate predictor, for example, donor type, number of transplants, time since transplant, age of donor and recipient, human leukocyte antigen mismatch, and so on.

n in significantly different populations.22 In addition, the effects of known confounders can be tested explicitly by adding clinical information to the multivariate predictor, for example, donor type, number of transplants, time since transplant, age of donor and recipient, human leukocyte antigen mismatch, and so on. WHAT MAKES A GOOD BIOMARKER STUDY? We will now consider the component parts of a well-designed biomarker study. In a recent review, Naesens and Sarwal21 argued that ‘omics' data are currently being used in clinical studies for two different purposes: predictive biomarker discovery, and elucidation of pathophysiological processes to identify therapeutic targets. The ultimate goals of biomarker studies are prediction and prognosis, and this needs to be recognized at all stages of the study, from design planning to data analysis and reporting (see glossary for a description of methodological terms in this section).

of pathophysiological processes to identify therapeutic targets. The ultimate goals of biomarker studies are prediction and prognosis, and this needs to be recognized at all stages of the study, from design planning to data analysis and reporting (see glossary for a description of methodological terms in this section). Design Prospective studies are preferred over cross-sectional studies, and the selection of the time points of interest is a major decision that cannot be taken lightly. It is our experience that involving a multidisciplinary team in the planning stages of the study is vital to reach an optimal design and ensure a smooth running of the study; for example, clinical team, translational research team, laboratory technicians, data manager, and statisticians. An ideal biomarker or set of biomarkers should show significant differences between tolerant and non-tolerant patients either in mean expression at a specified time point after transplant, or in the slope of expression estimated across a number of repeated measures.

oratory technicians, data manager, and statisticians. An ideal biomarker or set of biomarkers should show significant differences between tolerant and non-tolerant patients either in mean expression at a specified time point after transplant, or in the slope of expression estimated across a number of repeated measures. Data analysis Naesens and Sarwal21 propose an ‘integrative omics approach' for translational research. Although it is clear that ‘cross-platform' multivariate predictors that incorporate different types of ‘omics' data are the future, this imposes a serious statistical challenge. The issues associated with the processing and analysis of high-dimensional microarray data have been broadly discussed elsewhere.23, 24 In the context of biomarker studies, it is important to use a data-analysis protocol that places classification accuracy as the top priority. The feature selection method should ideally: use misclassification error or other measures of classification accuracy as criteria for feature selection (genes, molecules, and so on), rather than simply P-values, for example, predictive analysis of microarrays, significant analysis of microarrays, supervised principal components; consider the performance of the features in combination rather than individually in the selection process, for example, classification trees, principal components analysis, regularized regression; be sensitive to correlations and/or interactions between the features, for example, random forest or elastic net (see Hastie et al.25 for an extensive review of data mining methods).

consider the performance of the features in combination rather than individually in the selection process, for example, classification trees, principal components analysis, regularized regression; be sensitive to correlations and/or interactions between the features, for example, random forest or elastic net (see Hastie et al.25 for an extensive review of data mining methods). Subsequently, different classification algorithms can be used to build the final predictive model, and the most appropriate will depend on the data, the proficiency of the analysis team, and the complexity of interactions and correlations present in the selected features. Recently, the MicroArray Quality Control (MAQC) consortium24 demonstrated that ‘good modeling practices were more important than the choice of a particular algorithm' (p 834). Four key points are to be taken from this report: (1) maintain a reasonable ratio of sample size to classifier complexity; (2) use internal validation (cross-validation or bootstrapping) throughout feature selection and development of the classifier, not only at the end; (3) perform external validation in a completely independent sample; and (4) all the steps and decisions taken during the classifier-building procedure should be carefully documented and justified. This last point brings us to the importance of a full report of the analysis process, as well as the use of appropriate measures of performance, to facilitate replication by other research teams.

(4) all the steps and decisions taken during the classifier-building procedure should be carefully documented and justified. This last point brings us to the importance of a full report of the analysis process, as well as the use of appropriate measures of performance, to facilitate replication by other research teams. Reporting The value of a biomarker depends on its ability to discriminate different groups of patients, and in this case tolerant vs stable and/or chronic rejectors, but this should not be the only outcome. In a recent publication, Steyerberg et al.22 describe classic and novel ways to evaluate the predictive value of a new marker. According to these authors, it is essential to quantify the value added by the new marker to currently available, easier to obtain (clinical) variables. Furthermore, accuracy measures such as sensitivity and specificity, or area under the receiving operating curve, provide incomplete information regarding the performance of the new test. Calibration measures are needed to compare outcomes and predictions, especially in the external validation stage; measures related to reclassification provide detailed information regarding the gain of adding a new marker; more importantly, a good sensitivity is not necessarily translated into clinical usefulness, and decision-curve analysis can help to evaluate the value of a marker, taking into consideration the drawbacks and benefits of false positives and true positives, respectively.

mation regarding the gain of adding a new marker; more importantly, a good sensitivity is not necessarily translated into clinical usefulness, and decision-curve analysis can help to evaluate the value of a marker, taking into consideration the drawbacks and benefits of false positives and true positives, respectively. Overall, reports of discovery of new biomarkers, or validation of previous ones, need to focus around a comprehensive description of accuracy of classification and clinical usefulness added to existing clinical measures. A detailed disclosure of the final predictive model, including values of coefficients, is desired to allow for a complete replication and validation by others. Copyrights should be protected by patent applications, rather than by lack of transparency in scientific reports.

ological situations, and lead to killing of target cells. The major activating NK receptors in humans include the NKG2D, CD16, and the natural cytotoxic receptors (NKp46, NKp44, and NKp30). A great number of excellent reviews had been published recently that describe the structure and function of NK-related receptors.3 The ability of NK cells to recognize and kill allogeneic cells without prior sensitization is explained by the concept of ‘missing self'.4 Inhibitory receptors expressed on NK cells interact with self-MHC class I molecules, deliver an inhibitory signal, and lead to inhibition of NK activity. Thus, target cells that lost major histocompatibility complex (MHC) class I expression by virus infection or malignant cell transformation become sensitive to NK cell destruction. However, it is now known that the activation of NK cells requires more than the absence of inhibitory signals, and target cells have to express specific ligands for activating receptors. The NKG2D-activating receptor recognizes several ligands that are induced in response to cellular damage, activate NK cell functions, and lead to the lysis of target cells. In comparison with the missing self-hypothesis, this process was called ‘induced or stressed' self-hypothesis.

sefulness added to existing clinical measures. A detailed disclosure of the final predictive model, including values of coefficients, is desired to allow for a complete replication and validation by others. Copyrights should be protected by patent applications, rather than by lack of transparency in scientific reports. THE FUTURE: TRANSLATION OF BIOMARKERS INTO THE CLINIC Advances in our understanding of human immunological processes and developments in new therapeutic and diagnostic agents make the detection and/or induction of graft tolerance a real possibility in the near future. Many therapeutic agents with potential tolerogenic properties have been described, and some of them are currently undergoing clinical trials (reviewed in St Clair et al.26). However, the lack of well-defined biomarkers of tolerance represents the most significant barrier to the development of tolerance therapeutics. Without these tools, studies will miss an appropriate clinical endpoint to define the operational tolerance state. Long-term prospective studies could address the generation of tolerance but only in the context of IS withdrawal protocols. The only chance for a weaning study to be successful will be a carefully designed one, in which weaning is considered in the presence of increased surveillance and using validated biomarkers of both rejection and tolerance (Figure 1). Notwithstanding, this will imply in the case of kidney allografts a real risk of graft loss. The current efforts in establishing validated and stable biomarkers of tolerance in combination with validated and widely accepted biomarkers of rejection will allow such study to be ethically acceptable.

tolerance (Figure 1). Notwithstanding, this will imply in the case of kidney allografts a real risk of graft loss. The current efforts in establishing validated and stable biomarkers of tolerance in combination with validated and widely accepted biomarkers of rejection will allow such study to be ethically acceptable. The road to the discovery of biomarkers has been enormously boosted by the advances of new tools for high-throughput analysis, which enable us to interrogate the genome, epigenome, transcriptome, and proteome. Together with these advances, new methods for data analysis are also being developed to embrace this new multidimensional data in a holistic rather than reductionistic perspective. The discovery process implies the establishment of correlations between gold standard clinical parameters of disease and changes in biomarkers. To do so, there is a real need for a coordinated approach between international networks to create common biobanks associated with comprehensive clinical records. This is of particular importance in the process of biomarker validation, where large-scale prospective multicenter studies are required. New initiatives such as the Biomedical Research Centers27 in the United Kingdom are of outstanding importance in bringing together both clinical and research expertise, which is an essential step to optimize the resources and funding that translational projects require.

ale prospective multicenter studies are required. New initiatives such as the Biomedical Research Centers27 in the United Kingdom are of outstanding importance in bringing together both clinical and research expertise, which is an essential step to optimize the resources and funding that translational projects require. CONCLUDING REMARKS We aspire that validation of the biomarkers of tolerance will conclude in a clear and robust definition of the hidden phenotype of immune tolerance. This definition will have multiple benefits that will ultimately impact the management of kidney transplant recipients. We foresee a new paradigm in transplantation medicine, where patients will be stratified based on pretransplant-established risk factors such as human leukocyte antigen matching or cold ischemia times, and post-transplant monitoring of non-invasive biomarkers.28 Together, the studies described here provide evidence that it might be possible to develop biomarkers capable of detecting operational tolerance in kidney transplantation. We expect that, in the near future, together with clinical and histological information, better characterized molecular and cellular markers will help us predict the outcome in kidney transplantation, guide personalized IS and weaning protocols, and even develop new therapeutic targets.

rance in kidney transplantation. We expect that, in the near future, together with clinical and histological information, better characterized molecular and cellular markers will help us predict the outcome in kidney transplantation, guide personalized IS and weaning protocols, and even develop new therapeutic targets. MPH-F, EP, and PS acknowledge financial support from the Department of Health via the National Institute for Health Research (NIHR) comprehensive Biomedical Research Centre award to Guy's & St Thomas' NHS Foundation Trust in partnership with King's College London and King's College Hospital NHS Foundation Trust. MPH-F acknowledges financial support from the MRC (G0801537/ID: 88245) and Guy's & St Thomas's Charity (grant 080530). IR-M acknowledges financial support from the MRC. We thank Dr Mike Weale for his critical reading of the manuscript. TO CITE THIS ARTICLE: Perucha E, Rebollo-Mesa I, Sagoo P et al. Biomarkers of tolerance: searching for the hidden phenotype. Kidney Int Sup 2011; 1: 40–46. MPH-F has filed biomarkers of tolerance as intellectual property (IP); however, this has not generated income or royalties to date.

ic ligands for activating receptors. The NKG2D-activating receptor recognizes several ligands that are induced in response to cellular damage, activate NK cell functions, and lead to the lysis of target cells. In comparison with the missing self-hypothesis, this process was called ‘induced or stressed' self-hypothesis. During development, NK cells experience a process of functional maturation referred to as ‘NK-cell education' that involves the interaction with self-MHC class I molecules via inhibitory receptors. Two models have been proposed to explain this process. The ‘licensing' hypothesis postulates that the engagement of MHC class I to inhibitory receptors on NK cells during development allows these cells to become competent effector cells.5 Although the ‘disarming' model suggests that the absence of MHC class I molecules on normal cells would chronically stimulate NK cells and render them anergic, recent studies demonstrated that the educational process of NK cells is regulated by the number and the affinity of each inhibitory receptor for self-MHC class I molecules.

MPH-F, EP, and PS acknowledge financial support from the Department of Health via the National Institute for Health Research (NIHR) comprehensive Biomedical Research Centre award to Guy's & St Thomas' NHS Foundation Trust in partnership with King's College London and King's College Hospital NHS Foundation Trust. MPH-F acknowledges financial support from the MRC (G0801537/ID: 88245) and Guy's & St Thomas's Charity (grant 080530). IR-M acknowledges financial support from the MRC. We thank Dr Mike Weale for his critical reading of the manuscript. TO CITE THIS ARTICLE: Perucha E, Rebollo-Mesa I, Sagoo P et al. Biomarkers of tolerance: searching for the hidden phenotype. Kidney Int Sup 2011; 1: 40–46. MPH-F has filed biomarkers of tolerance as intellectual property (IP); however, this has not generated income or royalties to date. Figure 1 Biomarker-led management of kidney transplants. Pretransplant assessment: risk is assigned according to anti-donor immune responses, anti-donor antibodies, human leukocyte antigen (HLA) mismatches, and genetic risk. Biomarkers of rejection will assess whether acute rejection will develop in the initial months post-transplantation, the time period depending on the depletion agent used. To maximize organ function, the treatment could be modified before tissue injury occurs. Two time points use biomarkers of tolerance: first, to establish the success of any peritransplant tolerance-inducing protocol. Later, to establish whether a tolerance state has been reached, the main aim would be to wean those patients off with positive biomarkers of tolerance. Throughout weaning, biomarkers of rejection could be used to stop the process before tissue injury is evident. Obviously, if a patient is negative for the biomarkers of tolerance, then the standard protocol of immunosuppression should be maintained for life.

ean those patients off with positive biomarkers of tolerance. Throughout weaning, biomarkers of rejection could be used to stop the process before tissue injury is evident. Obviously, if a patient is negative for the biomarkers of tolerance, then the standard protocol of immunosuppression should be maintained for life. Table 1 Examples of current international efforts towards conducting translational research and biomarker discovery in solid organ transplantation Transplant tolerance-related clinical study Organ Lead institution Country Ref. Genetic analysis and monitoring of biomarkers of immunological tolerance (GAMBIT) Kidney King's College London United Kingdom 10 An observational study to assess the prevalence of a tolerance signature in renal transplant recipients (ARTIST) Kidney ITN USA 13 ITN registry of tolerant kidney transplant recipients Kidney ITN USA 14 Prediction and mechanisms of transplantation tolerance Kidney Stanford University (Sarwal lab) USA 15 Effect of immunosuppression drug weaning on hepatitis C virus (HCV)-induced liver damage after liver transplantation Liver Hospital Clinic of Barcelona Spain 16 Gradual withdrawal of immunosuppression in patients receiving a liver transplant (AWISH) Liver ITN USA 17 Abbreviation: ITN, Immune Tolerance Network.

Kidney transplantation is the treatment of choice for end-stage renal disease owing to the use of potent immunosuppressive therapy. Current immunosuppression is highly effective in avoiding acute rejection, but it is associated with nephrotoxicity, cardiovascular morbidity, infection, and cancer. Therefore, the improvement in short-term graft survival has not been reflected in improved long-term outcomes.1 Current immunosuppression strategies are primarily based on the use of an induction regimen using a monoclonal or polyclonal antibody, followed by maintenance immunosuppression based on a calcineurin inhibitor (CNI), an anti-proliferative agent, and low-dose corticosteroids.2 Several CNI minimization and withdrawal protocols have been attempted with varied results.3 The use of mammalian target of rapamycin inhibitors for CNI minimization or withdrawal has been hampered by their adverse side-effect profile.4, 5 Therefore, it is accepted that CNIs currently remain the cornerstone of maintenance immunosuppression in renal transplantation.6 Undoubtedly, new drugs dealing with new mechanisms, as well as minimizing comorbidities, are warranted in renal transplantation. The present trend in drug development is focused on preservation of long-term function and minimization of the adverse events of immunosuppressive drugs.7 Several small molecules and biological agents are currently being studied. Belatacept is a humanized antibody that inhibits T-cell co-stimulation and has shown encouraging results in Phase II and III trials. Moreover, two new small molecules are under clinical development: AEB071 or sotrastaurin (a protein kinase C (PKC) inhibitor) and CP-690550 (a Janus kinase (JAK) inhibitor). All three drugs are excellent examples to show how difficult it is to develop new immunosuppressants to overcome the CNI toxicities. Refinement in selecting the best combinations for the new and some current immunosuppressive agents is probably the main challenge for next few years.

50 (a Janus kinase (JAK) inhibitor). All three drugs are excellent examples to show how difficult it is to develop new immunosuppressants to overcome the CNI toxicities. Refinement in selecting the best combinations for the new and some current immunosuppressive agents is probably the main challenge for next few years. BELATACEPT In the near future, belatacept would be the first biological agent for use in long-term maintenance regimen in organ transplantation. Its parent molecule was CTLA4-Ig (abatacept), and resulted from the fusion of the extracellular domain of CTLA4 with the constant region fragment of human IgG1.8 As CTLA4 shows high affinity for CD80/86, CTLA4-Ig may block antigen-presenting cell co-stimulation of T cells through CD28, thereby abrogating the immune response. Abatacept was proved to be highly effective for autoimmune T-cell-mediated autoimmune disorders,9 but inadequate as a maintenance immunosuppressive agent in non-human primate models of transplantation.10 Potential explanation could be that abatacept is significantly less potent in inhibiting CD86-dependent as opposed to CD80 co-stimulation.11 A daughter molecule LEA29Y (belatacept) with two amino-acid substitutions (L104->E and A29->Y) was developed.12 Belatacept was found to bind four times more avidly to CD86 and two times more avidly to CD80 than the parent abatacept. This improved binding resulted in ∼10-fold more potent inhibition of T-cell activation.12 Belatacept efficacy was demonstrated in several non-human primate preclinical models of renal transplantation, particularly in combination with basiliximab and other immunosuppressants such as mycophenolate mofetil (MMF). In addition, belatacept was suggested as being capable of preventing development of donor-specific antibodies.13

icacy was demonstrated in several non-human primate preclinical models of renal transplantation, particularly in combination with basiliximab and other immunosuppressants such as mycophenolate mofetil (MMF). In addition, belatacept was suggested as being capable of preventing development of donor-specific antibodies.13 Clinical development of belatacept The first clinical trial on the use of belatacept in clinical renal transplantation was a Phase II non-inferiority trial comparing the efficacy of belatacept vs cyclosporine (CsA) for prevention of acute rejection at 6 months post-transplant.14 Belatacept was administered as less (LI) or more intensive (MI) schedule. All patients also received MMF and corticosteroids as maintenance immunosuppression, and induction with basiliximab. At 6 months, the incidence of acute rejection was similar in all three groups ranging from 6 to 8%. The LI group experienced a higher incidence of subclinical rejection and treated episodes of subclinical rejection compared with the MI and CsA groups. Glomerular filtration rate was significantly higher in the belatacept groups compared with the CsA arm. Protocol biopsies demonstrated a significant reduction in the incidence of chronic allograft nephropathy in the belatacept group. Cardiovascular profile improved in the belatacept groups as they had a statistically significant lower risk of developing diabetes, need for treatment of hyperlipidemia, and a lower incidence of hypertension.

demonstrated a significant reduction in the incidence of chronic allograft nephropathy in the belatacept group. Cardiovascular profile improved in the belatacept groups as they had a statistically significant lower risk of developing diabetes, need for treatment of hyperlipidemia, and a lower incidence of hypertension. Two Phase III studies have been recently published. The BENEFIT (Belatacept Evaluation of Nephroprotection and Efficacy as First line Immunosuppression Trial) study is a 3-year Phase III clinical trial, which randomized patients to three groups as previously described. Patient and graft survival are similar in three groups.15, 16 However, the incidence of acute rejection was greater in MI (22%) and LI (17%) belatacept compared with CsA (7%), although no apparent impact on graft survival was demonstrated. Most acute rejection episodes occurred within the first 3 months and severity was higher in belatacept compared with CsA-treated patients. At the end of 2 years, glomerular filtration rate continued to be significantly higher in the belatacept-treated patients. Belatacept-treated patients also had sustained benefits in their cardiovascular and metabolic risk profile. The BENEFIT-EXT (Belatacept Evaluation of Nephroprotection and Efficacy as First-line Immunosuppression Trial—EXTended criteria donors) study is a 3-year randomized Phase III study in patients receiving an extended-criteria-donor kidney allograft.17 Patient and graft survival were similar in all the three groups. Renal function was statistically superior in MI belatacept vs CsA but not in LI vs CsA group. The incidence of chronic allograft nephropathy was similar in the three groups, probably because of the presence of renal damage at baseline. Interestingly, cardiovascular risk factors were lower in the belatacept-treated patients. The 2-year results showed similar trend.16

belatacept vs CsA but not in LI vs CsA group. The incidence of chronic allograft nephropathy was similar in the three groups, probably because of the presence of renal damage at baseline. Interestingly, cardiovascular risk factors were lower in the belatacept-treated patients. The 2-year results showed similar trend.16 Vincenti et al.18 recently published the 5-year safety data of their initial Phase II study. Belatacept-treated patients did not have a higher frequency of serious infections or post-transplant lymphoproliferative disorder (PTLD) compared with CsA. Remarkably, major cardiac adverse events occurred more frequently with CsA (2% belatacept vs 12% CsA). Of the three major trials described above, a total of 13 patients in the belatacept groups have been diagnosed with PTLD (1.4%) compared with only one CsA patient (0.2%). Of the 13 cases of PTLD identified, six developed PTLD of the central nervous system. This is a diagnosis to be concerned about considering that this involvement is seldom encountered in solid organ transplantation.19 The majority of patients who developed PTLD had known risk factors for PTLD, including pre-transplant EBV-seronegative recipients, those receiving lymphocyte-cell-depleting agents, and those having a primary EBV infection.

d about considering that this involvement is seldom encountered in solid organ transplantation.19 The majority of patients who developed PTLD had known risk factors for PTLD, including pre-transplant EBV-seronegative recipients, those receiving lymphocyte-cell-depleting agents, and those having a primary EBV infection. In these lines, a recent Phase II analysis that excluded transplant recipients who were EBV seronegative before transplant has yet to report any cases of PTLD.20 The absence of PTLD in this analysis may be evidence enough to avoid use of belatacept in those recipients who are EBV seronegative before transplant. A total of 89 patients were randomized to receive belatacept-MMF, belatacept-sirolimus, or tacrolimus-MMF. All patients received thymoglobulin induction. Renal function was better in the belatacept-treated groups. Acute rejection occurred in four, one, and one patient in the belatacept-MMF, belatacept-SRL, and TAC-MMF groups, respectively. The authors concluded that the use of belatacept in renal transplantation may allow CNI and corticosteroid avoidance, with acceptable rates of acute rejection and improved glomerular filtration rate.

ejection occurred in four, one, and one patient in the belatacept-MMF, belatacept-SRL, and TAC-MMF groups, respectively. The authors concluded that the use of belatacept in renal transplantation may allow CNI and corticosteroid avoidance, with acceptable rates of acute rejection and improved glomerular filtration rate. Results from a Phase II study to evaluate conversion from a CNI-based regimen to belatacept have been recently published.21 This randomized, open-label study included 171 patients, 6 months to 3 years post-transplant, who were receiving CNI-based immunosuppression and had stable renal function. Patients were randomized into one of two treatment groups: conversion to belatacept 5 mg/kg with CNI discontinuation (n=83), or continued CNI therapy (n=88). At month 12, the mean change from baseline in glomerular filtration rate was higher in the belatacept group compared with the CNI group. Six patients in the belatacept group had acute rejection episodes, all within the first 6 months, and all resolved with no allograft loss. The overall safety profile was similar in each treatment group.

an change from baseline in glomerular filtration rate was higher in the belatacept group compared with the CNI group. Six patients in the belatacept group had acute rejection episodes, all within the first 6 months, and all resolved with no allograft loss. The overall safety profile was similar in each treatment group. In summary, some concerns appear with the use of belatacept. It was originally anticipated that co-stimulation blockade would be successful in achieving immunological allograft tolerance; however, based on higher acute rejection rates, this does not appear to be the case, probably because of its inhibitory effect on regulatory T-cell expansion by MI belatacept regime in combination with basiliximab. Another limitation of this medication is that administration requires an intravenous infusion. As there is increased PTLD risk in EBV-seronegative patients, belatacept should be prescribed for EBV-seropositive patients only. Nevertheless, clinical trials suggest that the use of belatacept can lead to better renal function along with a lower incidence of diabetes and cardiovascular risk factors. Currently, on the basis of the manufacturer's recommendation, the Food and Drug Administration is reviewing the LI-dosing regimen of belatacept as an immunosuppressive regimen in kidney transplant recipients. This LI belatacept protocol is associated with low acute rejection rates, but maintains a renal and cardiovascular favorable profile.

s of the manufacturer's recommendation, the Food and Drug Administration is reviewing the LI-dosing regimen of belatacept as an immunosuppressive regimen in kidney transplant recipients. This LI belatacept protocol is associated with low acute rejection rates, but maintains a renal and cardiovascular favorable profile. JANUS KINASE INHIBITORS (CP-690550 OR TASOCITINIB) Janus kinases are cytoplasmic tyrosine kinases involved in cell proliferation, growth, and survival by integrating extracellular signaling induced by cytokines.22 For instance, after co-stimulation, type I cytokines (interleukin (IL)-2, IL-4, IL-7, IL-9, IL-15, IL-2) bind to cell surface receptor members of the cytokine receptor common gamma (γc) chain family and activate JAKs. Nevertheless, JAKs participate in the signaling of many cytokine receptors in several cell types. Activation of JAKs induces cytokine receptor phosphorylation, as well as recruitment of signal transducers and activators of transcription, and catalyzes phosphorylation of signal transducers and activators of transcription that facilitates its dimerization and transportation to the nucleus where they regulate gene expression. There are four JAKs identified in mammals: JAK-1, 2, 3, and tyrosine kinase-2. JAK1 is activated by gp130 cytokines, type I interferon, interferon-γ, and βc cytokines; its deficiency causes neurological defects and severe combined immunodeficiency. JAK2 is activated by erythropoietin, thrombopoietin, prolactin, growth hormone, γc cytokines, interferon-γ, and IL-12; its deficiency is lethal because of defective erythropoiesis. JAK3 is mainly expressed in hematopoietic cells and just activated by γc cytokines; its deficiency causes severe combined immunodeficiency. Tyrosine kinase-2 is activated by gp130 cytokines, type I interferon, IL-12, and IL-23; its deficiency induces minor effects.23 Therefore, compared with other JAKs, which are ubiquitous and activated by several types of cytokines, its specificity makes JAK3 an interesting target for immunosuppression. However, there is high structural similarity between JAK2 and JAK3 that makes it difficult to synthesize compounds that are able to inhibit JAK3 without affecting JAK2. This is crucial as the safety profile depends on JAK3 inhibition selectivity.

, its specificity makes JAK3 an interesting target for immunosuppression. However, there is high structural similarity between JAK2 and JAK3 that makes it difficult to synthesize compounds that are able to inhibit JAK3 without affecting JAK2. This is crucial as the safety profile depends on JAK3 inhibition selectivity. CP-690550 or tasocitinib is a synthetic orally available inhibitor of JAK3 that maintains reasonable selectivity for JAK3. In vitro and animal studies demonstrated its potency and capacity to prevent rejection even in cynomolgus monkeys.24 Therefore, CP-690550 is currently under clinical development.

, its specificity makes JAK3 an interesting target for immunosuppression. However, there is high structural similarity between JAK2 and JAK3 that makes it difficult to synthesize compounds that are able to inhibit JAK3 without affecting JAK2. This is crucial as the safety profile depends on JAK3 inhibition selectivity. CP-690550 or tasocitinib is a synthetic orally available inhibitor of JAK3 that maintains reasonable selectivity for JAK3. In vitro and animal studies demonstrated its potency and capacity to prevent rejection even in cynomolgus monkeys.24 Therefore, CP-690550 is currently under clinical development. A double-blinded, placebo-controlled Phase I trial assessing safety and tolerability of CP-690550 (5, 15, 30 mg b.i.d.) in renal transplant recipients reported that most adverse events were gastrointestinal or infectious.25 In addition, high CP-690550 doses were associated with reduction of hemoglobin levels, demonstrating its inhibitory effect on JAK2. Further studies confirm that, although highly specific for JAK3, CP-690550 also inhibits JAK2 to some extent.26 A 6-month Phase II trial and its extension to 12 months have been published.27 In this trial, 61 adult renal transplant recipients were randomized to CP-690550 15 mg or 30 mg b.i.d., vs tacrolimus in combination with an IL-2 receptor antagonist, MMF, and steroids. In the high-dose arm, an increased incidence of BK virus nephropathy and cytomegalovirus infection required a protocol amendment, based on planned MMF withdrawal and rapid steroid taper. The consequence was 21.1% incidence of acute rejection in the high-dose arm. However, the low-dose arm provided excellent results that showed a 5.3% incidence of acute rejection and 76.9 ml/min glomerular filtration rate. These results were confirmed in the 12-month extension protocol in which CP-690550 was reduced to 15 mg b.i.d. In the CP-690550 arms, there was a trend toward more frequent anemia and neutropenia. Overall, the efficacy/safety profile of CP-690550 at 15 mg b.i.d. was comparable to tacrolimus, with the exception of a higher rate of viral infection. These results were used for designing ongoing protocols exploring the effects of a lower dose of CP-690550 in renal transplantation (5 and 10 mg b.i.d.). These preliminary data suggest that CP-690550 has the potential to improve current immunosuppression armamentarium. However, there still exist some concerns. Anemia is a common adverse event that has been reported in 30% of patients enrolled in the Phase II trial; lower doses and new combination strategies should be explored and, finally, new molecules with high JAK3 selectivity warranted.

ve current immunosuppression armamentarium. However, there still exist some concerns. Anemia is a common adverse event that has been reported in 30% of patients enrolled in the Phase II trial; lower doses and new combination strategies should be explored and, finally, new molecules with high JAK3 selectivity warranted. SOTRASTAURIN (AEB071) Protein kinase C has an important role in the immune response. It is well known that T-cell receptor activation with co-stimulation signaling leads to PKC activation and IL-2 production.28, 29, 30 On the basis of cofactor requirements, there are at least 10 PKC isoforms that can be divided into three categories: classical or conventional, novel, and atypical. The α, β, and θ isoforms appear to have clear roles in either T- or B-cell signaling, thus suggesting that inhibition of several isoforms are needed to achieve full immunosuppression. The best characterized is PKCθ, which is mostly restricted to T lymphocytes and mediates activation of the transcription factors activator protein-1 and nuclear factor κB, leading to IL-2 production. In fact, knockout of PKCθ impairs T-cell activation in mice.31

al isoforms are needed to achieve full immunosuppression. The best characterized is PKCθ, which is mostly restricted to T lymphocytes and mediates activation of the transcription factors activator protein-1 and nuclear factor κB, leading to IL-2 production. In fact, knockout of PKCθ impairs T-cell activation in mice.31 Sotrastaurin is a small molecule that inhibits PKC activity, including classical (α, β) and novel (δ, ɛ, η, θ) isoforms. Similar to CNIs, sotrastaurin principally inhibits PKCθ acting on IL-2 gene promoters. Nevertheless, it has insignificant effect on downstream targets of calcineurin, such as nuclear factor of activated T cells.32, 33 This feature led investigators to hypothesize that sotrastaurin can be as potent as CNIs without displaying nephrotoxicity. Non-human primate and healthy human volunteer studies have endorsed those in vitro sotrastaurin characteristics. Sotrastaurin, in monotherapy or in combination with other immunosuppressants, prolongs allograft survival in rats and cynomolgus monkeys.34, 35

rin can be as potent as CNIs without displaying nephrotoxicity. Non-human primate and healthy human volunteer studies have endorsed those in vitro sotrastaurin characteristics. Sotrastaurin, in monotherapy or in combination with other immunosuppressants, prolongs allograft survival in rats and cynomolgus monkeys.34, 35 Preclinical and early clinical safety data demonstrated no signs of nephrotoxicity or hepatotoxicity, and no metabolic or blood pressure effects at standard exposures.28, 29 Gastrointestinal effects were the dose-limiting toxicities in all species tested preclinically. In vitro tests indicated a modest potential for QT prolongation. However, in healthy volunteer studies, QT effects could not be confirmed at therapeutic doses. A reversible increase in mean ventricular heart rate was observed at a single dose of 500 mg, with mean heart rates remaining within the normal range.36 Similar to CNIs and mammalian target of rapamycin inhibitors, compensatory reduction in the dose of sotrastaurin is warranted when strong CYP3A4 inhibitors are coadministered. 37 In a proof-of-concept study in patients with psoriasis, clinical severity was reduced by 69% after a 2-week treatment with sotrastaurin. This effectiveness was dose dependent and achieved with good drug tolerability.38

Preclinical and early clinical safety data demonstrated no signs of nephrotoxicity or hepatotoxicity, and no metabolic or blood pressure effects at standard exposures.28, 29 Gastrointestinal effects were the dose-limiting toxicities in all species tested preclinically. In vitro tests indicated a modest potential for QT prolongation. However, in healthy volunteer studies, QT effects could not be confirmed at therapeutic doses. A reversible increase in mean ventricular heart rate was observed at a single dose of 500 mg, with mean heart rates remaining within the normal range.36 Similar to CNIs and mammalian target of rapamycin inhibitors, compensatory reduction in the dose of sotrastaurin is warranted when strong CYP3A4 inhibitors are coadministered. 37 In a proof-of-concept study in patients with psoriasis, clinical severity was reduced by 69% after a 2-week treatment with sotrastaurin. This effectiveness was dose dependent and achieved with good drug tolerability.38 Results in renal transplantation have not been as good as it was expected.36 In one trial, patients were initially placed on tacrolimus/sotrastaurin/steroids treatment and then underwent conversion from tacrolimus to sodium mycophenolate (MPA) at 3 months, which resulted in an increased incidence of the primary composite endpoint (acute rejection, graft loss, or death) in the sotrastaurin arm. In another trial, de novo CNI-free arm of sotrastaurin/MPA/steroids was compared with tacrolimus/MPA/steroids. Again, acute rejection rate was higher in the sotrastaurin arm. These studies were prematurely stopped and the results from the first sotrastaurin Phase II trial in renal transplantation have been recently published.36 A total of 216 patients were randomized, 74 were allocated to control-MPA+standard exposure tacrolimus, 76 to sotrastaurin 200 mg b.i.d.+standard exposure tacrolimus, and 66 to sotrastaurin 200 mg b.i.d.+reduced exposure tacrolimus. Sotrastaurin-treated recipients who met conversion criteria at month 3 were converted to a CNI-free regimen of sotrastaurin 200 mg b.i.d.+MPA 720 mg b.i.d. During the 3-month pre-conversion period, all regimens showed comparable efficacy to control for the composite endpoint (acute rejection, graft loss, or death). However, after conversion from tacrolimus to MPA, both sotrastaurin+MPA regimens were inferior to the control for the primary composite endpoint. Thus, composite efficacy failure rates were 7.8, 44.8, and 34.1% at study end in the control, sotrastaurin+standard exposure tacrolimus, and sotrastaurin+reduced exposure tacrolimus. The majority of biopsies in the sotrastaurin groups were Banff IA or IB. Therefore, the initial sotrastaurin-tacrolimus regimen was efficacious and well tolerated, but the post-conversion sotrastaurin-MPA regimen showed inadequate efficacy. With regard to adverse events, gastrointestinal toxicities were frequent, mild, and similar in all groups.36 Tachycardia occurred at a higher incidence in both sotrastaurin groups compared with control in the peritransplant period, but returned to baseline levels at 1 week.

aurin-MPA regimen showed inadequate efficacy. With regard to adverse events, gastrointestinal toxicities were frequent, mild, and similar in all groups.36 Tachycardia occurred at a higher incidence in both sotrastaurin groups compared with control in the peritransplant period, but returned to baseline levels at 1 week. Interestingly, the incidence of new-onset diabetes in the control group (14.9%) was nearly twice that in the sotrastaurin groups (6.7 and 7.7%). The sotrastaurin-tacrolimus combination seems to be as effective as tacrolimus-MPA, at least in the short term. This may be the rationale for designing studies of sotrastaurin with CNI minimization.28 In these lines, it has been recently reported that tacrolimus does not alter the pharmacokinetics of sotrastaurin; however, sotrastaurin increases tacrolimus area under the concentration-time curve by twofold.39 Given the lack of efficacy of the sotrastaurin+MPA regimen, attention has turned to explore sotrastaurin in combination with everolimus in a new Phase II trial. This trial is currently underway in Europe. There is some pharmacokinetic interaction between both drugs. Sotrastaurin exposure does not seem to be altered by everolimus, but sotrastaurin increases everolimus exposure by 20%.40

n has turned to explore sotrastaurin in combination with everolimus in a new Phase II trial. This trial is currently underway in Europe. There is some pharmacokinetic interaction between both drugs. Sotrastaurin exposure does not seem to be altered by everolimus, but sotrastaurin increases everolimus exposure by 20%.40 CONCLUSION This review clearly illustrates the difficulty faced in developing new immunosuppressants to overcome the CNI toxicities. On one hand, belatacept given in combination with basiliximab, steroids, and MMF is associated with both PTLD in EBV-negative recipients and more rejection. On the other hand, tasocitinib is highly effective in preventing rejection, but is associated with some over-immunosuppression complications. Finally, sotrastaurin shows lack of efficacy in combination with an antiproliferative agent. Altogether, these findings suggest that refinement in selecting the best combinations for the new and some current immunosuppressive agents is probably the main challenge for the next years. This work was supported by the Red de Investigación Renal (REDinREN, ISCIII 06/0016). TO CITE THIS ARTICLE: Cruzado JM, Bestard O, Melilli E et al. Targets of new immunosuppressants in renal transplantation. Kidney Int Sup 2011; 1: 47–51. All the authors declared no competing interests.

Natural killer (NK) cells, described by several groups in the early 1970s, are important components of the innate immune system and can kill target cells without a requirement for a prior exposure to these targets, in contrast to cytolytic T cells.1, 2 These cells mediate their effector functions through a complex integration of inhibitory and activating receptors that interact with different ligands expressed on host cells. Activated NK cells produce cytolytic molecules (perforin and granzymes) that can directly kill allogeneic cells and proinflammatory cytokines (tumour necrosis factor-α/β, interferon (IFN)-γ) that activate the adaptive immune responses involved in the rejection. The main inhibitory receptors, such as the killer Ig-like receptors, leukocyte Ig-like inhibitory receptors, and the C-type lectin receptor superfamily (CD94/NKG2A), recognize self-MHC class I molecules that are expressed on healthy cells and protect them from NK-induced lysis. In contrast, the activating receptors bind mainly to stress-induced molecules, or molecules found in pathological situations, and lead to killing of target cells. The major activating NK receptors in humans include the NKG2D, CD16, and the natural cytotoxic receptors (NKp46, NKp44, and NKp30). A great number of excellent reviews had been published recently that describe the structure and function of NK-related receptors.3

‘disarming' model suggests that the absence of MHC class I molecules on normal cells would chronically stimulate NK cells and render them anergic, recent studies demonstrated that the educational process of NK cells is regulated by the number and the affinity of each inhibitory receptor for self-MHC class I molecules. During bone marrow transplantation, NK cells have a major role in the rejection of donor bone marrow cells, but the impact of donor and recipient NK cells in solid organ allografts is a controversial point even today. First, studies showed that heart transplants in Rag−/− mice, which are deficient in T and B cells, but maintain an intact NK cell population, are accepted indefinitely and survive throughout the length of the experiment, suggesting that NK cells are not sufficient to reject a heart allograft. However, recent studies highlight the ability of NK cells to promote graft injury or rejection.6, 7, 8, 9 The cellular microenvironment and the interactions with other cells (dendritic cells (DCs), T and B cells) may modulate the immune responses developed against the donor and may contribute to these ambiguous effects of NK cells. Furthermore, the functions of NK cells are determined ultimately by integrated signals obtained from both inhibitory and activating receptors, which are expressed in a stochastic pattern. The inhibitory and activating receptor profiles on NK cells aid in identifying the many subsets of NK cells with different functional properties. Taken together, NK cells may have some functional plasticity during transplantation, so that they can contribute to promoting rejection or inducing tolerance on the basis of the surrounding milieu.

vating receptor profiles on NK cells aid in identifying the many subsets of NK cells with different functional properties. Taken together, NK cells may have some functional plasticity during transplantation, so that they can contribute to promoting rejection or inducing tolerance on the basis of the surrounding milieu. As we have previously reported, NKG2D is one of the most potent activating receptors expressed on NK cells, and induces a potent cytotoxic response in humans through its association with the adaptor molecule DAP10 (ref. 10). The interaction of NKG2D with its ligands has been involved in multiple processes and has an important role in infections, tumors, and autoimmune diseases. Recently, several studies reported the contribution of NKG2D and its ligands in the solid organ transplantation, and suggest a great potential for therapeutic intervention of this interaction. NKG2D LIGANDS IN SOLID ORGAN TRANSPLANTATION The ligands of NKG2D in humans are MHC class I chain-related A and B (MICA, MICB), and ULBP 1–5 (UL-16-binding protein).11 Mice lack MIC genes but express the ULBP homologous proteins, namely RAE-1, H60, and MULT-1. MICA/B genes are encoded in the MHC region and share 28–35% homology to human leukocyte antigen class I genes. The MICA/B genes are highly polymorphic, with more than 72 MICA and 31 MICB recognizable alleles (http://www.ebi.ac.uk/imgt/hla).

genes but express the ULBP homologous proteins, namely RAE-1, H60, and MULT-1. MICA/B genes are encoded in the MHC region and share 28–35% homology to human leukocyte antigen class I genes. The MICA/B genes are highly polymorphic, with more than 72 MICA and 31 MICB recognizable alleles (http://www.ebi.ac.uk/imgt/hla). The main findings have focused on the presence of anti-MICA antibodies in patients with a kidney or heart transplant and their direct involvement in the failure of the allograft. The development of anti-MICA antibodies after kidney transplantation is a significant risk factor in the outcome of the graft, and is independent of the presence of anti-HLA antibodies.12 Moreover, increased rates of allograft failure in human leukocyte antigen-well-matched patients were observed among those with antibodies against MICA before transplantation.13 Our group has also described a high correlation between the presence of anti-MICA antibodies and increased risk of acute rejection during the first year after heart transplantation.14 Because excellent reviews have been published regarding the humoral response against MICA; this article will focus on the cellular immune response.

as also described a high correlation between the presence of anti-MICA antibodies and increased risk of acute rejection during the first year after heart transplantation.14 Because excellent reviews have been published regarding the humoral response against MICA; this article will focus on the cellular immune response. NKG2D ligands are expressed at low levels in many normal cells. Their expression is increased on stressed cells by infection or malignant transformation, and it alerts the immune system of adverse cellular conditions. Increased MICA/B expression has been observed in several transplanted grafts with histological evidence of rejection and/or cellular injury14, 15, 16 (Table 1). Studies in animal models showed that ischemia/reperfusion injury (IRI), which is caused after transplantation, induces expression of RAE-1 on tubular epithelial cells, permits the infiltration of NK cells inside the graft, and increases the risk of acute allograft rejection by NKG2D-dependent mechanisms.17, 18 Furthermore, the simultaneous expression of Rae-1, H60, and NKG2D during acute heart allograft rejection support the role of this interaction in the immune responses associated with transplantation.19 Whether an analogous mechanism occurs in humans still needs to be elucidated; however, it is known that IRI leads to the activation of endothelial cells, which increase their expression of adhesion molecules and recruit effector cells into the tissue. Recently, hypoxia-inducible factor-1 was shown to increase the MICA expression on human renal tubular epithelial cells and cardiomyocytes during the hypoxia/reoxygenation process that occurs during IRI.20, 21 These findings provide the first insights on the IRI mechanisms that induce MICA expression, and can spur the development of new strategies to reduce the early renal inflammatory injury after transplantation.

ular epithelial cells and cardiomyocytes during the hypoxia/reoxygenation process that occurs during IRI.20, 21 These findings provide the first insights on the IRI mechanisms that induce MICA expression, and can spur the development of new strategies to reduce the early renal inflammatory injury after transplantation. One of the best known mechanisms of immune evasion in tumor cells is the proteolytic release of MICA from the cell surface. The endoplasmic reticulum protein 5 interacts with MICA in the cell membrane and induces a conformational change that is necessary for proteolytic cleavage of the molecule. We identified a soluble form of MICA in the serum samples, from heart transplant patients, obtained during the first year after transplant.22 Transplant patients with high levels of soluble MICA showed a lower incidence of acute rejection and had higher graft function and survival. In vitro studies demonstrated that soluble MICA engages NK cells expressing NKG2D, induces receptor internalization and degradation, and impairs the NKG2D-mediated allogenic cytolytic responses. This finding supports a new mechanism by which NKG2D ligands could contribute to thwarting allograft rejection, and elucidates its inhibition of effector functions mediated by NK and CD8+ T-cytotoxic cells.

, induces receptor internalization and degradation, and impairs the NKG2D-mediated allogenic cytolytic responses. This finding supports a new mechanism by which NKG2D ligands could contribute to thwarting allograft rejection, and elucidates its inhibition of effector functions mediated by NK and CD8+ T-cytotoxic cells. NKG2D MEDIATES ALLOGRAFT INJURY The NKG2D-activating receptor is expressed in all NK cells, γδ T, αβ CD8 T, and NKT cells.23 NKG2D functions as a primary activating receptor in NK cells, and a co-stimulatory receptor in CD8+ T cells. In these cells, a T-cell receptor-mediated stimulus is necessary for activation, although, under certain circumstances, NKG2D might be able to function as an activating receptor, independent of T-cell receptor stimulation. Recently, studies in mouse models deficient in NKG2D showed that this receptor is involved in the development, homeostasis, and survival of NK cells. NK cells from NKG2D-deficient mice show a higher proliferation rate of immature NK cells and are more susceptible to apoptosis.24 NKG2D signaling is also implicated in the effector functions of NKs as these NKG2D−/− NK cells showed a weaker cytolytic response and lower IFN-γ production against target cells expressing NKG2D ligands.

NKG2D-deficient mice show a higher proliferation rate of immature NK cells and are more susceptible to apoptosis.24 NKG2D signaling is also implicated in the effector functions of NKs as these NKG2D−/− NK cells showed a weaker cytolytic response and lower IFN-γ production against target cells expressing NKG2D ligands. The activation of NK cells through the NKG2D receptor bound to its ligands may regulate multiple immunological pathways involved in the rejection or the outcome of allograft transplantation (Table 1). The best known example of the NKG2D function is reported in bone marrow transplantation of mice.25 NK cells reject the bone marrow cells expressing the NKG2D ligand, Rae-1, and the blockade with a neutralizing non-depleting NKG2D monoclonal antibody prevents rejection. Similar results were obtained using a mouse model with severe combined immunodeficiency disease recipients bearing MHC class II-deficient skin allografts.8 Transferred CD4 T cells, which recognize alloantigens only through the indirect pathway, mediate rejection by a NK cell-dependent route. Inflammation induced by indirectly primed CD4+ T cells leads to the upregulation of NKG2D ligands in the allografts. Concurrently, these CD4+ T cells recruit and trigger activation of NK cells through the interactions of the NKG2D-activating receptor with its ligands on donor cells. These results were corroborated so that NKG2D blocking significantly prolonged survival but did not induce a permanent acceptance, probably because of the involvement of other activating NK receptors.

igger activation of NK cells through the interactions of the NKG2D-activating receptor with its ligands on donor cells. These results were corroborated so that NKG2D blocking significantly prolonged survival but did not induce a permanent acceptance, probably because of the involvement of other activating NK receptors. Only some indirect evidence has reported the involvement of NK cells during rejection of kidney transplants. Accumulation of CD56+ NK cells expressing granzyme in kidney biopsies of patients undergoing acute rejection suggests a role of their cytolytic activity in kidney allograft rejection.26 Furthermore, IRI likely induces nonspecific recruitment of NK cells via early graft infiltration.27 A recent study associated the presence of NK cells with the mechanisms of microcirculation injury during antibody-mediated rejection in kidney transplants.9 They proposed that donor-specific antibodies are able to bind to the endothelium and recruit NK cells that produce IFN-γ and trigger antibody-dependent cellular cytotoxicity. The increased expression of NK-cell transcripts, such as CX3CR1, indicated the recruitment of NK cells to allograft endothelium when donor-specific antibodies are present.

pecific antibodies are able to bind to the endothelium and recruit NK cells that produce IFN-γ and trigger antibody-dependent cellular cytotoxicity. The increased expression of NK-cell transcripts, such as CX3CR1, indicated the recruitment of NK cells to allograft endothelium when donor-specific antibodies are present. The NK-cell receptors that are directly involved in the immune responses after kidney transplantation have not been well defined, but various studies suggest that NKG2D could have an important role. An elevated NKG2D mRNA expression was correlated to severity of acute rejection, and NKG2D+ cells were detected in clusters around tubules in biopsies derived from patients diagnosed with acute and chronic rejection.28 NKG2D gene expression was also detected in urinary sediments obtained at 2–3 days before the acute rejection episode; these findings raise the possibility that NKG2D may be able to serve as an additional informative biomarker of transplantation outcome. That is, the NKG2D activating receptor might participate not only in the immune responses that occur initially after transplantation and is promoted by IRI, but it may also contribute to the development of acute rejection. However, further studies are needed to elucidate whether the source of NKG2D receptors is predominantly NK cells or CD8+ T cells.

ceptor might participate not only in the immune responses that occur initially after transplantation and is promoted by IRI, but it may also contribute to the development of acute rejection. However, further studies are needed to elucidate whether the source of NKG2D receptors is predominantly NK cells or CD8+ T cells. NKG2D RECEPTOR, A BRIGDE BETWEEN INNATE AND ADAPTIVE IMMUNITY Maier et al.7 were the first to demonstrate that although NK cells may not be sufficient to directly reject a solid allograft, they can participate in acute rejection by promoting the actions of alloreactive T cells. They demonstrated that CD28−/− mice, which have an impaired T-cell co-stimulation, reject fully MHC-mismatched allogeneic hearts in a manner similar to CD28+/+ recipients. However, CD28−/− mice depleted of NK cells show significant prolongation of allograft survival.

on by promoting the actions of alloreactive T cells. They demonstrated that CD28−/− mice, which have an impaired T-cell co-stimulation, reject fully MHC-mismatched allogeneic hearts in a manner similar to CD28+/+ recipients. However, CD28−/− mice depleted of NK cells show significant prolongation of allograft survival. The bidirectional crosstalk between NK cells and DCs is involved in the maturation of DCs and the activation of an adaptive immune response. DCs secrete cytokines such as interleukin-12 and interleukin-15, which upregulate both the cytotoxic ability and cytokine production (IFN-γ and tumor necrosis factor-α) of NK cells. Concurrently, these proinflammatory cytokines induce DC maturation and the subsequent activation of T cells, promoting a Th1-like alloresponse. NKG2D has a critical role in the NK–DC interaction, as MICA and MICB expression is upregulated on DCs induced to mature by INF-α.29 Furthermore, cytokines such as interleukin-2, interleukin-15, or IFN-α increase the expression of NKG2D, and, consequently, the NKG2D-mediated effector functions. In short, the NK–DC crosstalk through interaction of NKG2D and NKG2D ligands contributes to the feedback regulation of T-cell-mediated immune responses.

α.29 Furthermore, cytokines such as interleukin-2, interleukin-15, or IFN-α increase the expression of NKG2D, and, consequently, the NKG2D-mediated effector functions. In short, the NK–DC crosstalk through interaction of NKG2D and NKG2D ligands contributes to the feedback regulation of T-cell-mediated immune responses. After transplantation, IRI leads to a rapid upregulation of NKG2D ligands that decreases within a few days. However, a second phase of upregulation of NKG2D ligands was only observed in allogenic transplantation,30 that is, Rag−/− mice (deficient in T and B cells but intact NK cells) fail to express Rae-1 in the late phase, suggesting that the innate response is insufficient to maintain the expression of these ligands after the initial injury. These data suggest that initial interactions of NKG2D+ cells with its ligands amplify the adaptive response, enhance the effector functions of CD8+ T cells and NK cells, and increase graft injury. Blockade of NKG2D significantly prolongs graft survival, prevents CD28-independent rejection of cardiac allografts, and inhibits NK effector functions without depleting NK cells or reducing their migration to the graft. Prolonged treatment with anti-NKG2D is critical in maintaining the blockage, as single doses of monoclonal antibody were ineffective in the same cardiac transplantation model.6 Activated NK cells after transplantation may be able to directly or indirectly provide T-cell co-stimulatory signals that can promote allograft rejection.

reatment with anti-NKG2D is critical in maintaining the blockage, as single doses of monoclonal antibody were ineffective in the same cardiac transplantation model.6 Activated NK cells after transplantation may be able to directly or indirectly provide T-cell co-stimulatory signals that can promote allograft rejection. In this way, the increased expression of NKG2D ligands in transplanted allograft might recruit NK cells that directly enhance the NKG2D-mediated cytotoxicity against the graft, function as a bridge between innate and adaptive immunity, and lead to the rejection of the graft. IMMUNOSUPPRESSION AND NKG2D RECEPTOR NK cells also have the ability to contribute to allograft tolerance in patients who have undergone organ transplantation, through elimination or inhibition of donor DCs that subsequently reduce the activation of alloreactive T cells or modulate their response by immunosuppressants. Current immunosuppression in solid organ transplantation involves the use of pharmacological and biological agents that mainly prevent the activation of T cells and development of immune responses against the donor. However, the effects of immunosuppressive agents on NK cells have not been clearly defined.

uppressants. Current immunosuppression in solid organ transplantation involves the use of pharmacological and biological agents that mainly prevent the activation of T cells and development of immune responses against the donor. However, the effects of immunosuppressive agents on NK cells have not been clearly defined. Conflicting results on the influence of calcineurin inhibitors on NK cell phenotype and function have been recently reported. In vitro studies demonstrated that cyclosporine and tacrolimus (FK506) produce dose-dependent inhibition of NK cell degranulation and IFN-γ production.31 Moreover, blood samples from calcineurin inhibitor-treated transplant patients show a reduction of NK cell numbers and impaired effector functions. However, other studies have reported that mycophenolic acid and rapamycin inhibit the acquisition of NKG2A, reduce the expression of NKG2D- and natural cytotoxicity receptor-activating receptors, and lead to the loss of cytotoxicity against target cells,32, 33 but cyclosporine had no effects on the NK receptor repertoire and leaves the cytolytic capacity intact.32, 33 These findings suggest that the distinct mechanisms of action of the immunosuppressive reagents may contribute to their differential effects. As the effects of cyclosporine are known to be reversible, the distinct experimental designs of these studies may also contribute to the disparate results.

capacity intact.32, 33 These findings suggest that the distinct mechanisms of action of the immunosuppressive reagents may contribute to their differential effects. As the effects of cyclosporine are known to be reversible, the distinct experimental designs of these studies may also contribute to the disparate results. Kidney transplant patients receiving induction therapy with polyclonal rabbit anti-thymocyte globulin showed normal levels of NK cells at 1 month after transplantation, but the repertoire of NK cell receptors was modified.34 These cells had an increased expression of the inhibitory receptor NKG2A, and reduced expression of killer Ig-like receptors and NKG2D. The global functions of NK cells were not affected and probably were maintained by a compensatory effect of other receptors. The low levels of NKG2D during the first months after transplantation are of special interest because this receptor is involved in the adaptive immune response against cytomegalovirus (CMV) infection, one of the most common viral complications following solid organ transplantation. NK cells have an important role in the initial stages of viral infections, especially when adaptive immunity is not fully active or is damaged by the use of immunosuppressive therapy. NKG2D interacts with its ligand MICA, which is upregulated in CMV-infected cells, and NKG2D functions as a co-stimulatory molecule in the activation of human CD4+ T lymphocytes against CMV.35 Hadaya et al.36 recently demonstrated an expansion of NKG2D+ NK cell population during acute CMV infection, and these NK cells may have a role similar to that of CD4+ T cells. For resolving CMV infections, it is essential to maintain a proper number and function of NK cells during the early time points after transplantation, and decipher how immunosuppression may affect NK activity to minimize the risks.

acute CMV infection, and these NK cells may have a role similar to that of CD4+ T cells. For resolving CMV infections, it is essential to maintain a proper number and function of NK cells during the early time points after transplantation, and decipher how immunosuppression may affect NK activity to minimize the risks. CONCLUDING REMARKS In summary, the finding that expression of NKG2D ligands is induced by IRI injury during acute and chronic rejection and the discovery of their interactions with their activating receptor NKG2D open a new route of intervention for improving the outcome of solid organ transplantation. NKG2D+ cells were found in kidney biopsies during acute and chronic allograft dysfunction, and blockage of this interaction prolonged the survival in skin and heart-transplanted mice models. Moreover, immunosuppression may influence NK cell functionality and the ability of NK cells to resolve opportunistic viral infections in addition to modulating DC and T-cell responses to the donor. In the future, it is important to determine the in vivo effect of these drugs on the repertoire of inhibitory and activating receptors, such as NKG2D, and their modulation of the effector functions of NK sub-populations. In conclusion, blockade of NKG2D with monoclonal antibodies or soluble NKG2D ligands can contribute to prevent the effector functions of NKG2D+ cells that participate in the innate and adaptive immune response involved in rejection after transplantation. The interaction of NKG2D–NKG2D ligands would be an interesting target for the development of new therapeutic strategies for transplantation.

D ligands can contribute to prevent the effector functions of NKG2D+ cells that participate in the innate and adaptive immune response involved in rejection after transplantation. The interaction of NKG2D–NKG2D ligands would be an interesting target for the development of new therapeutic strategies for transplantation. This work was supported by Spanish grants from the Red de Investigación Renal (REDinREN) and the FIS PI08/0566 from Instituto ‘Carlos III'. TO CITE THIS ARTICLE: Suárez-Álvarez B, Fernández-Sánchez A, López-Vázquez A et al. NKG2D and its ligands: active factors in the outcome of solid organ transplantation? Kidney Int Sup 2011; 1: 52–57. All the authors declared no competing interests. Table 1 Effects of NKG2D and its ligands in solid organ transplantation Transplant Animal model/samples Biological effects Reference NKG2D Heart CD28−/− heart allotransplantation model Increase of NKG2D ligands within the graft amplify the adaptive immune response and increase graft injury Blockage of NKG2D prolongs the cardiac allograft survival 29 Skin SCID mice bearing MHC class II-deficient skin allograft Increased expression of NKG2D ligands on indirectly primed CD4+ T cells in the rejecting grafts Activation of NK cells through NKG2D leads to cytotoxicity against graft cells and rejection Blockage of NKG2D prolongs graft survival but does not induce permanent graft acceptance 8 Kidney Human biopsies and urine Elevated levels of NKG2D mRNA and NKG2D+ cells in urine and kidney biopsies, respectively, diagnosed with acute and chronic rejection 27

KG2D leads to cytotoxicity against graft cells and rejection Blockage of NKG2D prolongs graft survival but does not induce permanent graft acceptance 8 Kidney Human biopsies and urine Elevated levels of NKG2D mRNA and NKG2D+ cells in urine and kidney biopsies, respectively, diagnosed with acute and chronic rejection 27 NKG2D ligands Kidney TEC IRI induces the expression of NKG2D ligand, Rae-1, in TEC NK cell engagement through NKG2D allows NK cell-induced apoptosis 17 Kidney Renal IRI mice model IRI increases the expression of Rae-1 and H60 and activates NK and CD8+ T cells 18 Heart C57BL/6 heart allotransplant model Simultaneous increase of Rae-1, H60, and NKG2D during acute cardiac allograft rejection 19 Kidney Human renal proximal tubular epithelial cells (HK2 cell line) Hypoxia during IRI increases MICA expression through a HIF-1 pathway Enhance cytotoxicity and IFN-γ secretion by NK cells, leading to graft injury 20 Kidney and pancreas Biopsies Increase of MICA/B in epithelial cells of kidney and pancreas allografts during acute and chronic rejection 16 Kidney Biopsies Upregulation of MICB molecules following renal transplantation associated with evidence of cellular stress 15 Heart Biopsies Increase of MICA in endomyocardial biopsies with histological evidence of acute rejection 14 Abbreviations: HIF, hypoxia-inducible factor; IFN, interferon; IRI: ischemia/reperfusion injury; MHC, major histocompatibility complex; MICA, MHC class I chain-related A; MICB, MHC class I chain-related B; NK, natural killer; SCID, severe combined immunodeficiecy disease; TEC, tubular epithelial cells.

The current immunosuppressive therapy in solid organ transplantation uses a combination of several drugs that function on multiple pathways of the immune response. These drugs are classified by their mechanism of action, such as calcineurin inhibitors (cyclosporine A, tacrolimus (Tac)), inhibitors of purine synthesis (mycophenolate mofetil), and mammalian target of rapamycin inhibitors (sirolimus, everolimus). These drugs are frequently combined with glucocorticoids (methylprednisolone, prednisone), monoclonal (muromonab, basiliximab, daclizumab), and polyclonal (anti-thymocyte globulin) antibodies. Tacrolimus is an immunosuppressive drug used to prevent solid organ rejection, and also to treat autoimmune diseases. Tac, similar to cyclosporine, is a calcineurin inhibitor and suppresses the activation, proliferation, and differentiation of T cells. Calcineurin inhibitors prevents the transcription of several cytokine genes involved in immune responses.1, 2, 3 Tac has gradually replaced cyclosporine as the first-choice immunosuppressive drug, mainly because of its higher immunosuppressive activity and fewer adverse effects. However, Tac has also been associated with a higher risk of developing dyslipidemia, hypercholesterolemia, hypertension, post-transplant (PT) nephrotoxicity, and new-onset diabetes after transplantation.4, 5, 6, 7

immunosuppressive drug, mainly because of its higher immunosuppressive activity and fewer adverse effects. However, Tac has also been associated with a higher risk of developing dyslipidemia, hypercholesterolemia, hypertension, post-transplant (PT) nephrotoxicity, and new-onset diabetes after transplantation.4, 5, 6, 7 Tac shows an interindividual pharmacokinetic variability that affects the dose required to reach the target concentration in blood.8, 9 The current therapeutic approach is based on an initial daily dose of 0.2 mg/kg (given in two equal 12-h doses in the case of Prograf (Astellas Pharma, Deerfield, IL)). The blood level is measured 12 h after dose (immediately before receiving the next dose) and is known as the trough or C0 level. The clinician uses the C0 level in each individual patient to decide whether to maintain, increase, or reduce the dose.9, 10 The target C0, which is 10–15 ng/ml in the period 0 to 3 months PT, and 5–10 ng/ml thereafter, is considered as the optimal concentration to avoid rejection (a concentration too low) and toxicity (a concentration too high).10

l in each individual patient to decide whether to maintain, increase, or reduce the dose.9, 10 The target C0, which is 10–15 ng/ml in the period 0 to 3 months PT, and 5–10 ng/ml thereafter, is considered as the optimal concentration to avoid rejection (a concentration too low) and toxicity (a concentration too high).10 Mainly, Tac is metabolized by two enzymes of the cytochrome P450 family, CYP3A5 and CYP3A4, whereas other P450 isoforms are much less effective.11, 12, 13 Most of the Tac biotransformation occurs in the liver, and to a lesser extent in the small intestine. In vitro studies with human liver microsomes showed that CYP3A5 had high Tac catalytic efficiency, and its contribution was stronger in microsomes from individuals with low CYP3A4 concentrations.11 Several factors influence the blood concentration of Tac. Some factors are under the patients' control, such as diet or the co-administration of drugs that share the same metabolic pathways with Tac (i.e., fluconazole and ketoconazole).14, 15 However, some of the major determinants of Tac bioavailability reside in genes implicated in its absorption and metabolization. Several studies have reported that polymorphisms at the ABCB1/MDR-1, CYP3A4, and CYP3A5 affect Tac dose requirements, as discussed below.

s with Tac (i.e., fluconazole and ketoconazole).14, 15 However, some of the major determinants of Tac bioavailability reside in genes implicated in its absorption and metabolization. Several studies have reported that polymorphisms at the ABCB1/MDR-1, CYP3A4, and CYP3A5 affect Tac dose requirements, as discussed below. CYP3A5 IN Tac DOSE The effect of the CYP3A5 genotype on Tac bioavailability has been demonstrated by several laboratories.16, 17, 18, 19, 20, 21, 22, 23, 24 The main determinant of this pharmacogenetic effect is a single-nucleotide polymorphism (SNP) in intron 3 of CYP3A5 (6986 A>G; SNP rs776746), also known as CYP3A5*3 (for a complete list of the CYP variants, see the home page of the Human Cytochrome P450 Allele Nomenclature, http://www.cypalleles.ki.se).25, 26 Most studies examined the effect of CYP3A5*3 on the twice-daily dose formulation of Tac (Prograf) at several PT times. The mean dose-adjusted blood Tac concentration was significantly higher among CYP3A5*3 homozygotes than that of carriers of the wild-type allele (CYP3A5*1). The CYP3A5*3 allele affects splicing of the pre-mRNA and greatly reduces P450-3A5 activity.11, 12 The poor metabolizing phenotype of CYP3A5*3/*3 homozygotes explains why they would require a lower Tac dose to reach the blood target concentration compared with carriers of the CYP3A5*1 allele.

ild-type allele (CYP3A5*1). The CYP3A5*3 allele affects splicing of the pre-mRNA and greatly reduces P450-3A5 activity.11, 12 The poor metabolizing phenotype of CYP3A5*3/*3 homozygotes explains why they would require a lower Tac dose to reach the blood target concentration compared with carriers of the CYP3A5*1 allele. We recently reported the results of a multicenter study of Tac-pharmacogenetics in Spanish patients who received a first cadaveric kidney graft (the REDinREN pharmacogenetic study).24 A total of 400 patients were treated with a standard triple immunosuppressive therapy with Tac (Prograf), prednisone, and mycophenolate mofetil. The initial oral dose of Tac was 0.2 mg/kg per day and was adjusted to reach a C0 of 10–15 ng/ml in the period from 0 to 3 months PT, and 5–10 ng/ml thereafter. Tac was measured in human whole blood with an automated chemiluminescent immunoassay and the Arquitect Tacrolimus assay (Abbott Laboratories, Chicago, IL).27 Compared with CYP3A5*1 carriers (n=80), patients who were CYP3A5*3 homozygotes (n=320) received lower median Tac (mg/kg per day) at 1 week (0.14 vs 0.12), at 6 months (0.10 vs 0.06), and at 1 year (0.08 vs 0.05) PT. These values were similar to those reported by others.

limus assay (Abbott Laboratories, Chicago, IL).27 Compared with CYP3A5*1 carriers (n=80), patients who were CYP3A5*3 homozygotes (n=320) received lower median Tac (mg/kg per day) at 1 week (0.14 vs 0.12), at 6 months (0.10 vs 0.06), and at 1 year (0.08 vs 0.05) PT. These values were similar to those reported by others. Assessing the impact of the CYP3A5*3 allele on Tac pharmacogenetics needs to consider the genotype frequencies among populations of various ethnic origins. Approximately 80% of Caucasians, but only 30% of African Americans, are CYP3A5*3 homozygotes (non-expressors).28 These differences in genotype frequencies could explain part of the observed variability in Tac dose requirements among different populations.29 CYP3A4 IN Tac DOSE A number of CYP3A4 SNPs have been identified. Most of the interindividual variability in CYP3A4 activity may be due to differences in transcript levels, and results from nucleotide changes in the promoter region.30 In particular, the CYP3A4*1B (−392A>G; SNP rs2740574) is a common allele located in the promoter region, is associated with differences in transcriptional activity, and correlates with increased hepatic expression of CYP3A4.31, 32

ences in transcript levels, and results from nucleotide changes in the promoter region.30 In particular, the CYP3A4*1B (−392A>G; SNP rs2740574) is a common allele located in the promoter region, is associated with differences in transcriptional activity, and correlates with increased hepatic expression of CYP3A4.31, 32 Its expression varies in liver and other tissues, and its inherent concentration has a role on Tac metabolism in liver microsomes, particularly in microsomes from individuals who did not express CYP3A5.11 However, none of CYP3A4 SNPs has shown a clear influence on Tac pharmacokinetics.33 In our study, carriers of the −392 A>G variant had significantly higher Tac doses.24 A higher gene expression linked to this allele (compared with the wild type, CYP3A4*1) could explain the lower dose requirements among CYP3A4*1 homozygotes. Although our work confirmed the results from other studies,24 the significance of our study was limited by the low frequency of the CYP3A4*1B allele (only 6% of the patients were CYP3A4*1B carriers, and no patient was homozygous for this allele). However, CYP3A4 and CYP3A5 are closely linked, and the effect of the CYP3A4 polymorphisms on Tac pharmacokinetics could be due to linkage disequilibrium with CYP3A5*3. A way to solve this dilemma is to analyze the effect of CYP3A4 variation on patients with different CYP3A5 genotypes.

t was homozygous for this allele). However, CYP3A4 and CYP3A5 are closely linked, and the effect of the CYP3A4 polymorphisms on Tac pharmacokinetics could be due to linkage disequilibrium with CYP3A5*3. A way to solve this dilemma is to analyze the effect of CYP3A4 variation on patients with different CYP3A5 genotypes. CYP3A4*1B carriers had significantly higher median Tac C0 values at 3 and 1 year PT, but not at 7 days PT than CYP3A4*1 homozygotes did. The same modifying effect of the CYP3A4 genotype was observed among CYP3A5*1 carriers.24 In contrast, Kuypers et al.23 reported similar Tac C0 values for the two CYP3A5*1 groups. However, no patient in their study was a CYP3A5*3 homozygote+CYP4A4*1B carrier. Because the conclusions of these studies are hampered by the low number of patients who were CYP3A5*1B carriers, additional studies with larger cohorts of patients are necessary to determine the value of genotyping CYP3A4 in addition to CYP3A5.23, 24

o patient in their study was a CYP3A5*3 homozygote+CYP4A4*1B carrier. Because the conclusions of these studies are hampered by the low number of patients who were CYP3A5*1B carriers, additional studies with larger cohorts of patients are necessary to determine the value of genotyping CYP3A4 in addition to CYP3A5.23, 24 ABCB1 POLYMORPHISMS IN Tac DOSE The ABCB1 gene (also known as the multidrug resistance-1 gene, MDR-1) encodes the P-glycoprotein (P-gp), which is a pump that drives the efflux of many drugs in the intestinal wall and other cell types. The amount of the drug that reaches the blood stream could depend on the P-gp activity, and ABCB1 polymorphisms linked to differences in P-gp expression/function could have an important role on dose requirements.33, 34, 35, 36, 37, 38 The role of P-gp expression on Tac bioavailability was reported by Masuda et al.,39 who found a strong correlation between ABCB1 mRNA levels in intestinal biopsies and the dose-adjusted Tac concentrations. The effect of several ABCB1 SNPs on Tac pharmacokinetics has been investigated, with conflicting results.17, 18, 19, 20, 21, 22, 23 We did not find a significant effect of the common c.3435 C/T polymorphism (exon 26 SNP rs1045642) on Tac bioavailability.24 In addition, this SNP did not modify the effect of the CYP3A5 genotype.

ct of several ABCB1 SNPs on Tac pharmacokinetics has been investigated, with conflicting results.17, 18, 19, 20, 21, 22, 23 We did not find a significant effect of the common c.3435 C/T polymorphism (exon 26 SNP rs1045642) on Tac bioavailability.24 In addition, this SNP did not modify the effect of the CYP3A5 genotype. However, the donor ABCB1 3435TT genotype was significantly associated with susceptibility to chronic allograft damage.40 The 3435 T homozygosity likely increased the renal expression of P-gp, which resulted in intrarenal accumulation of Tac.40 If this result is confirmed by others, the donor ABCB1 genotype could be a valuable tool to predict Tac-induced nephrotoxicity.

icantly associated with susceptibility to chronic allograft damage.40 The 3435 T homozygosity likely increased the renal expression of P-gp, which resulted in intrarenal accumulation of Tac.40 If this result is confirmed by others, the donor ABCB1 genotype could be a valuable tool to predict Tac-induced nephrotoxicity. OTHER GENE VARIANTS IN Tac DOSE Although the CYP3A5*3 is the main genetic determinant of Tac pharmacokinetics, this SNP explains ∼50% of the total variability.20 Thus, other genetic variants could affect Tac metabolism and dose requirements. The effect of other nucleotide variants could also explain the variability between individuals with the same CYP3A5 genotype. For instance, 41% of our CYP3A5*3/*3 and 26% of the CYP3A5*1 carriers had C0 values in the target range (10–15 ng/μl) at 1 week PT. Although these frequencies diminished with time, 10% of the patients remained out of the target range (5–10 ng/μl) after 6 months PT. Data regarding the possible role of several polymorphisms on Tac pharmacogenetics have been recently presented.41 We assessed the effect of 96 DNA variants in 16 metabolizing enzymes on Tac dose requirements.24 In addition to CYP3A4, CYP3A5, and ABCB1, several P450, glutathione and N-acetyl transferases, and thiopurine S-methyltransferase gene variants were studied. We did not detect any significant effects of these SNPs on Tac dose requirements. Moreover, none of these polymorphisms had a significant effect after correcting for the CYP3A5 genotype.

YP3A5, and ABCB1, several P450, glutathione and N-acetyl transferases, and thiopurine S-methyltransferase gene variants were studied. We did not detect any significant effects of these SNPs on Tac dose requirements. Moreover, none of these polymorphisms had a significant effect after correcting for the CYP3A5 genotype. The CYP3A4 polymorphisms may also affect Tac pharmacokinetics. As discussed above, our data suggested an effect of the CYP3A4*1B allele on Tac metabolism. At 1 year PT, the patients who were CYP3A5*3*3+CYP3A4*1B carriers had Tac C0 values in the target range, whereas 6% of the CYP3A5*3/*3+CYP3A4*1/*1 remained out of the target range.24 Most of the CYP3A4 variants found in the coding region have an allele frequency <1%. An exception was CYP3A4*2, a missense SNP (Ser222Pro) with a frequency of 5% among the Caucasians. This allele was linked to a lower clearance of the CYP3A4 substrate nifedipine, and carriers of this allele can thus be classified as ‘slow metabolizers'.42, 43 The effect of this variant on Tac bioavailability has not been established. The sequencing of CYP3A4 may be very informative in patients whose C0 values cannot be explained by the CYP3A5 genotype, and the sequencing can help determine the overall contribution of CYP3A4 to Tac dose requirements. The same argument applies for the sequencing of CYP3A5 in those patients who are CYP3A5*1 carriers with C0 values that were above the target range. These patients could harbor one of several CYP3A5 variants that are linked to a reduced catalytic activity and a slow to null metabolizing phenotype.

Tac dose requirements. The same argument applies for the sequencing of CYP3A5 in those patients who are CYP3A5*1 carriers with C0 values that were above the target range. These patients could harbor one of several CYP3A5 variants that are linked to a reduced catalytic activity and a slow to null metabolizing phenotype. READY FOR CLINICAL TRANSLATION? The ultimate goal of the pharmacogenetics of Tac is to provide a tool to predict the dose for each patient before transplantation, and prevent the effects induced by an over/underdose. Haufroid et al.20 proposed a loading dose of 0.075 mg/kg and 0.150 mg/kg body weight twice a day among CYP3A5 non-expressors and expressors, respectively. These values were derived from a study of 19 volunteers (nine expressors, 10 non-expressors) who received a standard dose (0.1 mg/kg body weight twice a day). This and other studies paved the way toward clinical trials that evaluate the benefit of dosing according to the genotype.44

essors and expressors, respectively. These values were derived from a study of 19 volunteers (nine expressors, 10 non-expressors) who received a standard dose (0.1 mg/kg body weight twice a day). This and other studies paved the way toward clinical trials that evaluate the benefit of dosing according to the genotype.44 The first prospective study has been recently reported by Thervet et al.45 A group of 280 patients received a Tac dose, either according to the CYP3A5 genotype (the adapted-dose group; n=116) or to the standard regimen (the control group; n=120). All patients received induction therapy with mycophenolate mofetil (Cell-Cept; Roche Farma, Basel, Switzerland), corticosteroids, and either basiliximab (Simulect; Novartis, Basel, Switzerland) or intravenous anti-thymocyte globulin (Thymoglobulin; Genzyme, Cambridge, MA). No drugs known to interact with CYP3A5 were administered. A limitation of this study was that Tac administration began on day 7 PT (a time required to determine the genotype), and the effect of the pharmacogenetic adaptation was thus not evaluated in patients treated with Tac from day 0. This delay in Tac dosing could affect the main clinical and analytical findings.

istered. A limitation of this study was that Tac administration began on day 7 PT (a time required to determine the genotype), and the effect of the pharmacogenetic adaptation was thus not evaluated in patients treated with Tac from day 0. This delay in Tac dosing could affect the main clinical and analytical findings. At day 7 PT, the patients in the adapted-dose group who were CYP3A5 expressors (n=26) received an initial Prograf dose of 0.30 mg/kg per day, compared with 0.15 mg/kg per day among the CYP3A5 non-expressors (n=90). The control group was treated with an initial dose of 0.20 mg/kg per day. The first measurement of the Tac C0 concentration was recorded after the sixth Tac dose (on day 10 PT). Patients in the adapted-dose group had Tac C0 values in the target range (10–15 ng/l) more frequently than the control group (43.3 vs 29.2% P=0.003). Moreover, the adapted-dose group required 3–8 days (median 6 days) to reach the target range compared with 3–25 days (median 7 days) in the control group (P=0.001). The total number of dose modifications was also lower in the adapted-dose group (281 vs 420; P=0.004). This study provided the first evidence that the genotyping-based dose adaptation reduces the time to reach the blood target concentration, but was this the only benefit?

(median 7 days) in the control group (P=0.001). The total number of dose modifications was also lower in the adapted-dose group (281 vs 420; P=0.004). This study provided the first evidence that the genotyping-based dose adaptation reduces the time to reach the blood target concentration, but was this the only benefit? Tac blood levels are routinely monitored several times in the first few weeks PT to adjust the dose to the target concentration, which is achieved within the first 2 weeks in ∼90% of the patients. Therefore, it should not be surprising that clinicians may consider that the adapted-dose method requires too much effort if the only benefit is to more rapidly (few days) determine the right dose.46 The significant delay in achieving the target blood concentration among CYP3A5 expressors has been linked to higher risk for early acute rejection.47, 48, 49 However, some authors failed to confirm the association between the CYP3A5*1 allele and acute rejection.50, 51 Thervet et al.45 did not find significant differences in the incidence of delayed graft function, the number of PT dialysis sessions per patient, or the number of acute rejection episodes between the adapted and control groups. Their findings suggested that the Tac dose according to the CYP3A5 genotype might contribute minimally to the reduction of early acute rejection. However, the fact that their patients received biological induction therapy coupled with high dose of MMF during the first week could significantly reduce the incidence of acute rejection, affecting the results of the study. It is thus important to replicate this study on patients treated with an adapted dose from day 0, and also to determine whether the pharmacogenetic approach could help reduce the necessity for induction therapy and co-immunosuppressors.

cantly reduce the incidence of acute rejection, affecting the results of the study. It is thus important to replicate this study on patients treated with an adapted dose from day 0, and also to determine whether the pharmacogenetic approach could help reduce the necessity for induction therapy and co-immunosuppressors. Finally, recent studies have also demonstrated a significant effect of the CYP3A5 genotype on the pharmacokinetic and dose requirements for the once-daily Tac formulation (Advagraf, Astellas Pharma, Staines, UK).52, 53 These results suggested that the pharmacogenetic approach for the twice-daily formulation could be also applied to this once-daily formulation, which may improve patient compliance. In conclusion, genotyping of CYP3A5 may be useful to predict the Tac dose immediately after transplantation, and reduce the time required to reach the target concentration. However, the effect of the genotype-adapted dose on acute rejection and other clinical outcomes seems less clear. Therefore, trials to determine whether this pharmacogenetic approach could reduce the incidence of acute rejection and delayed graft function are necessary, particularly in patients without biological induction therapy. This work was supported by the Instituto de Salud Carlos III-Fondos Feder European Union, grants FIS-08/0566, and by ETS-08/90008. TO CITE THIS ARTICLE: Coto E, Tavira B, Suárez-Álvarez B et al. Pharmacogenetics of tacrolimus: ready for clinical translation? Kidney Int Sup 2011; 1: 58–62. All the authors declared no competing interests.

Biological discoveries and technological advancement allowing synchronous, detailed analyses of metabolic systems as well as sophisticated informatics will soon lead to a profound restructuring of medical specialties. New inter-specialty areas are generated in the modern magmatic scenario, areas where the clinical investigator and the basic scientist will effectively join forces to create a novel integrative approach to clinical research. Nephrology and Cardiology are sister specialties. The several existing international societies focusing on hypertension represent concrete proof of the efforts by internists, nephrologists and cardiologists to build a common approach to a condition which is of core interest to these medical specialties. Chronic kidney disease (CKD) is now an established public health priority and, mainly because of the high risk for cardiovascular complications secondary to renal function loss, it represents one of the most challenging problems of modern medicine. The bidirectional link that associates renal and cardiovascular diseases and the high risk of the death signaled by their coexistence is at the basis of a new discipline aiming at making the borders between nephrology and cardiovascular medicine even more permeable than before. Cardiovascular and Renal Medicine is a new, shared territory for clinicians and investigators with a prominent interest on the kidney-cardiovascular system interface.

xistence is at the basis of a new discipline aiming at making the borders between nephrology and cardiovascular medicine even more permeable than before. Cardiovascular and Renal Medicine is a new, shared territory for clinicians and investigators with a prominent interest on the kidney-cardiovascular system interface. To respond to the growing interest in this clinical research area, the European Renal Association–European Dialysis and Transplant Association (ERA–EDTA) has recently created an EUropean working group (WG) focusing on REnal and CArdiovascular Medicine (EURECA-m). The scope of EURECA-m is that of promoting collaborations among European centres pursuing research in the overlapping area of cardiovascular and renal medicine. The specific goals of the WG were delineated in a paper (NDT Plus 2010; 2: 522–525) deposited in the WEB site of the same WG (http://www.era-edta.org/eureca-m_publications.htm).

A-m is that of promoting collaborations among European centres pursuing research in the overlapping area of cardiovascular and renal medicine. The specific goals of the WG were delineated in a paper (NDT Plus 2010; 2: 522–525) deposited in the WEB site of the same WG (http://www.era-edta.org/eureca-m_publications.htm). The clinical research questions to be faced are numerous and the WG agreed that at this stage it is fundamental to identify research priorities. For this reason members of the Board of EURECA-m met twice in 2010 to delineate concrete plans for research initiatives and for scoping questions to be shared with EURECA-m members and interested investigators and clinicians. This Supplement of Kidney International is the result of the joint effort of the EURECA-m Board. We hope that the doubts and questions listed in these manuscripts may serve as a stimulus for channeling research into priority themes and as a starting point to frankly debate the many intriguing problems posed by this fascinating research area.

The heart and kidney are frequently affected by similar, highly prevalent risk factors such as diabetes and hypertension, and both are profoundly affected by the aging process. Atherosclerosis is a generalized arterial disease of the arterial intima, characterized by the presence of plaques and occlusive arterial lesions. Atherosclerosis extends from the coronaries and the thoracic aorta to the renal circulation. Nephrosclerosis, that is, the renal expression of intimal disease (either alone or associated with occlusive renal artery disease), is the most frequent renal disease underlying the high prevalence of chronic kidney disease (CKD) in the general population. Even though most patients with CKD die because of atherosclerotic complications before they reach end-stage renal disease (ESRD), the dialysis population is composed mainly of elderly patients with a high burden of cardiovascular complications. Kidney failure accelerates the progression of atherosclerosis and modifies the morphology of atherosclerosis lesions by increasing the propensity to calcification.1

ey reach end-stage renal disease (ESRD), the dialysis population is composed mainly of elderly patients with a high burden of cardiovascular complications. Kidney failure accelerates the progression of atherosclerosis and modifies the morphology of atherosclerosis lesions by increasing the propensity to calcification.1 Aging and age-associated arterial changes underlie fundamentally different alterations from atherosclerosis, and alterations attributable to the aging process may have a relevant effect on cardiac and renal disease. Aging per se is associated with a parallel decrease in renal function and is accompanied by relevant changes in the properties of large- and medium-sized arteries. The main structural changes attributable to aging include arterial dilatation and tortuousness, wall hypertrophy, and increased collagen-to-elastin ratio with fragmentation and calcification of elastic fibers. The functional consequence of these structural alterations is hardening/sclerosis of vessel walls (arteriosclerosis) and loss of distensibility, that is, increased stiffness.2 Arterial aging mainly involves the aorta and major central arteries, whereas peripheral muscular conduit arteries undergo only modest changes.3 In young and middle-aged subjects the aorta is more distensible than peripheral arteries. Physiologically, the higher distensibility of the aorta coupled with a progressively lower distensibility in peripheral vessels creates a ‘stiffness gradient' that works as a ‘hydraulic filter' and acts to buffer pressure pulsations and their transmission to microcirculation and capillary network principally in the main parenchymal organs such as the kidney and the brain.4 In brief, during systole, the stroke volume interacts with aortic characteristics to produce a pulsatile pressure wave (forward pressure) that normally travels from the aorta toward peripheral arteries at a pulse wave velocity (PWV) that accelerates centrifugally. PWV is low in the distensible aorta and accelerates in progressively stiffer peripheral arteries. During heart contraction, only a part of the stroke volume is forwarded directly to the peripheral tissues. The pressure generated by the left ventricular (LV) systole distends the elastic elements of the arterial walls and is transformed into an elastic force, while a part of the stroke volume is accommodated in the distended aorta.

ntraction, only a part of the stroke volume is forwarded directly to the peripheral tissues. The pressure generated by the left ventricular (LV) systole distends the elastic elements of the arterial walls and is transformed into an elastic force, while a part of the stroke volume is accommodated in the distended aorta. During diastole, this elastic force recoils the aorta and squeezes the blood forward into the peripheral tissues, thereby ensuring a continuous flow.5, 6 For this function to be efficient, the energy necessary for arterial distension and recoil should be as low as possible; that is, for a given stroke volume, the pulse pressure should be as low as possible. In other words, the more distensible the arterial wall (that is, the lower the stiffness), the smoother the provision of proper flow to peripheral tissues. As the forward pressure wave travels toward the less distensible and smaller peripheral arteries, impedance mismatches generate a reflected wave, that is, a wave that travels backward toward the aorta (reflected wave). In physiological conditions, the reflected wave returns normally to the aorta in late systole and early diastole (distensible aorta with low PWV), producing a favorable increase in diastolic pressure and in coronary perfusion and effectively limiting the transmission of high pulsatile energy to microcirculation.5, 6, 7

In physiological conditions, the reflected wave returns normally to the aorta in late systole and early diastole (distensible aorta with low PWV), producing a favorable increase in diastolic pressure and in coronary perfusion and effectively limiting the transmission of high pulsatile energy to microcirculation.5, 6, 7 Aortic stiffening manifests as high systolic and pulse pressures with increased cardiac afterload and arterial circumferential stress, all factors promoting LV hypertrophy (LVH), which may evolve toward heart failure. Arterial rigidity increases the PWV and determines an early return of the reflected wave. This wave reaches the central circulation during early rather than late systole and late diastole, thereby increasing systolic and pulse pressure and decreasing diastolic pressure.5, 7 Stiff aorta cannot be stretched, and therefore the stroke volume flows through the arterial system toward peripheral tissues principally during systole, decreasing capillary transit time and metabolic exchanges. This is an energy-demanding hemodynamic pattern implying a high cardiac energy expenditure and a high oxygen consumption in the myocardium, thereby favoring cardiac ischemia. Importantly, in such a situation, the lack of proper aortic buffering also determines a direct transmission of pulsatile energy into the peripheral microvessels and microcirculatory network.4, 8 Arterioles are the ‘last barrier' protecting the capillary microcirculation from high pulsatile energy.

ing cardiac ischemia. Importantly, in such a situation, the lack of proper aortic buffering also determines a direct transmission of pulsatile energy into the peripheral microvessels and microcirculatory network.4, 8 Arterioles are the ‘last barrier' protecting the capillary microcirculation from high pulsatile energy. In the presence of central stiffness and the associated loss of the arterial system distensibility gradient, organs characterized by high blood flow and low resistance, such as the kidney or the brain, are particularly exposed to the damaging effect of high pulsatile pressure.8 Strong associations have been described between aortic stiffness (aortic PWV) and LVH and LV dysfunction, as well as between the same parameters and indicators of renal dysfunction (glomerular filtration rate and microalbuminuria).9 In addition, aortic stiffness and pulse pressure have been associated with cognitive impairment and dementia.10 Age-associated aortic stiffening notoriously occurs at a much accelerated rate in ESRD patients.11 In this population, arterial rigidity is typically associated with calcifications, and PWV in ESRD reaches the highest level observed in human diseases.12 PWV represents one of the strongest markers for the risk of death and cardiovascular outcomes in dialysis patients.11, 12 Similar to the general population, arterial stiffening in ESRD is mainly confined to the aorta.3, 4, 11, 13

th calcifications, and PWV in ESRD reaches the highest level observed in human diseases.12 PWV represents one of the strongest markers for the risk of death and cardiovascular outcomes in dialysis patients.11, 12 Similar to the general population, arterial stiffening in ESRD is mainly confined to the aorta.3, 4, 11, 13 The rational schema laid down above provides a general framework for interpretation of the role of age-associated arterial changes in cardiovascular and renal diseases and for framing urgent research questions in this area. Indeed, the causes and consequences of age-dependent arterial abnormalities are not well understood and should be the focus of interest in future studies. In this regard, many critical questions still remain to be properly addressed (Figure 1). Aortic stiffening could be analyzed by several techniques, including aortic PWV.14 Information on time-related changes in aortic stiffness as related to hemodynamic changes (24-h ambulatory blood pressure monitoring, extracellular volume, and indicators of salt and volume excess) and renal function is still scarce, which is why it is critical to probe the general relevance of the stiffening process in cardiovascular and renal health.

s in aortic stiffness as related to hemodynamic changes (24-h ambulatory blood pressure monitoring, extracellular volume, and indicators of salt and volume excess) and renal function is still scarce, which is why it is critical to probe the general relevance of the stiffening process in cardiovascular and renal health. Longitudinal analyses of the relationship between the typical biochemical alterations of CKD and ESRD, such as high levels of oxidative stress, inflammation, endothelial dysfunction (see previous section), and markers of mineral disorders, PWV, and clinical outcomes, are needed to identify causal risk factors for arterial stiffening and cardiovascular risk excess in CKD and ESRD. Equally important is investigating the relationship between the biochemical alterations of CKD and PWV in peripheral arteries (carotido-radial or femoro-tibial) in order to understand the role of dissipation of the distensibility gradient in target organ damage (that is, kidney, brain, and heart disease). Studies of the microvascular structure and reactivity in the kidney, heart, and brain are the ‘parent pauvre' in CKD and ESRD. Techniques exist to study the microvascular territory at the anatomic level (cutaneous, nail, retinal capillary density) or at the functional level (in terms of postischemic forearm vasodilation and flow), but the best way to measure microvascular reactivity within the kidney circulation is an open question (see previous section).

xist to study the microvascular territory at the anatomic level (cutaneous, nail, retinal capillary density) or at the functional level (in terms of postischemic forearm vasodilation and flow), but the best way to measure microvascular reactivity within the kidney circulation is an open question (see previous section). Even though progress has been made in this area, we still ignore whether measures of arterial stiffness such as PWV are useful in clinical practice to monitor therapies that broadly interfere with the cardiovascular system and arterial function, such as drugs antagonizing the effects of the renin–angiotensin–aldosterone system or calcium-channel blockers or drugs impinging upon alterations in mineral metabolism. Vascular calcifications are a prominent feature of arterial disease in CKD and ESRD;15, 16 however, it should be emphasized that it is largely undefined whether calcifications represent a valid surrogate end point that can be applied in intervention studies in these patients. In other words, we ignore to check whether interventions that reduce vascular calcifications unequivocally translate into better clinical outcomes, and this is true both for interventions based on drugs impinging upon dyslipidemia and for interventions aimed at correcting alterations in mineral metabolism in CKD (phosphate binders, vitamin D, calcimimetics, vitamin K). Finally, arterial changes are tightly associated with cardiac morphological (LVH) and functional changes (LV dysfunction)17, 18, 19 in cross-sectional studies; however, the longitudinal relationship between arterial and cardiac changes is not well documented in CKD/ESRD and the role of arterial stiffening on heart disease in these patients remains to be defined, in parallel with the evaluation of senescence rates in these CKD/ESRD populations.20

19 in cross-sectional studies; however, the longitudinal relationship between arterial and cardiac changes is not well documented in CKD/ESRD and the role of arterial stiffening on heart disease in these patients remains to be defined, in parallel with the evaluation of senescence rates in these CKD/ESRD populations.20 TO CITE THIS ARTICLE: London G, Covic A, Goldsmith D et al. Arterial aging and arterial disease: interplay between central hemodynamics, cardiac work, and organ flow—implications for CKD and cardiovascular disease. Kidney Int Sup 2011; 1: 10–12.

19 in cross-sectional studies; however, the longitudinal relationship between arterial and cardiac changes is not well documented in CKD/ESRD and the role of arterial stiffening on heart disease in these patients remains to be defined, in parallel with the evaluation of senescence rates in these CKD/ESRD populations.20 TO CITE THIS ARTICLE: London G, Covic A, Goldsmith D et al. Arterial aging and arterial disease: interplay between central hemodynamics, cardiac work, and organ flow—implications for CKD and cardiovascular disease. Kidney Int Sup 2011; 1: 10–12. AC has received consulting fees from Abbott Laboratories and received lecture fees from F. Hoffmann-La Roche, Amgen, and Fresenius Medical Care Holdings. AM-C has received consulting fees from Abbott Laboratories, Roche Spain, and Abbott Spain. AO has received grant support from the Spanish Government. AW has received lecture fees from Amgen, F. Hoffmann-La Roche, and Janssen-Cileg. AW has also received grant support from Astellas Pharma. DF has received funding from the EU. DG has received consulting fees and lecture fees from Shire, Genzyme, Novartis AG, Sandoz, Pfizer, and Fresenius Medical Care Holdings. FWD has received funding from Amgen and Baxter. GL has received consulting fees from Amgen and Sandoz. GL has also received lecture fees from Amgen, Sandoz, Genzyme, and Shire. PJB has received consulting fees from Medtronic and has received grant support from Ardian and Novartis AG. RA has received consulting fees from Amgen, Abbott Laboratories, Merck, Affymax, Takeda Pharmaceutical Company, Daiichi Sankyo, Celgene, Watson Pharmaceuticals, and Rockwell Medical. RA has also received lecture fees from Abbott Laboratories, Merck, and Medscape. ZM has received lecture fees from Amgen, Shire, Genzyme, FMC, and Merck Sharp & Dohme. ZM has received grant support from Baxter, Amgen, FMC, Shire, and Genzyme. The remaining authors declared no competing interests.

and Rockwell Medical. RA has also received lecture fees from Abbott Laboratories, Merck, and Medscape. ZM has received lecture fees from Amgen, Shire, Genzyme, FMC, and Merck Sharp & Dohme. ZM has received grant support from Baxter, Amgen, FMC, Shire, and Genzyme. The remaining authors declared no competing interests. Figure 1 Remaining questions to be addressed.

Hypertension is both a cause and effect of chronic kidney disease (CKD). There is little doubt about this bidirectional and causal relationship, the evidence for which comes from epidemiological, clinical, and research models. Although the modern techniques to measure blood pressure (BP) were described over a 100 years ago by Riva Rocci and Nikolai Korotkoff, how to best assess hypertension and the level to which it should be lowered among those with CKD is mired in controversy. This controversy is especially vexing among patients with end-stage renal disease on chronic hemodialysis. In these patients, large volume shifts from before to after dialysis cause wide BP variations. These variations make the optimal timing and definition of hypertension problematic. The gaps in our knowledge that need to be addressed in patients with CKD and those on dialysis are discussed further.

chronic hemodialysis. In these patients, large volume shifts from before to after dialysis cause wide BP variations. These variations make the optimal timing and definition of hypertension problematic. The gaps in our knowledge that need to be addressed in patients with CKD and those on dialysis are discussed further. CKD PATIENTS NOT ON DIALYSIS Joint National Commission-6 recommended lowering BP to <125/75 mm Hg among those with >1 g proteinuria. However, these recommendations were removed as they were based on post hoc analyses. Among patients with CKD who are not on dialysis, Joint National Commission-7 guidelines recommend lowering BP to <130/80 mm Hg.1 However, even these recommendations are based on largely observational data or post hoc analyses of randomized controlled trial data. In fact, the three randomized controlled clinical trials that targeted BP to two different levels among patients with CKD have failed to confirm the superiority of one level over the other with respect to renal or cardiovascular outcomes.2, 3, 4 Each of these three clinical trials targeted BP measured in the clinic. It is now becoming increasingly apparent that BP levels assessed in the clinic do not agree well with the usual level of BP; the usual level of BP is commonly assessed using 24-h ambulatory BP monitoring.5 Using 24-h ambulatory BP monitoring as the reference standard, a recent meta-analysis revealed that ∼20% of patients with CKD have white coat hypertension and about 5–10% have masked hypertension.6 Other studies using more liberal definitions of masked hypertension have found a much higher prevalence of masked hypertension. The classification of patients into these two categories of masked hypertension and white coat hypertension is of more than statistical importance.

n and about 5–10% have masked hypertension.6 Other studies using more liberal definitions of masked hypertension have found a much higher prevalence of masked hypertension. The classification of patients into these two categories of masked hypertension and white coat hypertension is of more than statistical importance. Patients with white coat hypertension, whether assessed using ambulatory BP or home BP recordings, have a prognosis that is substantially better than those with sustained hypertension.7 On the other hand, patients with masked hypertension have a prognosis that is substantially worse than those with persistent normotension.8 These data suggest that the diagnosis and treatment of hypertension based on home BP recordings would be superior to those based on clinical recordings alone.9 In fact, both European and US guidelines suggest home BP monitoring for all patients, including those with CKD.10, 11 Despite these recommendations, the precise method of how to measure home BP, how frequently to measure it, and how low to target the BP goal requires more study. Accordingly, home measurements should be systematically tested in larger populations of patients with CKD.

g for all patients, including those with CKD.10, 11 Despite these recommendations, the precise method of how to measure home BP, how frequently to measure it, and how low to target the BP goal requires more study. Accordingly, home measurements should be systematically tested in larger populations of patients with CKD. Although many trials have demonstrated that management of hypertension in the population without CKD with home BP monitoring leads to better BP control, only two such trials have been conducted in those with CKD.12, 13 Accordingly, there is an urgent need to conduct such trials in those with CKD to evaluate the risks and benefits of this simple and effective therapy to treat hypertension. If trials demonstrate that superior outcomes can be obtained with home BP monitoring, it could transform the management of patients with CKD. It has long been recognized that the presence of even the slightest kidney disease causes blunting of the usual nocturnal decline in systolic BP with sleep.14, 15 This phenomenon of non-dipping among patients with CKD has been associated with poor outcomes in some but not all studies.16, 17, 18 BP patterns, besides the usual level of BP, may or may not contain prognostic information.19 Determination of the independent prognostic significance of BP patterns among patients with CKD needs to be further studied.20 In particular, it is unknown whether non-dipping is a mediator or a marker of poor outcomes. This notion can only be tested in randomized trials.

ay or may not contain prognostic information.19 Determination of the independent prognostic significance of BP patterns among patients with CKD needs to be further studied.20 In particular, it is unknown whether non-dipping is a mediator or a marker of poor outcomes. This notion can only be tested in randomized trials. CKD is a state of accelerated vascular aging. Many studies have used pulse pressure as a proxy of vascular age. These studies have shown a strong and linear relationship between pulse pressure and mortality among dialysis patients. However, pulse pressure is not the best proxy of vascular age. Vascular age is better reflected by increased arterial stiffness; this can be easily measured by carotid to femoral pulse wave velocity. There is an excellent relationship between directly measured intra-aortic pulse wave velocity and systolic interdialytic ambulatory BP.21 There is also ample evidence to draw a direct relationship between pulse wave velocity and adverse outcomes among dialysis patients.20 How the evaluation of arterial stiffness adds to the management of hypertension among patients with CKD now needs to be better defined. As arterial stiffness through the measurement of pulse wave velocity and central BP can be measured using the same equipment, it is possible to evaluate the role of central BP and pulse wave velocity simultaneously.

stiffness adds to the management of hypertension among patients with CKD now needs to be better defined. As arterial stiffness through the measurement of pulse wave velocity and central BP can be measured using the same equipment, it is possible to evaluate the role of central BP and pulse wave velocity simultaneously. END-STAGE RENAL DISEASE PATIENTS Among patients on chronic dialysis, the clinical guidelines for the level to which BP should be lowered are opinion based. These opinions suggest lowering predialysis BP to <140/90 mm Hg and postdialysis BP to <130/80 mm Hg.22 However, some data suggest that achieving these targets is associated with increased episodes of intradialytic hypotension.23 In fact, collective evidence suggests that predialysis and postdialysis BP measurements are poor estimates of interdialytic ambulatory BP measurements.24 In contrast to peridialytic BP measurements, BP measurements outside the dialysis unit are associated with target organ damage and prognosis.25, 26, 27 Furthermore, at least one randomized trial has suggested that when BP is targeted using home BP rather than pre- or postdialysis measurements, better ambulatory BP control is achieved at 6 months.12 These data further support the use of home BP monitoring in the management of hypertension among hemodialysis patients.

7 Furthermore, at least one randomized trial has suggested that when BP is targeted using home BP rather than pre- or postdialysis measurements, better ambulatory BP control is achieved at 6 months.12 These data further support the use of home BP monitoring in the management of hypertension among hemodialysis patients. A vexing problem especially among chronic hemodialysis patients is that of assessment of volume and its relationship with BP. The concept of dry weight has evolved over time and its definition has changed. Although there is no consensus on its definition, one such definition defines dry weight as the lowest tolerated postdialysis weight achieved by a gradual change in postdialysis weight at which there are minimal signs or symptoms of either hypovolemia or hypervolemia. Although clinical examination does not adequately detect latent increase in dry weight, several technologies such as relative plasma volume monitoring and body impedance analysis are emerging that may help in assessing dry weight in the future. There is a need to better evaluate these technologies.

mia or hypervolemia. Although clinical examination does not adequately detect latent increase in dry weight, several technologies such as relative plasma volume monitoring and body impedance analysis are emerging that may help in assessing dry weight in the future. There is a need to better evaluate these technologies. Among dialysis patients, volume overload, often subclinical, is the primary cause of resistant hypertension. Sodium restriction is a modifiable risk factor that can lead to better BP control. However, dietary sodium restriction requires lifestyle modifications that are difficult to implement, and even harder to sustain over the long term. Newer options may include resins that bind dietary sodium. Restricting dialysate sodium is a simpler but underexplored strategy that can reduce thirst, limit interdialytic weight gain, and assist in the achievement of dry weight. However, larger studies with firm end points are needed. Achievement of dry weight can improve interdialytic BP, reduce pulse pressure, and limit hospitalizations. Accordingly, reduction of volume overload with dietary and dialysate sodium restriction and periodic probing of dry weight is believed to be good clinical practice.28 Avoiding medication-directed control of BP may enhance the opportunity to probe dry weight, facilitate removal of volume, and limit the risk for pressure–volume overload, which may be a significant concern leading to myocardial remodeling in the hemodialysis patient. Probing dry weight among patients with end-stage renal disease has the potential to improve dismal cardiovascular outcomes.

to probe dry weight, facilitate removal of volume, and limit the risk for pressure–volume overload, which may be a significant concern leading to myocardial remodeling in the hemodialysis patient. Probing dry weight among patients with end-stage renal disease has the potential to improve dismal cardiovascular outcomes. The use of drugs to improve BP control among chronic hemodialysis patients is even more debatable. Epidemiological studies demonstrate that a lower BP and decline in BP over months or years are associated with higher mortality in dialysis patients. In contrast, randomized, controlled trials so far available have a low power to establish the benefits of antihypertensive therapy. A meta-analysis of five studies among 1202 hemodialysis patients demonstrated the overall benefit of antihypertensive therapy compared with the control or placebo group. It found a combined hazard ratio for cardiovascular events of 0.69 (95% confidence interval: 0.56–0.84).29 This meta-analysis suggests benefit but does not establish the value of the antihypertensive drug among hemodialysis patients. It remains unclear whether strategies to control BP with dry weight or drugs have associated risks such as increased episodes of intradialytic hypotension, subclinical myocardial ischemia, access dysfunction, and an accelerated demise of residual renal function. Thus, the risks and benefits of these techniques on cardiovascular outcomes need to be evaluated in adequately powered randomized trials.

e associated risks such as increased episodes of intradialytic hypotension, subclinical myocardial ischemia, access dysfunction, and an accelerated demise of residual renal function. Thus, the risks and benefits of these techniques on cardiovascular outcomes need to be evaluated in adequately powered randomized trials. In conclusion, despite many advances in the management of patients with CKD, both on and off dialysis, there exist several gaps in our knowledge. These relate to the definition of hypertension, assessment of volume, and evaluation of outcomes (see Figure 1). TO CITE THIS ARTICLE: Agarwal R, Martinez-Castelao A, Wiecek A et al. The lingering dilemma of arterial pressure in CKD: what do we know, where do we go? Kidney Int Sup 2011; 1: 17–20.

In conclusion, despite many advances in the management of patients with CKD, both on and off dialysis, there exist several gaps in our knowledge. These relate to the definition of hypertension, assessment of volume, and evaluation of outcomes (see Figure 1). TO CITE THIS ARTICLE: Agarwal R, Martinez-Castelao A, Wiecek A et al. The lingering dilemma of arterial pressure in CKD: what do we know, where do we go? Kidney Int Sup 2011; 1: 17–20. AC has received consulting fees from Abbott Laboratories and received lecture fees from F. Hoffmann-La Roche, Amgen, and Fresenius Medical Care Holdings. AM-C has received consulting fees from Abbott Laboratories, Roche Spain, and Abbott Spain. AO has received grant support from the Spanish Government. AW has received lecture fees from Amgen, F. Hoffmann-La Roche, and Janssen-Cileg. AW has also received grant support from Astellas Pharma. DF has received funding from the EU. DG has received consulting fees and lecture fees from Shire, Genzyme, Novartis AG, Sandoz, Pfizer, and Fresenius Medical Care Holdings. FWD has received funding from Amgen and Baxter. GL has received consulting fees from Amgen and Sandoz. GL has also received lecture fees from Amgen, Sandoz, Genzyme, and Shire. PJB has received consulting fees from Medtronic and has received grant support from Ardian and Novartis AG. RA has received consulting fees from Amgen, Abbott Laboratories, Merck, Affymax, Takeda Pharmaceutical Company, Daiichi Sankyo, Celgene, Watson Pharmaceuticals, and Rockwell Medical. RA has also received lecture fees from Abbott Laboratories, Merck, and Medscape. ZM has received lecture fees from Amgen, Shire, Genzyme, FMC, and Merck Sharp & Dohme. ZM has received grant support from Baxter, Amgen, FMC, Shire, and Genzyme. The remaining authors declared no competing interests.

and Rockwell Medical. RA has also received lecture fees from Abbott Laboratories, Merck, and Medscape. ZM has received lecture fees from Amgen, Shire, Genzyme, FMC, and Merck Sharp & Dohme. ZM has received grant support from Baxter, Amgen, FMC, Shire, and Genzyme. The remaining authors declared no competing interests. Figure 1 Research questions.

The endothelium is the innermost (single) cell lining of all blood vessels within the body; however, endothelial cell phenotypes may vary considerably in structure and function within different vascular regions.1, 2 For example, even between glomerular and peritubular capillaries, endothelial function differs significantly because of their high specialization. In this respect, the integrity of the endothelial cell layer has a pivotal role in many aspects of vascular function, for example, control of vasomotor tone and permeability, the latter being of paramount importance particularly for glomerular capillaries. However, despite the functional diversity of endothelial cells in different vascular compartments, a key common feature is their ability to synthesize and secrete a variety of factors impinging upon vascular tone and on vascular protection.3

latter being of paramount importance particularly for glomerular capillaries. However, despite the functional diversity of endothelial cells in different vascular compartments, a key common feature is their ability to synthesize and secrete a variety of factors impinging upon vascular tone and on vascular protection.3 The endothelium produces a range of vasorelaxant factors, the most significant and well characterized of which is nitric oxide (NO). NO is a fundamental gas that stimulates relaxation of vascular smooth muscle cells and inhibits their proliferation, and prevents leukocyte attachment and migration into the arterial wall, and platelet adhesion and aggregation to the endothelium. Prostacyclin and endothelium-derived hyperpolarizing factor are also important endothelium-derived vasorelaxants, with the latter contributing to endothelium-dependent vasodilatation in resistant arteries. The vast majority of studies on endothelial dysfunction have concentrated on the mechanisms responsible for the decreased bioavailability of NO, which may result from a decrease in NO production, from a decrease in activation of guanylyl cyclase, and/or from an increase in NO degradation. A decrease in NO production may result from reduced availability of substrates and cofactors for NO synthases, such as L-arginine or tetrahydrobiopterin; from a decreased expression of endothelial NO synthase (eNOS) or from a decreased activation of eNOS, such as phosphorylation of the enzyme or interactions with proteins (for example, heat shock protein 90 or calmodulin); or from high levels of endogenous inhibitors of eNOS, such as asymmetric dimethylarginine in particular. Finally, reduced NO bioavailability levels may be caused by the binding of NO to hemoglobin or from oxidative stress, which gives rise to peroxynitrite, a vasculotoxic substance. On the other hand, endothelial cells produce several vasoconstrictors, including endothelin-1, cyclooxygenase-derived prostanoids, reactive oxygen species, dinucleotide uridine adenosine tetraphosphate, and angiotensin II. When the balance between endothelium-derived vasorelaxants and vasoconstrictors is altered, endothelial dysfunction ensues.

lial cells produce several vasoconstrictors, including endothelin-1, cyclooxygenase-derived prostanoids, reactive oxygen species, dinucleotide uridine adenosine tetraphosphate, and angiotensin II. When the balance between endothelium-derived vasorelaxants and vasoconstrictors is altered, endothelial dysfunction ensues. Because of its enormous surface area within the body, the endothelium has an important role in major diseases such as hypertension and diabetes. In these conditions, the endothelium undergoes functional and structural alterations, eventually resulting in loss of its role as a protective barrier. Endothelial dysfunction is the earliest—merely functional—step in the cascade of events leading to atherosclerosis, and the fundamental feature of this condition is impaired NO bioavailability.4, 5 If perpetuated long enough, dysfunction of endothelial cells is followed by their apoptosis, which can finally result in functional and structural disintegration of the endothelial cell layer. This is paralleled by vascular ‘microinflammation' as a result of leukocyte and thrombocyte activation and adhesion, which further accelerates the vessel wall damage.6 This process leads to progressive atherosclerotic disease in larger vessels and/or complete disruption of smaller (tissue) blood vessels. Finally, vessel disappearance (‘vascular rarefaction') may terminate in malperfusion and hypoxia of tissues and whole organs. For this reason, hypertension and diabetes are recognized not only as main (cardio) vascular risk factors resulting in death due to atherosclerotic complications but also as the most important conditions leading to end-stage kidney disease as a consequence of progressive glomerulosclerosis and rarefaction of postglomerular capillaries. Here, the kidney pays the definite price for its generous vascular supply as the organ with the largest total endothelial surface area.

ications but also as the most important conditions leading to end-stage kidney disease as a consequence of progressive glomerulosclerosis and rarefaction of postglomerular capillaries. Here, the kidney pays the definite price for its generous vascular supply as the organ with the largest total endothelial surface area. In patients with chronic kidney disease (CKD), endothelial dysfunction may have a dual role. On the one hand, it is a crucial step in the development of cardiovascular disease (CVD). On the other hand, activation, dysfunction, and disintegration of endothelial cells in glomerular capillaries and particularly in the capillaries that nurture the renal medulla pave the way for CKD progression. It has become clear from experimental studies that vascular rarefaction in this capillary system is a crucial step toward renal tissue hypoxia and kidney damage.7 A myriad of factors are thought to be involved in the process of endothelial dysfunction and disintegration, which, through further steps of vascular injury, finally results in end-stage CVD and/or CKD. However, the competing twin risk of patients with CKD—development of CVD and/or CKD progression—is not well elucidated. In other words, we do not understand yet why some patients progress to end-stage kidney disease without significant cardiovascular events, whereas others die before reaching terminal renal failure due to complications of severe (larger-vessel) atherosclerosis. Furthermore, in patients without CKD, a large body of evidence supports the hypothesis that endothelial dysfunction and microinflammation represent major promoters for atherosclerosis and independently predict the risk of future cardiovascular events,8, 9, 10 whereas in patients with CKD firm evidence for this relationship has not been provided so far. Finally, the relationship between endothelial dysfunction of peripheral and renal vessels in CKD patients has not been explored in detail, and the prognostic value of endothelial dysfunction in the renal circulation for CKD progression is almost unknown. Here, the assessment of endothelial dysfunction is mostly restricted to functional tests such as the response of the renal (micro)circulation to vasodilator and/or vasoconstrictor stimuli, for example, NO inhibitors or angiotensin II—usually assessed by a change in the para-aminohippurate clearance.

is almost unknown. Here, the assessment of endothelial dysfunction is mostly restricted to functional tests such as the response of the renal (micro)circulation to vasodilator and/or vasoconstrictor stimuli, for example, NO inhibitors or angiotensin II—usually assessed by a change in the para-aminohippurate clearance. Furthermore, although the number of studies focusing on NO bioavailability and disturbed NO vasoregulation in CKD is on the rise, we still lack studies looking at the balance between vasorelaxant and vasoconstrictor factors rather than on single-factor perturbation.

is almost unknown. Here, the assessment of endothelial dysfunction is mostly restricted to functional tests such as the response of the renal (micro)circulation to vasodilator and/or vasoconstrictor stimuli, for example, NO inhibitors or angiotensin II—usually assessed by a change in the para-aminohippurate clearance. Furthermore, although the number of studies focusing on NO bioavailability and disturbed NO vasoregulation in CKD is on the rise, we still lack studies looking at the balance between vasorelaxant and vasoconstrictor factors rather than on single-factor perturbation. Many risk factors—traditional and non-traditional—are thought to have a more or less important role in the development of CVD and progression in CKD patients (Table 1). Some of these are established cardiovascular risk factors, for example, hypertension and smoking, and their successful treatment or cessation results in reduced cardiovascular events and in slowing down progression. Others only seem to identify patients at risk, that is, they are only markers of risk, such as high serum homocysteine. In patients with progressive CKD, the issue is further complicated because of the appearance of uremia-specific risk factors with the potential of contributing to endothelial and vascular dysfunction and damage (Table 2). In this respect, many novel putative ‘biomarkers' of risk—either for CVD or for progression—have been discovered in the last two decades. However, for many of them causality has not been proven yet, even in experimental studies, and for almost all of them the definitive confirmation of their pathophysiological role and clinical relevance from intervention trials in CKD patients is still pending. This will certainly be one of the most important future challenges in the field of (cardio)vascular research in nephrology (Figure 1). In addition, the relative contribution of these (risk) factors and markers to the twin risk of CVD and progression in CKD patients has not been appropriately investigated so far. In the face of the many discovered putative risk factors and markers in recent years, the above questions may be of greater importance than the search for further biomarkers with uncertain significance for CVD and progression in patients with CKD.

gression in CKD patients has not been appropriately investigated so far. In the face of the many discovered putative risk factors and markers in recent years, the above questions may be of greater importance than the search for further biomarkers with uncertain significance for CVD and progression in patients with CKD. TO CITE THIS ARTICLE: Fliser D, Wiecek A, Suleymanlar G et al. The dysfunctional endothelium in CKD and in cardiovascular disease: mapping the origin(s) of cardiovascular problems in CKD and of kidney disease in cardiovascular conditions for a research agenda. Kidney Int Sup 2011; 1: 6–9.

gression in CKD patients has not been appropriately investigated so far. In the face of the many discovered putative risk factors and markers in recent years, the above questions may be of greater importance than the search for further biomarkers with uncertain significance for CVD and progression in patients with CKD. TO CITE THIS ARTICLE: Fliser D, Wiecek A, Suleymanlar G et al. The dysfunctional endothelium in CKD and in cardiovascular disease: mapping the origin(s) of cardiovascular problems in CKD and of kidney disease in cardiovascular conditions for a research agenda. Kidney Int Sup 2011; 1: 6–9. AC has received consulting fees from Abbott Laboratories and received lecture fees from F. Hoffmann-La Roche, Amgen, and Fresenius Medical Care Holdings. AM-C has received consulting fees from Abbott Laboratories, Roche Spain, and Abbott Spain. AO has received grant support from the Spanish Government. AW has received lecture fees from Amgen, F. Hoffmann-La Roche, and Janssen-Cileg. AW has also received grant support from Astellas Pharma. DF has received funding from the EU. DG has received consulting fees and lecture fees from Shire, Genzyme, Novartis AG, Sandoz, Pfizer, and Fresenius Medical Care Holdings. FWD has received funding from Amgen and Baxter. GL has received consulting fees from Amgen and Sandoz. GL has also received lecture fees from Amgen, Sandoz, Genzyme, and Shire. PJB has received consulting fees from Medtronic and has received grant support from Ardian and Novartis AG. RA has received consulting fees from Amgen, Abbott Laboratories, Merck, Affymax, Takeda Pharmaceutical Company, Daiichi Sankyo, Celgene, Watson Pharmaceuticals, and Rockwell Medical. RA has also received lecture fees from Abbott Laboratories, Merck, and Medscape. ZM has received lecture fees from Amgen, Shire, Genzyme, FMC, and Merck Sharp & Dohme. ZM has received grant support from Baxter, Amgen, FMC, Shire, and Genzyme. The remaining authors declared no competing interests.

and Rockwell Medical. RA has also received lecture fees from Abbott Laboratories, Merck, and Medscape. ZM has received lecture fees from Amgen, Shire, Genzyme, FMC, and Merck Sharp & Dohme. ZM has received grant support from Baxter, Amgen, FMC, Shire, and Genzyme. The remaining authors declared no competing interests. Figure 1 Some open questions on the role of endothelium in the cardio-renal connection. Table 1 Risk factors and putative ‘biomarkers' for cardiovascular disease and progression in patients with chronic kidney disease Traditional Age Gender (male) Family history (genetic background) High blood pressure Obesity/physical inactivity Hyper- and dyslipidemia Increased fibrinogen/other coagulation disorders Hyperinsulinemia Glucose intolerance/hyperglycemia/diabetes Smoking ?

disease and progression in patients with chronic kidney disease Traditional Age Gender (male) Family history (genetic background) High blood pressure Obesity/physical inactivity Hyper- and dyslipidemia Increased fibrinogen/other coagulation disorders Hyperinsulinemia Glucose intolerance/hyperglycemia/diabetes Smoking ? Non-traditional Albuminuria/proteinuria Increased homocysteine Increased asymmetric dimethylarginine and other endogenous nitric oxide inhibitors Increased high-sensitivity C-reactive protein and other inflammatory markers Increased adhesion molecules Oxidative stress/increased production of reactive oxygen species Increased fatty acids/high lipoprotein a Increased advanced glycation endproducts Reduced adiponectin and/or increased leptin Reduced vitamin D Increased natriuretic peptides (e.g., NT-proBNP) ? Table 2 CKD-specific risk factors and putative ‘biomarkers' for cardiovascular disease and for progression in patients with CKD Volume overload/increased natriuretic peptides (e.g., NT-proBNP) Proteinuria Increased parathormone and calcium/phosphate product Increased fibroblast growth factor 23 Reduced vitamin D Acidosis Anemia Hypoalbuminemia Reduced fetuin A and other inhibitors of calcification Increased asymmetric dimethylarginine and other endogenous NO inhibitors Increased high-sensitivity C-reactive protein and other inflammatory markers Oxidative stress/increased production of reactive oxygen species Increased susceptibility to infections ? Abbreviations: CKD, chronic kidney disease; NO, nitric oxide.

on Increased asymmetric dimethylarginine and other endogenous NO inhibitors Increased high-sensitivity C-reactive protein and other inflammatory markers Oxidative stress/increased production of reactive oxygen species Increased susceptibility to infections ? Abbreviations: CKD, chronic kidney disease; NO, nitric oxide. Some of these factors are present also in patients without CKD, but they accumulate/disperse significantly with declining kidney function.

The increased interest toward mesenchymal stem cells (MSCs) is associated with the prominent role that these cells have recently provoked in tissue regeneration. Mainly, they represent an important component of the hematopoietic stem cell niche in the bone marrow (BM), where they contribute to hematopoietic stem cell maturation.1, 2 In kidney regeneration, in which the quest for a pharmacological therapy in acute kidney injury (AKI) has been largely unsuccessful, BM-MSCs might represent a valid therapeutic tool.

component of the hematopoietic stem cell niche in the bone marrow (BM), where they contribute to hematopoietic stem cell maturation.1, 2 In kidney regeneration, in which the quest for a pharmacological therapy in acute kidney injury (AKI) has been largely unsuccessful, BM-MSCs might represent a valid therapeutic tool. Acknowledged evidence has proved that BM supplies kidney with cells for physiological turnover or regeneration of tubular epithelial cells.3 Our group investigated the use of BM-MSCs as cell therapy for AKI and documented for the first time that murine BM-MSCs contributed to renal repair and recovery from AKI.4 By means of intravenous injection, murine BM-MSCs ameliorated renal function and tubular injury of mice with AKI induced by the nephrotoxic anticancer agent cisplatin (Table 1).4, 5 In the same experimental model, hematopoietic stem cells had no protective effect.4 Therefore, it is important to understand which BM-MSCs properties make these cells responsible for their decisive role in kidney repair. In kidneys of AKI-injured mice supplied with BM-MSCs, we observed, by Ki67 staining, a marked increase of tubular cell proliferation, a fundamental step by which kidney restores normal architecture after acute damage. Proliferating cells were identified as endogenous renal cells, as the vast majority of the cells lacked Y chromosome, the marker used for identifying BM-MSC male cells given to female mice, and were negative for PKH-26, the cell tracker used to stain BM-MSCs before their administration.4, 5

itecture after acute damage. Proliferating cells were identified as endogenous renal cells, as the vast majority of the cells lacked Y chromosome, the marker used for identifying BM-MSC male cells given to female mice, and were negative for PKH-26, the cell tracker used to stain BM-MSCs before their administration.4, 5 However, the mechanism responsible for the repair was still unclear. Indeed, in the face of a remarkable renoprotective and regenerative effect, in the kidney, the number of BM-MSCs was quantitatively very low, with an engraftment of PKH-26 MSC in the renal tissue that averaged 1.1±0.7 BM-MSC/105 renal cells.5 Moreover, the occasional differentiation of these cells into tubular cells could not explain and sustain per se such a therapeutic effect.

rative effect, in the kidney, the number of BM-MSCs was quantitatively very low, with an engraftment of PKH-26 MSC in the renal tissue that averaged 1.1±0.7 BM-MSC/105 renal cells.5 Moreover, the occasional differentiation of these cells into tubular cells could not explain and sustain per se such a therapeutic effect. Two considerations have been useful to advance a working hypothesis on the mechanism for regeneration of tubular epithelium: the first is that growth factors have the leading role in induction of a proliferative repair,6, 7 and the second is that BM-MSCs are responsible for the secretion of multiple bioactive factors.8, 9, 10, 11 Particularly, in the kidney, studies on rats with ischemia/reperfusion injury have indicated that BM-MSCs-mediated renal repair was associated with a high renal production of growth factors such as hepatocyte growth factor, vascular growth factor, and insulin-like growth factor-1 (IGF-1).8 Moreover, the effect of BM-MSCs was accompanied by the tissue downregulation of proinflammatory cytokines and upregulation of prosurvival mediators.8 These reasons prompted us to hypothesize that MSCs afford protection primarily through a paracrine pathway, and to investigate, among potential candidates, the contribution of IGF-1 to tissue protection. IGF-1 is constitutively expressed and secreted by BM-MSCs,8, 12 possesses mitogenic and antiapoptotic properties,13, 14 and is implicated as an important mediator in kidney regeneration in models of AKI.13, 15, 16 Our studies showed that in vitro, murine BM-MSCs induce proliferation of cisplatin-damaged tubular cells and protect tubular cells from cisplatin-induced apoptosis via IGF-1.5 Direct blocking of IGF-1, by specific IGF-1 small interfering RNAs or by specific antibody, confirmed that BM-MSC-derived IGF-1 mediated the proliferation of cisplatin-damaged tubular cells (Figure 1). Although partially, the inhibition of cisplatin-induced apoptosis on proximal tubular cells was also found to be IGF-1 mediated (Figure 1). In vivo, we have found confirmation of this evidence. Administration of IGF-1 gene-silenced BM-MSCs failed to protect renal function and tubular structure of mice injured with cisplatin. When BM-MSC localization within the kidney was explored, these cells were preferentially found in peritubular areas, further supporting the notion that their mitogenic and antiapoptotic action was exerted through a paracrine mechanism.

iled to protect renal function and tubular structure of mice injured with cisplatin. When BM-MSC localization within the kidney was explored, these cells were preferentially found in peritubular areas, further supporting the notion that their mitogenic and antiapoptotic action was exerted through a paracrine mechanism. These findings have opened up new extensive investigations on the effects of BM-MSCs principally in the surroundings of the tubuli. In the future perspective to use BM-MSCs in clinic, we used MSCs obtained from human BM aspirates to treat cisplatin-injured immunodeficient nonobese diabetes/severe combined immunodeficiency mice.17 Similar to the corresponding murine cells, human BM-MSCs were engaged to engraft the kidney and to preserve its tubular integrity and renal function, ultimately leading to a proliferation and reduced apoptosis of tubular cells. Moreover, we reported a clear effect on survival of mice receiving human BM-MSCs as compared with AKI mice that were administered saline.17 It is fully recognized that the main target of cisplatin-induced damage is tubular epithelial cell with DNA damage, mitochondrial dysfunction, and reactive oxygen species production, followed by apoptosis. However, cisplatin also induces intrarenal vasoactive mediators18 and proinflammatory factors, specifically tumor necrosis factor-α, which perturbs the peritubular endothelium19 leading to inflammatory cell migration and leukocyte-mediated changes of vascular tone and perfusion. The persistent vasoconstriction and reduction of blood flow, which develop later than the tubular epithelial damage, have been suggested to amplify the deleterious effects of tubular cell injury in AKI.20 These observations were confirmed in our AKI experimental model, where peritubular capillaries were significantly reduced in volume density and diameter. Notably, human BM-MSCs treatment not only protected against tubular cell damage but also almost completely normalized the endothelium and lumen density, as well as the capillary diameters (Figure 2).17 These findings add new intriguing information about the role of MSCs in kidney recovery from AKI. Amelioration of hemodynamic changes, likely consequence of a more pervious capillary, should yield advantageous effects increasing tissue oxygenation, reducing endothelial cell activation, and preserving microvascular integrity.

findings add new intriguing information about the role of MSCs in kidney recovery from AKI. Amelioration of hemodynamic changes, likely consequence of a more pervious capillary, should yield advantageous effects increasing tissue oxygenation, reducing endothelial cell activation, and preserving microvascular integrity. Finally, an active role of BM-MSCs on endothelial cells cannot be excluded considering that these cells might induce prosurvival pathways and antioxidant mechanisms. However, several of these pathways still need to find experimental confirmation. At the moment, the overall findings can only suggest a hypothesis for the complex link of BM-MSCs-mediated regeneration. Following MSC recruitment to damaged tissues, the recovery process starts and passes through MSC-secreted growth factors, one of which is certainly IGF-1, which promotes prosurvival pathways and stimulates tubular cell proliferation (Figure 3).

suggest a hypothesis for the complex link of BM-MSCs-mediated regeneration. Following MSC recruitment to damaged tissues, the recovery process starts and passes through MSC-secreted growth factors, one of which is certainly IGF-1, which promotes prosurvival pathways and stimulates tubular cell proliferation (Figure 3). Studies conducted by our group on MSCs obtained from a different source, the human cord blood (CB-MSCs), confirmed and extended some of the findings highlighted for BM-MSCs.21 Specifically, data in murine model of cisplatin-induced AKI showed the great potential of CB-MSC in terms of protection from renal function impairment and remarkable prolongation of animal survival. CB-MSC treatment led to reduction of apoptosis and tubular cell proliferation. Moreover, the favorable effect on renal tissues was associated with inhibition of tubular oxidative damage, in terms of nitrotyrosine expression and induction of the phosphorylation of the prosurvival factor Akt.21 Similar to the MSCs derived from BM, CB-MSCs almost exclusively localized in the peritubular areas where they are likely to promote regeneration through paracrine action. In support to this statement are in vitro experiments showing that the proregenerative growth factors fibroblast growth factor, heparin binding-epidermal growth factor-like growth factor, vascular endothelial growth factor, and hepatocyte growth factor are increased in the supernatant of cisplatin-treated proximal tubular cells cocultured with CB-MSCs. The release of inflammatory cytokines, such as interleukin-1α and transforming growth factor-β, was found to be decreased.21

h factor-like growth factor, vascular endothelial growth factor, and hepatocyte growth factor are increased in the supernatant of cisplatin-treated proximal tubular cells cocultured with CB-MSCs. The release of inflammatory cytokines, such as interleukin-1α and transforming growth factor-β, was found to be decreased.21 Studies that aimed to further clarify the mechanism responsible for renoprotection by MSCs obtained from different sources are now essential to support an educated answer to the question of which cell type will definitely represent the best therapeutic strategy for kidney regeneration. We are really indebted to Professor Giuseppe Remuzzi for valuable comments and suggestions. TO CITE THIS ARTICLE: Imberti B, Morigi M, Benigni A. Potential of mesenchymal stem cells in the repair of tubular injury. Kidney inter., Suppl. 2011; 1: 90–93. All the authors declared no competing interests.

Studies that aimed to further clarify the mechanism responsible for renoprotection by MSCs obtained from different sources are now essential to support an educated answer to the question of which cell type will definitely represent the best therapeutic strategy for kidney regeneration. We are really indebted to Professor Giuseppe Remuzzi for valuable comments and suggestions. TO CITE THIS ARTICLE: Imberti B, Morigi M, Benigni A. Potential of mesenchymal stem cells in the repair of tubular injury. Kidney inter., Suppl. 2011; 1: 90–93. All the authors declared no competing interests. Figure 1 Bone marrow-mesenchymal stem cells (BM-MSCs) stimulated proximal tubular cell (PTEC) proliferation and inhibited cisplatin-induced apoptosis via insulin-like growth factor-1 (IGF-1). (a) Proliferation of PTECs treated with cisplatin (2.5 μM, 6 h), alone or in coculture for 4 days with BM-MSCs transfected with irrelevant (irrel) or IGF-1 small interfering (si) RNAs. Blocking of IGF-1 by RNA silencing (si-IGF-1) led to a strong reduction in the proliferation of cisplatin-treated PTECs as compared with BM-MSCs transfected with si-irrel. °P<0.01 versus PTECs; *P<0.01 versus PTECs+cisplatin; #P<0.05 versus si-irrel BM-MSCs. Data are means±s.e.m. (b) Cisplatin-induced apoptosis on PTECs is reduced by BM-MSCs treatment via IGF-1. Untreated PTECs and cisplatin-treated PTECs, alone or in coculture for 4 days with si-irrel BM-MSCs or si-IGF-1 BM-MSCs, were analyzed by fluorescence-activated cell sorter to determine late apoptosis (expression of caspase 3 and 7, propidium iodide). °P<0.01 versus PTECs; *P<0.01 versus PTECs+cisplatin; #P<0.05 versus si-irrel BM-MSCs and PTECs+cisplatin. Data are means±s.e.m.

ulture for 4 days with si-irrel BM-MSCs or si-IGF-1 BM-MSCs, were analyzed by fluorescence-activated cell sorter to determine late apoptosis (expression of caspase 3 and 7, propidium iodide). °P<0.01 versus PTECs; *P<0.01 versus PTECs+cisplatin; #P<0.05 versus si-irrel BM-MSCs and PTECs+cisplatin. Data are means±s.e.m. Figure 2 Effect of human bone marrow-mesenchymal stem cells (hBM-MSCs) on peritubular capillaries in immunodeficient mice with cisplatin-induced acute kidney injury. Representative micrografts of kidney tissues of control mouse and cisplatin-treated mice that were administered saline or hBM-MSCs at 4 days after cisplatin. The peritubular capillary endothelium was labeled with MECA-32 (red), whereas renal structures were stained with fluorescein isothiocyanate-labeled lectin wheat germ agglutinin (green). Volume density of endothelial cells and capillary lumen was markedly reduced in cisplatin-treated mice that were administered saline as compared with control mice, as well as their capillary diameter. Treatment with hBM-MSCs normalized all these parameters. Original magnification, × 630.

wheat germ agglutinin (green). Volume density of endothelial cells and capillary lumen was markedly reduced in cisplatin-treated mice that were administered saline as compared with control mice, as well as their capillary diameter. Treatment with hBM-MSCs normalized all these parameters. Original magnification, × 630. Figure 3 Suggested mechanism for mesenchymal stem cell (MSC)-mediated tubular repair after acute injury. Administered MSCs are attracted to the site of injury following cytokine and chemokine release from damaged tubular cells. Recruited MSCs release growth factors such as insulin-like growth factor-1 (IGF-1), which may affect tubular functional and structural repair by induction of cell proliferation and inhibition of apoptosis. MAP-kinase, mitogen-activated protein kinase; PI3K, phosphatidylinositol 3-kinase. Table 1 Bone marrow-derived mesenchymal stem cells (BM-MSCs) protected cisplatin-treated mice from renal function deterioration and renal histology changes Control Cisplatin+saline (4d) Cisplatin+BM- MSCs (4d) Renal function BUN (mg/dl) 17.02±1.00 82.54±5.12* 33.43±3.06°°

Figure 3 Suggested mechanism for mesenchymal stem cell (MSC)-mediated tubular repair after acute injury. Administered MSCs are attracted to the site of injury following cytokine and chemokine release from damaged tubular cells. Recruited MSCs release growth factors such as insulin-like growth factor-1 (IGF-1), which may affect tubular functional and structural repair by induction of cell proliferation and inhibition of apoptosis. MAP-kinase, mitogen-activated protein kinase; PI3K, phosphatidylinositol 3-kinase. Table 1 Bone marrow-derived mesenchymal stem cells (BM-MSCs) protected cisplatin-treated mice from renal function deterioration and renal histology changes Control Cisplatin+saline (4d) Cisplatin+BM- MSCs (4d) Renal function BUN (mg/dl) 17.02±1.00 82.54±5.12* 33.43±3.06°° Renal histology Casts 0 1.77±0.20 0.50±0.19°° Tubular cell degeneration 0 1.54±0.14 0.62±0.18° Cell loss 0 1.38±0.18 0.12±0.12°° Mice were subcutaneously administered with cisplatin (12.7 mg/kg) and 24 h later BM-MSCs (2 × 105 cells) were given intravenously in the tail vein. Animals were killed at day 4 (4d). Renal function was measured as blood urea nitrogen (BUN). Renal histology was evaluated as periodic acid Shiff-positive droplets, cell debris in tubular lumens, and tubular degeneration consisting of brush border loss, nuclear changes, and vacuolization. BUN and histology data are mean values±s.e.m. and mean score±s.e.m., respectively. *P<0.01 versus control; °P<0.05, °°P<0.01 versus cisplatin+saline.

SECTION I: USE OF THE CLINICAL PRACTICE GUIDELINE This Clinical Practice Guideline document is based upon systematic literature searches last conducted in October 2010, supplemented with additional evidence through March 2012. It is designed to provide information and assist decision making. It is not intended to define a standard of care, and should not be construed as one, nor should it be interpreted as prescribing an exclusive course of management. Variations in practice will inevitably and appropriately occur when clinicians take into account the needs of individual patients, available resources, and limitations unique to an institution or type of practice. Every health-care professional making use of these recommendations is responsible for evaluating the appropriateness of applying them in any particular clinical situation. The recommendations for research contained within this document are general and do not imply a specific protocol.

n institution or type of practice. Every health-care professional making use of these recommendations is responsible for evaluating the appropriateness of applying them in any particular clinical situation. The recommendations for research contained within this document are general and do not imply a specific protocol. SECTION II: DISCLOSURE Kidney Disease: Improving Global Outcomes (KDIGO) makes every effort to avoid any actual or reasonably perceived conflicts of interest that may arise as a result of an outside relationship or a personal, professional, or business interest of a member of the Work Group. All members of the Work Group are required to complete, sign, and submit a disclosure and attestation form showing all such relationships that might be perceived or actual conflicts of interest. This document is updated annually and information is adjusted accordingly. All reported information will be printed in the final publication and are on file at the National Kidney Foundation (NKF), Managing Agent for KDIGO.

It is our hope that this document will serve several useful purposes. Our primary goal is to improve patient care. We hope to accomplish this, in the short term, by helping clinicians know and better understand the evidence (or lack of evidence) that determines current practice. By providing comprehensive evidence-based recommendations, this guideline will also help define areas where evidence is lacking and research is needed. Helping to define a research agenda is an often neglected, but very important, function of clinical practice guideline development. We used the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) system to rate the strength of evidence and the strength of recommendations. In all, there were only 2 (5.4%) recommendations in this guideline for which the overall quality of evidence was graded ‘A,' whereas 9 (24.3%) were graded ‘B,' 14 (37.8%) were graded ‘C,' and 12 (32.4%) were graded ‘D.' Although there are reasons other than quality of evidence to make a grade 1 or 2 recommendation, in general, there is a correlation between the quality of overall evidence and the strength of the recommendation. Thus, there were 15 (40.5%) recommendations graded ‘1' and 22 (59.5%) graded ‘2.' There were 2 (5.4%) recommendations graded ‘1A,' 8 (21.6%) were ‘1B,' 1 (2.7%) were ‘1C,' and 4 (10.8%) were ‘1D.' There were 0 (0%) graded ‘2A,' 1 (2.7%) were ‘2B,' 13 (35.1%) were ‘2C,' and 8 (21.6%) were ‘2D.' There were 22 (37.3%) statements that were not graded.

ecommendations graded ‘1' and 22 (59.5%) graded ‘2.' There were 2 (5.4%) recommendations graded ‘1A,' 8 (21.6%) were ‘1B,' 1 (2.7%) were ‘1C,' and 4 (10.8%) were ‘1D.' There were 0 (0%) graded ‘2A,' 1 (2.7%) were ‘2B,' 13 (35.1%) were ‘2C,' and 8 (21.6%) were ‘2D.' There were 22 (37.3%) statements that were not graded. Some argue that recommendations should not be made when evidence is weak. However, clinicians still need to make clinical decisions in their daily practice, and they often ask, ‘What do the experts do in this setting?' We opted to give guidance, rather than remain silent. These recommendations are often rated with a low strength of recommendation and a low strength of evidence, or were not graded. It is important for the users of this guideline to be cognizant of this (see Notice). In every case these recommendations are meant to be a place for clinicians to start, not stop, their inquiries into specific management questions pertinent to the patients they see in daily practice. We wish to thank the Work Group Co-Chairs, Drs John McMurray and Pat Parfrey, along with all of the Work Group members who volunteered countless hours of their time developing this guideline. We also thank the Evidence Review Team members and staff of the National Kidney Foundation who made this project possible. Finally, we owe a special debt of gratitude to the many KDIGO Board members and individuals who volunteered time reviewing the guideline, and making very helpful suggestions. Bertram L Kasiske, MD KDIGO Co-Chair David C Wheeler, MD, FRCP KDIGO Co-Chair

Work Group Co-Chairs John J V McMurray, MD, FRCP, FESC BHF Glasgow Cardiovascular Research Centre Glasgow, United Kingdom Patrick S Parfrey, MD, FRCPC, FRSC Memorial University Medical School St John's, Canada Work Group John W Adamson, MD University of California at San Diego San Diego, CA, USA Pedro Aljama, MD, PhD Hospital Universitario Reina Sofía Córdoba, Spain Jeffrey S Berns, MD The Perelman School of Medicine at the University of Pennsylvania Philadelphia, PA, USA Julia Bohlius, MD, MScPH University of Bern Bern, Switzerland Tilman B Drüeke, MD, FRCP Université de Picardie Jules Verne Amiens, France Fredric O Finkelstein, MD Yale University New Haven, CT, USA Steven Fishbane, MD North Shore-LIJ Health System Manhasset, NY, USA Tomas Ganz, PhD, MD David Geffen School of Medicine at UCLA Los Angeles, CA, USA Iain C Macdougall, BSc, MD, FRCP King's College Hospital London, United Kingdom Ruth A McDonald, MD Seattle Children's Hospital Seattle, WA, USA Lawrence P McMahon, MBBS, MD Monash University Box Hill, Australia Gregorio T Obrador, MD, MPH Universidad Panamericana School of Medicine Mexico City, Mexico Giovanni FM Strippoli, MD, PhD, MPH Consorzio Mario Negri Sud Chieti, Italy Günter Weiss, MD Medical University of Innsbruck Innsbruck, Austria Andrzej Wie?cek, MD, PhD, FRCP Silesian University School of Medicine Katowice, Poland Evidence Review Team Tufts Center for Kidney Disease Guideline Development and Implementation Tufts Medical Center, Boston, MA, USA: Ethan M Balk, MD, MPH; Project Director; Program Director, Evidence-based Medicine Ashish Upadhyay, MD, Assistant Project Director Dana C Miskulin, MD, MS, Staff Nephrologist

Andrzej Wie?cek, MD, PhD, FRCP Silesian University School of Medicine Katowice, Poland Evidence Review Team Tufts Center for Kidney Disease Guideline Development and Implementation Tufts Medical Center, Boston, MA, USA: Ethan M Balk, MD, MPH; Project Director; Program Director, Evidence-based Medicine Ashish Upadhyay, MD, Assistant Project Director Dana C Miskulin, MD, MS, Staff Nephrologist Amy Earley, BS, Project Coordinator Shana Haynes, MS, DHSc, Research Assistant Jenny Lamont, MS, Project Manager In addition, support and supervision were provided by: Katrin Uhlig, MD, MS; Director, Guideline Development

The 2012 Kidney Disease: Improving Global Outcomes (KDIGO) Clinical Practice Guideline for Anemia in Chronic Kidney Disease aims to provide guidance on diagnosis, evaluation, management and treatment for all CKD patients (non-dialysis, dialysis, kidney transplant recipients and children) at risk of or with anemia. Guideline development followed an explicit process of evidence review and appraisal. The guideline contains chapters addressing diagnosis and evaluation of anemia in CKD and the use of various therapeutic agents (iron, ESAs and other agents) and red cell transfusion as means of treatment. Treatment approaches are addressed in each chapter and guideline recommendations are based on systematic reviews of relevant trials. Appraisal of the quality of the evidence and the strength of recommendations followed the GRADE approach. Ongoing areas of controversies and limitations of the evidence are discussed and additional suggestions are also provided for future research.

Chapter 1: Diagnosis and evaluation of anemia in CKD TESTING FOR ANEMIA Frequency of testing for anemia 1.1.1: For CKD patients without anemia (as defined below in Recommendation 1.2.1 for adults and Recommendation 1.2.2 for children), measure Hb concentration when clinically indicated and (Not Graded): at least annually in patients with CKD 3 at least twice per year in patients with CKD 4–5ND at least every 3 months in patients with CKD 5HD and CKD 5PD 1.1.2: For CKD patients with anemia not being treated with an ESA, measure Hb concentration when clinically indicated and (Not Graded): at least every 3 months in patients with CKD 3–5ND and CKD 5PD at least monthly in patients with CKD 5HD [See Recommendations 3.12.1–3.12.3 for measurement of Hb concentration in patients being treated with ESA.] Diagnosis of anemia 1.2.1: Diagnose anemia in adults and children >15 years with CKD when the Hb concentration is <13.0 g/dl (<130 g/l) in males and <12.0 g/dl (<120 g/l) in females. (Not Graded) 1.2.2: Diagnose anemia in children with CKD if Hb concentration is <11.0 g/dl (<110 g/l) in children 0.5–5 years, <11.5 g/dl (115 g/l) in children 5–12 years, and <12.0 g/dl (120 g/l) in children 12–15 years. (Not Graded) Investigation of anemia 1.3: In patients with CKD and anemia (regardless of age and CKD stage), include the following tests in initial evaluation of the anemia (Not Graded): Complete blood count (CBC), which should include Hb concentration, red cell indices, white blood cell count and differential, and platelet count Absolute reticulocyte count Serum ferritin level Serum transferrin saturation (TSAT)

1.3: In patients with CKD and anemia (regardless of age and CKD stage), include the following tests in initial evaluation of the anemia (Not Graded): Complete blood count (CBC), which should include Hb concentration, red cell indices, white blood cell count and differential, and platelet count Absolute reticulocyte count Serum ferritin level Serum transferrin saturation (TSAT) Serum vitamin B12 and folate levels Chapter 2: Use of iron to treat anemia in CKD TREATMENT WITH IRON AGENTS 2.1.1: When prescribing iron therapy, balance the potential benefits of avoiding or minimizing blood transfusions, ESA therapy, and anemia-related symptoms against the risks of harm in individual patients (e.g., anaphylactoid and other acute reactions, unknown long-term risks). (Not Graded) 2.1.2: For adult CKD patients with anemia not on iron or ESA therapy we suggest a trial of IV iron (or in CKD ND patients alternatively a 1–3 month trial of oral iron therapy) if (2C): an increase in Hb concentration without starting ESA treatment is desired* and TSAT is ≤30% and ferritin is ≤500 ng/ml (≤500 μg/l) 2.1.3: For adult CKD patients on ESA therapy who are not receiving iron supplementation, we suggest a trial of IV iron (or in CKD ND patients alternatively a 1–3 month trial of oral iron therapy) if (2C): an increase in Hb concentration** or a decrease in ESA dose is desired*** and TSAT is ≤30% and ferritin is ≤500 ng/ml (≤500 μg/l)

2.1.3: For adult CKD patients on ESA therapy who are not receiving iron supplementation, we suggest a trial of IV iron (or in CKD ND patients alternatively a 1–3 month trial of oral iron therapy) if (2C): an increase in Hb concentration** or a decrease in ESA dose is desired*** and TSAT is ≤30% and ferritin is ≤500 ng/ml (≤500 μg/l) 2.1.4: For CKD ND patients who require iron supplementation, select the route of iron administration based on the severity of iron deficiency, availability of venous access, response to prior oral iron therapy, side effects with prior oral or IV iron therapy, patient compliance, and cost. (Not Graded) 2.1.5: Guide subsequent iron administration in CKD patients based on Hb responses to recent iron therapy, as well as ongoing blood losses, iron status tests (TSAT and ferritin), Hb concentration, ESA responsiveness and ESA dose in ESA treated patients, trends in each parameter, and the patient's clinical status. (Not Graded) 2.1.6: For all pediatric CKD patients with anemia not on iron or ESA therapy, we recommend oral iron (or IV iron in CKD HD patients) administration when TSAT is ≤20% and ferritin is ≤100 ng/ml (≤100 μg/l). (1D) 2.1.7: For all pediatric CKD patients on ESA therapy who are not receiving iron supplementation, we recommend oral iron (or IV iron in CKD HD patients) administration to maintain TSAT >20% and ferritin >100 ng/ml (>100 μg/l). (1D) *Based on patient symptoms and overall clinical goals, including avoidance of transfusion, improvement in anemia-related symptoms, and after exclusion of active infection.

2.1.7: For all pediatric CKD patients on ESA therapy who are not receiving iron supplementation, we recommend oral iron (or IV iron in CKD HD patients) administration to maintain TSAT >20% and ferritin >100 ng/ml (>100 μg/l). (1D) *Based on patient symptoms and overall clinical goals, including avoidance of transfusion, improvement in anemia-related symptoms, and after exclusion of active infection. **Consistent with Recommendations #3.4.2 and 3.4.3. ***Based on patient symptoms and overall clinical goals including avoidance of transfusion and improvement in anemia-related symptoms, and after exclusion of active infection and other causes of ESA hyporesponsiveness. IRON STATUS EVALUATION 2.2.1: Evaluate iron status (TSAT and ferritin) at least every 3 months during ESA therapy, including the decision to start or continue iron therapy. (Not Graded) 2.2.2: Test iron status (TSAT and ferritin) more frequently when initiating or increasing ESA dose, when there is blood loss, when monitoring response after a course of IV iron, and in other circumstances where iron stores may become depleted. (Not Graded) CAUTIONS REGARDING IRON THERAPY 2.3: When the initial dose of IV iron dextran is administered, we recommend (1B) and when the initial dose of IV non-dextran iron is administered, we suggest (2C) that patients be monitored for 60 minutes after the infusion, and that resuscitative facilities (including medications) and personnel trained to evaluate and treat serious adverse reactions be available. Iron during infection

CAUTIONS REGARDING IRON THERAPY 2.3: When the initial dose of IV iron dextran is administered, we recommend (1B) and when the initial dose of IV non-dextran iron is administered, we suggest (2C) that patients be monitored for 60 minutes after the infusion, and that resuscitative facilities (including medications) and personnel trained to evaluate and treat serious adverse reactions be available. Iron during infection 2.4: Avoid administering IV iron to patients with active systemic infections. (Not Graded) Chapter 3: Use of ESAs and other agents to treat anemia in CKD ESA INITIATION 3.1: Address all correctable causes of anemia (including iron deficiency and inflammatory states) prior to initiation of ESA therapy. (Not Graded) 3.2: In initiating and maintaining ESA therapy, we recommend balancing the potential benefits of reducing blood transfusions and anemia-related symptoms against the risks of harm in individual patients (e.g., stroke, vascular access loss, hypertension). (1B) 3.3: We recommend using ESA therapy with great caution, if at all, in CKD patients with active malignancy—in particular when cure is the anticipated outcome—(1B), a history of stroke (1B), or a history of malignancy (2C). 3.4.1: For adult CKD ND patients with Hb concentration ≥10.0 g/dl (≥100 g/l), we suggest that ESA therapy not be initiated. (2D)

3.3: We recommend using ESA therapy with great caution, if at all, in CKD patients with active malignancy—in particular when cure is the anticipated outcome—(1B), a history of stroke (1B), or a history of malignancy (2C). 3.4.1: For adult CKD ND patients with Hb concentration ≥10.0 g/dl (≥100 g/l), we suggest that ESA therapy not be initiated. (2D) 3.4.2: For adult CKD ND patients with Hb concentration <10.0 g/dl (<100 g/l) we suggest that the decision whether to initiate ESA therapy be individualized based on the rate of fall of Hb concentration, prior response to iron therapy, the risk of needing a transfusion, the risks related to ESA therapy and the presence of symptoms attributable to anemia. (2C) 3.4.3: For adult CKD 5D patients, we suggest that ESA therapy be used to avoid having the Hb concentration fall below 9.0 g/dl (90 g/l) by starting ESA therapy when the hemoglobin is between 9.0–10.0 g/dl (90–100 g/l). (2B) 3.4.4: Individualization of therapy is reasonable as some patients may have improvements in quality of life at higher Hb concentration and ESA therapy may be started above 10.0 g/dl (100 g/l). (Not Graded) 3.4.5: For all pediatric CKD patients, we suggest that the selection of Hb concentration at which ESA therapy is initiated in the individual patient includes consideration of potential benefits (e.g., improvement in quality of life, school attendance/performance, and avoidance of transfusion) and potential harms. (2D)

3.4.4: Individualization of therapy is reasonable as some patients may have improvements in quality of life at higher Hb concentration and ESA therapy may be started above 10.0 g/dl (100 g/l). (Not Graded) 3.4.5: For all pediatric CKD patients, we suggest that the selection of Hb concentration at which ESA therapy is initiated in the individual patient includes consideration of potential benefits (e.g., improvement in quality of life, school attendance/performance, and avoidance of transfusion) and potential harms. (2D) ESA MAINTENANCE THERAPY 3.5.1: In general, we suggest that ESAs not be used to maintain Hb concentration above 11.5 g/dl (115 g/l) in adult patients with CKD. (2C) 3.5.2: Individualization of therapy will be necessary as some patients may have improvements in quality of life at Hb concentration above 11.5 g/dl (115 g/l) and will be prepared to accept the risks. (Not Graded) 3.6: In all adult patients, we recommend that ESAs not be used to intentionally increase the Hb concentration above 13 g/dl (130 g/l). (1A) 3.7: In all pediatric CKD patients receiving ESA therapy, we suggest that the selected Hb concentration be in the range of 11.0 to 12.0 g/dl (110 to 120 g/l). (2D) ESA DOSING 3.8.1: We recommend determining the initial ESA dose using the patient's Hb concentration, body weight, and clinical circumstances. (1D) 3.8.2: We recommend that ESA dose adjustments be made based on the patient's Hb concentration, rate of change in Hb concentration, current ESA dose and clinical circumstances. (1B)

ESA DOSING 3.8.1: We recommend determining the initial ESA dose using the patient's Hb concentration, body weight, and clinical circumstances. (1D) 3.8.2: We recommend that ESA dose adjustments be made based on the patient's Hb concentration, rate of change in Hb concentration, current ESA dose and clinical circumstances. (1B) 3.8.3: We suggest decreasing ESA dose in preference to withholding ESA when a downward adjustment of Hb concentration is needed. (2C) 3.8.4: Re-evaluate ESA dose if (Not Graded): The patient suffers an ESA-related adverse event The patient has an acute or progressive illness that may cause ESA hyporesponsiveness (See Recommendations 3.13.1–3.13.2) ESA ADMINISTRATION 3.9.1: For CKD 5HD patients and those on hemofiltration or hemodiafiltration therapy, we suggest either intravenous or subcutaneous administration of ESA. (2C) 3.9.2: For CKD ND and CKD 5PD patients, we suggest subcutaneous administration of ESA. (2C) Frequency of administration 3.10: We suggest determining the frequency of ESA administration based on CKD stage, treatment setting, efficacy considerations, patient tolerance and preference, and type of ESA. (2C) TYPE OF ESA 3.11.1: We recommend choosing an ESA based on the balance of pharmacodynamics, safety information, clinical outcome data, costs, and availability. (1D) 3.11.2: We suggest using only ESAs that have been approved by an independent regulatory agency. Specifically for ‘copy' versions of ESAs, true biosimilar products should be used. (2D) EVALUATING AND CORRECTING PERSISTENT FAILURE TO REACH OR MAINTAIN INTENDED HEMOGLOBIN CONCENTRATION Frequency of monitoring

TYPE OF ESA 3.11.1: We recommend choosing an ESA based on the balance of pharmacodynamics, safety information, clinical outcome data, costs, and availability. (1D) 3.11.2: We suggest using only ESAs that have been approved by an independent regulatory agency. Specifically for ‘copy' versions of ESAs, true biosimilar products should be used. (2D) EVALUATING AND CORRECTING PERSISTENT FAILURE TO REACH OR MAINTAIN INTENDED HEMOGLOBIN CONCENTRATION Frequency of monitoring 3.12.1: During the initiation phase of ESA therapy, measure Hb concentration at least monthly. (Not Graded) 3.12.2: For CKD ND patients, during the maintenance phase of ESA therapy measure Hb concentration at least every 3 months. (Not Graded) 3.12.3: For CKD 5D patients, during the maintenance phase of ESA therapy measure Hb concentration at least monthly. (Not Graded) Initial ESA hyporesponsiveness 3.13.1: Classify patients as having ESA hyporesponsiveness if they have no increase in Hb concentration from baseline after the first month of ESA treatment on appropriate weight-based dosing. (Not Graded) 3.13.2: In patients with ESA hyporesponsiveness, we suggest avoiding repeated escalations in ESA dose beyond double the initial weight-based dose. (2D) Subsequent ESA hyporesponsiveness 3.14.1: Classify patients as having acquired ESA hyporesponsiveness if after treatment with stable doses of ESA, they require 2 increases in ESA doses up to 50% beyond the dose at which they had been stable in an effort to maintain a stable Hb concentration. (Not Graded)

3.13.2: In patients with ESA hyporesponsiveness, we suggest avoiding repeated escalations in ESA dose beyond double the initial weight-based dose. (2D) Subsequent ESA hyporesponsiveness 3.14.1: Classify patients as having acquired ESA hyporesponsiveness if after treatment with stable doses of ESA, they require 2 increases in ESA doses up to 50% beyond the dose at which they had been stable in an effort to maintain a stable Hb concentration. (Not Graded) 3.14.2: In patients with acquired ESA hyporesponsiveness, we suggest avoiding repeated escalations in ESA dose beyond double the dose at which they had been stable. (2D) Management of poor ESA responsiveness 3.15.1: Evaluate patients with either initial or acquired ESA hyporesponsiveness and treat for specific causes of poor ESA response. (Not Graded) 3.15.2: For patients who remain hyporesponsive despite correcting treatable causes, we suggest individualization of therapy, accounting for relative risks and benefits of (2D): decline in Hb concentration continuing ESA, if needed to maintain Hb concentration, with due consideration of the doses required, and blood transfusions ADJUVANT THERAPIES 3.16.1: We recommend not using androgens as an adjuvant to ESA treatment. (1B) 3.16.2: We suggest not using adjuvants to ESA treatment including vitamin C, vitamin D, vitamin E, folic acid, L-carnitine, and pentoxifylline. (2D)

continuing ESA, if needed to maintain Hb concentration, with due consideration of the doses required, and blood transfusions ADJUVANT THERAPIES 3.16.1: We recommend not using androgens as an adjuvant to ESA treatment. (1B) 3.16.2: We suggest not using adjuvants to ESA treatment including vitamin C, vitamin D, vitamin E, folic acid, L-carnitine, and pentoxifylline. (2D) EVALUATION FOR PURE RED CELL APLASIA (PRCA) 3.17.1: Investigate for possible antibody-mediated PRCA when a patient receiving ESA therapy for more than 8 weeks develops the following (Not Graded): Sudden rapid decrease in Hb concentration at the rate of 0.5 to 1.0 g/dl (5 to 10 g/l) per week OR requirement of transfusions at the rate of approximately 1 to 2 per week, AND Normal platelet and white cell counts, AND Absolute reticulocyte count less than 10,000/μl 3.17.2: We recommend that ESA therapy be stopped in patients who develop antibody-mediated PRCA. (1A) 3.17.3: We recommend peginesatide be used to treat patients with antibody-mediated PRCA. (1B) Chapter 4: Red cell transfusion to treat anemia in CKD USE OF RED CELL TRANSFUSION IN CHRONIC ANEMIA 4.1.1: When managing chronic anemia, we recommend avoiding, when possible, red cell transfusions to minimize the general risks related to their use. (1B) 4.1.2: In patients eligible for organ transplantation, we specifically recommend avoiding, when possible, red cell transfusions to minimize the risk of allosensitization. (1C)

Chapter 4: Red cell transfusion to treat anemia in CKD USE OF RED CELL TRANSFUSION IN CHRONIC ANEMIA 4.1.1: When managing chronic anemia, we recommend avoiding, when possible, red cell transfusions to minimize the general risks related to their use. (1B) 4.1.2: In patients eligible for organ transplantation, we specifically recommend avoiding, when possible, red cell transfusions to minimize the risk of allosensitization. (1C) 4.1.3: When managing chronic anemia, we suggest that the benefits of red cell transfusions may outweigh the risks in patients in whom (2C): ESA therapy is ineffective (e.g., hemoglobinopathies, bone marrow failure, ESA resistance) The risks of ESA therapy may outweigh its benefits (e.g., previous or current malignancy, previous stroke) 4.1.4: We suggest that the decision to transfuse a CKD patient with non-acute anemia should not be based on any arbitrary Hb threshold, but should be determined by the occurrence of symptoms caused by anemia. (2C) URGENT TREATMENT OF ANEMIA 4.2: In certain acute clinical situations, we suggest patients are transfused when the benefits of red cell transfusions outweigh the risks; these include (2C): When rapid correction of anemia is required to stabilize the patient's condition (e.g., acute hemorrhage, unstable coronary artery disease) When rapid pre-operative Hb correction is required

TESTING FOR ANEMIA BACKGROUND In any individual, anemia may be the initial laboratory sign of an underlying medical problem. Consequently, a complete blood count, including the hemoglobin (Hb) concentration, is routinely part of global health assessment in most adults, whether or not they have chronic kidney disease (CKD). In patients with CKD but stable kidney function, the appearance or progression of anemia may herald a new problem that is causing blood loss or is interfering with red cell production. The anemia should be evaluated independently of CKD stage in order to identify any reversible process contributing to the anemia. The causes of acquired anemia are myriad and too many to include in a guideline such as this. A comprehensive list of causes and the approach to diagnosis can be found in a standard textbook of medicine or hematology. The most commonly encountered reversible cause of chronic anemia or worsening anemia in CKD patients, other than anemia related directly to CKD, is iron deficiency anemia. Frequency of testing for anemia 1.1.1: For CKD patients without anemia (as defined below in Recommendation 1.2.1 for adults and Recommendation 1.2.2 for children), measure Hb concentration when clinically indicated and (Not Graded): at least annually in patients with CKD 3 at least twice per year in patients with CKD 4–5ND at least every 3 months in patients with CKD 5HD and CKD 5PD 1.1.2: For CKD patients with anemia not being treated with an ESA, measure Hb concentration when clinically indicated and (Not Graded): at least every 3 months in patients with CKD 3–5ND and CKD 5PD

1.1.1: For CKD patients without anemia (as defined below in Recommendation 1.2.1 for adults and Recommendation 1.2.2 for children), measure Hb concentration when clinically indicated and (Not Graded): at least annually in patients with CKD 3 at least twice per year in patients with CKD 4–5ND at least every 3 months in patients with CKD 5HD and CKD 5PD 1.1.2: For CKD patients with anemia not being treated with an ESA, measure Hb concentration when clinically indicated and (Not Graded): at least every 3 months in patients with CKD 3–5ND and CKD 5PD at least monthly in patients with CKD 5HD [See Recommendations 3.12.1–3.12.3 for measurement of Hb concentration in patients being treated with ESA.]

1.1.2: For CKD patients with anemia not being treated with an ESA, measure Hb concentration when clinically indicated and (Not Graded): at least every 3 months in patients with CKD 3–5ND and CKD 5PD at least monthly in patients with CKD 5HD [See Recommendations 3.12.1–3.12.3 for measurement of Hb concentration in patients being treated with ESA.] RATIONALE Relatively little is known about the development and progression of anemia in patients with CKD. Consequently, one cannot determine precisely the optimal frequency at which Hb levels should be monitored. The recommendation that patients with CKD be periodically evaluated for anemia rests on observations that, in the absence of use of erythropoiesis-stimulating agents (ESAs), there often is a gradual decline in Hb over time in patients with CKD as the level of glomerular filtration rate (GFR) declines,1 suggesting the need for regular surveillance of Hb concentration. The frequency of Hb monitoring, regardless of CKD stage, should be influenced by the Hb level (i.e., more frequent monitoring may be appropriate in patients with more severe anemia) and rate of decline in Hb level. As kidney function declines and in patients with more advanced CKD stages, the incidence and prevalence of anemia increases. Thus, in order to identify CKD patients who may need intervention with iron administration, an ESA, or even require a transfusion, more frequent monitoring of the Hb concentration will be necessary at later CKD stages.

ines and in patients with more advanced CKD stages, the incidence and prevalence of anemia increases. Thus, in order to identify CKD patients who may need intervention with iron administration, an ESA, or even require a transfusion, more frequent monitoring of the Hb concentration will be necessary at later CKD stages. More frequent monitoring is recommended for adult CKD 5HD and CKD 5PD patients with anemia who are not receiving an ESA; at least monthly in CKD 5HD patients and at least every 3 months in CKD 5PD patients. In CKD 5HD patients, Hb monitoring is traditionally performed prior to a mid-week hemodialysis (HD) session. While this is not essential it probably does tend to minimize Hb variability due to the longer inter-dialytic interval between the last treatment of one week and the first of the next. As in all patients, Hb testing should be performed whenever clinically indicated, such as after a major surgical procedure, hospitalization, or bleeding episode.

l it probably does tend to minimize Hb variability due to the longer inter-dialytic interval between the last treatment of one week and the first of the next. As in all patients, Hb testing should be performed whenever clinically indicated, such as after a major surgical procedure, hospitalization, or bleeding episode. In the pediatric population with CKD, there is no direct evidence to recommend a different frequency of monitoring for anemia than for adults. In the Chronic Kidney Disease in Children Prospective Cohort Study (CKiD), which evaluated 340 North American children with CKD using iohexol-determined GFR,2 below a GFR threshold of 43 ml/min per 1.73 m2, there was a linear relationship between Hb and GFR, with Hb 0.3 g/dl (3 g/l) lower per 5 ml/min per 1.73 m2 lower GFR. Above that threshold, there was a nonsignificant association of 0.1 g/dl (1 g/l) lower Hb for every 5 ml/min per 1.73 m2 lower GFR. Because serum creatinine-based estimated glomerular filtration rate (eGFR) using the Schwartz formula may overestimate the true GFR in the children3 providers need to consider the potential for Hb decline and anemia even at early stages of CKD and monitor accordingly. In children with CKD 5HD and CKD 5PD, monthly monitoring for anemia is standard clinical practice. Diagnosis of anemia 1.2.1: Diagnose anemia in adults and children >15 years with CKD when the Hb concentration is <13.0 g/dl (<130 g/l) in males and <12.0 g/dl (<120 g/l) in females. (Not Graded)

In the pediatric population with CKD, there is no direct evidence to recommend a different frequency of monitoring for anemia than for adults. In the Chronic Kidney Disease in Children Prospective Cohort Study (CKiD), which evaluated 340 North American children with CKD using iohexol-determined GFR,2 below a GFR threshold of 43 ml/min per 1.73 m2, there was a linear relationship between Hb and GFR, with Hb 0.3 g/dl (3 g/l) lower per 5 ml/min per 1.73 m2 lower GFR. Above that threshold, there was a nonsignificant association of 0.1 g/dl (1 g/l) lower Hb for every 5 ml/min per 1.73 m2 lower GFR. Because serum creatinine-based estimated glomerular filtration rate (eGFR) using the Schwartz formula may overestimate the true GFR in the children3 providers need to consider the potential for Hb decline and anemia even at early stages of CKD and monitor accordingly. In children with CKD 5HD and CKD 5PD, monthly monitoring for anemia is standard clinical practice. Diagnosis of anemia 1.2.1: Diagnose anemia in adults and children >15 years with CKD when the Hb concentration is <13.0 g/dl (<130 g/l) in males and <12.0 g/dl (<120 g/l) in females. (Not Graded) 1.2.2: Diagnose anemia in children with CKD if Hb concentration is <11.0 g/dl (<110 g/l) in children 0.5–5 years, <11.5 g/dl (115 g/l) in children 5–12 years, and <12.0 g/dl (120 g/l) in children 12–15 years. (Not Graded)

1.2.1: Diagnose anemia in adults and children >15 years with CKD when the Hb concentration is <13.0 g/dl (<130 g/l) in males and <12.0 g/dl (<120 g/l) in females. (Not Graded) 1.2.2: Diagnose anemia in children with CKD if Hb concentration is <11.0 g/dl (<110 g/l) in children 0.5–5 years, <11.5 g/dl (115 g/l) in children 5–12 years, and <12.0 g/dl (120 g/l) in children 12–15 years. (Not Graded) RATIONALE The Hb concentration values that define anemia and should lead to initiation of an evaluation for the cause of anemia are dependent on sex and age. The recommended Hb values for adults and children represent the World Health Organization (WHO) definition of anemia and establish a benchmark for anemia workup that has been applied across populations.4 An alternative source for Hb concentration values that define anemia in children between 1 and 19 years is based on US National Health and Nutrition Examination Survey III (NHANES III) data from 1988–945 (Table 1). For children between birth and 24 months, the data are taken from normal reference values6 (Table 2). These thresholds for diagnosis of anemia and evaluation for the causes of anemia should not be interpreted as being thresholds for treatment of anemia. Rather than relying on a single laboratory test value, in patients without an apparent cause for a low Hb level, the value should be confirmed to be below the threshold values for diagnosis of anemia prior to initiating a diagnostic work up. Investigation of anemia

These thresholds for diagnosis of anemia and evaluation for the causes of anemia should not be interpreted as being thresholds for treatment of anemia. Rather than relying on a single laboratory test value, in patients without an apparent cause for a low Hb level, the value should be confirmed to be below the threshold values for diagnosis of anemia prior to initiating a diagnostic work up. Investigation of anemia 1.3: In patients with CKD and anemia (regardless of age and CKD stage), include the following tests in initial evaluation of the anemia (Not Graded): Complete blood count (CBC), which should include Hb concentration, red cell indices, white blood cell count and differential, and platelet count Absolute reticulocyte count Serum ferritin level Serum transferrin saturation (TSAT) Serum vitamin B12 and folate levels RATIONALE Complete blood count The complete blood count (CBC) provides information about the severity of anemia and adequacy of bone marrow function. Severity of anemia is assessed best by measuring Hb concentration rather than hematocrit. The latter measurement is a relatively unstable analyte and its measurement lacks standardization and is instrumentation dependent, since it is derived indirectly by automated analyzers.7, 8, 9 There is no evidence to support any different recommendation for the initial evaluation of anemia for children compared to adults.

latter measurement is a relatively unstable analyte and its measurement lacks standardization and is instrumentation dependent, since it is derived indirectly by automated analyzers.7, 8, 9 There is no evidence to support any different recommendation for the initial evaluation of anemia for children compared to adults. In addition to Hb concentration, other reported results of the CBC may convey important clinical information. The anemia of CKD is hypoproliferative, and in general, normochromic and normocytic. In this regard it is morphologically indistinguishable from the anemia of chronic disease.10 Folate or vitamin B12 deficiencies may lead to macrocytosis, whereas iron deficiency or inherited disorders of Hb formation (e.g., α- or β-thalassemia) may produce microcytosis. Iron deficiency, especially if long-standing, is associated with hypochromia (low mean corpuscular hemoglobin [MCH]). Macrocytosis with leukopenia or thrombocytopenia suggests a generalized disorder of hematopoiesis caused by toxins (e.g., alcohol), nutritional deficit (vitamin B12 or folate deficiency), or myelodysplasia. When these findings are present, further diagnostic evaluation may be indicated.

n corpuscular hemoglobin [MCH]). Macrocytosis with leukopenia or thrombocytopenia suggests a generalized disorder of hematopoiesis caused by toxins (e.g., alcohol), nutritional deficit (vitamin B12 or folate deficiency), or myelodysplasia. When these findings are present, further diagnostic evaluation may be indicated. The low erythropoietic activity that characterizes the anemia of CKD is consistent with insufficient erythropoietin stimulation. Erythropoietin levels are not routinely used in distinguishing erythropoietin deficiency from other causes of anemia in patients with CKD in most clinical settings and their measurement is generally not recommended.11, 12 Effective erythropoietic proliferative activity is most simply assessed by determination of the absolute reticulocyte count. Abnormalities of the white blood cell count and differential or platelet count are not typical of the anemia of CKD and should prompt investigation for other processes. Reticulocyte count, which may be obtained with automated CBC testing, may be high in patients who have active blood loss or hemolysis, and may be low in hypoproliferative erythropoiesis with anemia.

The low erythropoietic activity that characterizes the anemia of CKD is consistent with insufficient erythropoietin stimulation. Erythropoietin levels are not routinely used in distinguishing erythropoietin deficiency from other causes of anemia in patients with CKD in most clinical settings and their measurement is generally not recommended.11, 12 Effective erythropoietic proliferative activity is most simply assessed by determination of the absolute reticulocyte count. Abnormalities of the white blood cell count and differential or platelet count are not typical of the anemia of CKD and should prompt investigation for other processes. Reticulocyte count, which may be obtained with automated CBC testing, may be high in patients who have active blood loss or hemolysis, and may be low in hypoproliferative erythropoiesis with anemia. Iron status There are two important and distinct aspects of the assessment of iron status testing: the presence or absence of storage iron and the availability of iron to support ongoing erythropoiesis. The serum ferritin is the most commonly used test for evaluation of storage iron, for which the ‘gold standard' remains examination of a bone marrow aspiration stained for iron.13 The transferrin saturation (TSAT; serum iron × 100 divided by total iron binding capacity) is the most commonly used measure of the availability of iron to support erythropoiesis. The serum ferritin is affected by inflammation and is an ‘acute phase reactant'13 and, thus, ferritin values have to be interpreted with caution in CKD patients, especially those on dialysis in whom subclinical inflammation may be present.14

t commonly used measure of the availability of iron to support erythropoiesis. The serum ferritin is affected by inflammation and is an ‘acute phase reactant'13 and, thus, ferritin values have to be interpreted with caution in CKD patients, especially those on dialysis in whom subclinical inflammation may be present.14 Serum ferritin values ≤30 ng/ml (≤30 μg/l) indicate severe iron deficiency and are highly predictive of absent iron stores in bone marrow.15, 16 Ferritin values >30 ng/ml (>30 μg/l), however, do not necessarily indicate the presence of normal or adequate bone marrow iron stores. Studies assessing ferritin levels above which all or nearly all patients with CKD have normal bone marrow iron stores have produced varied results but most CKD patients, including those who are on HD, will have normal bone marrow iron stores when their serum ferritin level is ≥300 ng/ml (≥300 μg/l). Even at serum ferritin levels of 100 ng/ml (100 μg/l) most CKD patients have stainable bone marrow iron stores.16, 17, 18, 19, 20, 21 As will be discussed in Chapter 2, the serum ferritin and TSAT values are often used together to assess iron status, diagnose iron deficiency, and predict an erythropoietic response to iron supplementation (Supplementary Table 1 online).

l) most CKD patients have stainable bone marrow iron stores.16, 17, 18, 19, 20, 21 As will be discussed in Chapter 2, the serum ferritin and TSAT values are often used together to assess iron status, diagnose iron deficiency, and predict an erythropoietic response to iron supplementation (Supplementary Table 1 online). Other tests of iron status, such as percentage of hypochromic red blood cells and reticulocyte Hb content may be used instead of, or in addition to, TSAT and ferritin levels if available. Measurement of hepcidin levels has not been shown to be clinically useful or superior to more standard iron status tests in patients with CKD.22, 23 Vitamin B12 and folate Folate and vitamin B12 deficiency are uncommon but important causes of treatable anemia, typically associated with macrocytic red blood cell (RBC) indices. Limited data indicate a prevalence of vitamin B12 and folate deficiency in ≤10% of HD patients; the prevalence in CKD patients is not known. Nonetheless, since these deficiencies are easily correctable, and in the case of vitamin B12 may indicate other underlying disease processes, assessment of folate and vitamin B12 levels are generally considered standard components of anemia evaluation, especially in the presence of macrocytosis. Folate deficiency is best detected in most patients with serum folate level testing; RBC folate levels can be measured when serum folate levels are equivocal or when there is concern that recent dietary intake may obscure underlying folate deficiency using serum levels alone.24

pecially in the presence of macrocytosis. Folate deficiency is best detected in most patients with serum folate level testing; RBC folate levels can be measured when serum folate levels are equivocal or when there is concern that recent dietary intake may obscure underlying folate deficiency using serum levels alone.24 Additional tests Other tests, in addition to those indicated above, may be appropriate in individual patients and in certain specific clinical settings. For instance measurement of high sensitivity C-reactive protein (CRP) may be indicated if occult inflammation is a concern. In certain countries and/or in patients of specific nationalities or ethnicities, testing for hemoglobinopathies, parasites, and other conditions may be appropriate.

in specific clinical settings. For instance measurement of high sensitivity C-reactive protein (CRP) may be indicated if occult inflammation is a concern. In certain countries and/or in patients of specific nationalities or ethnicities, testing for hemoglobinopathies, parasites, and other conditions may be appropriate. DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. SUPPLEMENTARY MATERIAL Supplemental Table 1: Association between iron status and level of anemia in multivariable analyses. Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php

DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. SUPPLEMENTARY MATERIAL Supplemental Table 1: Association between iron status and level of anemia in multivariable analyses. Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php Table 1 Hb levels in children between 1–19 years for initiation of anemia workupa All races/ethnic groups Number of subjects Mean Hb g/dl (g/l) Standard deviation g/dl (g/l) Anemia definition met if value is <5th percentile g/dl (g/l) Boys 1 yr and over 12,623 14.7 (147) 1.4 (14) 12.1 (121) 1–2 yr 931 12.0 (120) 0.8 (8) 10.7 (107) 3–5 yr 1,281 12.4 (124) 0.8 (8) 11.2 (112) 6–8 yr 709 12.9 (129) 0.8 (8) 11.5 (115) 9–11 yr 773 13.3 (133) 0.8 (8) 12.0 (120) 12–14 yr 540 14.1 (141) 1.1 (11) 12.4 (124) 15–19 yr 836 15.1 (151) 1.0 (10) 13.5 (135)

dl (g/l) Boys 1 yr and over 12,623 14.7 (147) 1.4 (14) 12.1 (121) 1–2 yr 931 12.0 (120) 0.8 (8) 10.7 (107) 3–5 yr 1,281 12.4 (124) 0.8 (8) 11.2 (112) 6–8 yr 709 12.9 (129) 0.8 (8) 11.5 (115) 9–11 yr 773 13.3 (133) 0.8 (8) 12.0 (120) 12–14 yr 540 14.1 (141) 1.1 (11) 12.4 (124) 15–19 yr 836 15.1 (151) 1.0 (10) 13.5 (135) Girls 1 yr and over 13,749 13.2 (132) 1.1 (11) 11.4 (114) 1–2 yr 858 12.0 (120) 0.8 (8) 10.8 (108) 3–5 yr 1,337 12.4 (124) 0.8 (8) 11.1 (111) 6–8 yr 675 12.8 (128) 0.8 (8) 11.5 (115) 9–11 yr 734 13.1 (131) 0.8 (8) 11.9 (119) 12–14 yrb 621 13.3 (133) 1.0 (10) 11.7 (117) 15–19 yrb 950 13.2 (132) 1.0 (10) 11.5 (115) Hb, hemoglobin; yr, year. a Based on NHANES III data, United States, 1988–94.5 b Menstrual losses contribute to lower mean and 5th percentile Hb values for group. Table 2 Hb levels in children between birth and 24 months for initiation of anemia workupa Age Mean Hb g/dl (g/l) −2 SDb g/dl (g/l) Term (cord blood) 16.5 (165) 13.5 (135) 1–3 d 18.5 (185) 14.5 (145) 1 wk 17.5 (175) 13.5 (135) 2 wk 16.5 (165) 12.5 (125) 1 mo 14.0 (140) 10.0 (100) 2 mo 11.5 (115) 9.0 (90) 3–6 mo 11.5 (115) 9.5 (95) 6–24 mo 12.0 (120) 10.5 (105) d, day; Hb, hemoglobin; mo, month; SD, standard deviation; wk, week. a Data taken from normal reference values. This was published in Nathan DG, Orkin SH. Appendix 11: Normal hematologic values in children. In: Nathan DG, Orkin SH, Ginsburg D et al. (eds). Nathan and Oski's Hematology of Infancy and Childhood, 6th edn. p 1841, © Elsevier, 2003.6 b Values 2 standard deviations below the mean are equivalent to <2.5th percentile.

TREATMENT WITH IRON AGENTS BACKGROUND Correction of iron deficiency with oral or intravenous iron supplementation can reduce the severity of anemia in patients with CKD.25, 26 Untreated iron deficiency is an important cause of hyporesponsiveness to ESA treatment.27, 28 It is important to diagnose iron deficiency because treatment can readily correct the associated anemia and investigation for the cause of iron deficiency, which should follow its detection, can lead to important diagnoses. In the absence of menstrual bleeding, iron depletion and iron deficiency usually result from blood loss from the gastrointestinal tract. There are additional considerations in CKD patients with iron deficiency. For instance, hemodialysis patients are subject to repeated blood loss due to retention of blood in the dialyzer and blood lines. Other contributing causes in hemodialysis and other CKD patients include frequent blood sampling for laboratory testing, blood loss from surgical procedures (such as creation of vascular access), interference with iron absorption due to medications such as gastric acid inhibitors and phosphate binders, and reduced iron absorption due to inflammation.29 The reader is referred to standard textbooks of medicine and pediatrics for more extensive discussions on the diagnosis and evaluation of patients with known or suspected iron deficiency.

orption due to medications such as gastric acid inhibitors and phosphate binders, and reduced iron absorption due to inflammation.29 The reader is referred to standard textbooks of medicine and pediatrics for more extensive discussions on the diagnosis and evaluation of patients with known or suspected iron deficiency. Iron supplementation is widely used in CKD patients to treat iron deficiency, prevent its development in ESA-treated patients, raise Hb levels in the presence or absence of ESA treatment, and reduce ESA doses in patients receiving ESA treatment. Iron administration is appropriate when bone marrow iron stores are depleted or in patients who are likely to have a clinically meaningful erythropoietic response. It is prudent, however to avoid iron therapy in patients in whom it is unlikely to provide meaningful clinical benefit, i.e., avoid transfusion and reduce anemia-related symptoms, and in those in whom potential benefit is outweighed by risks of treatment.23, 30, 31, 32 There are relatively few data on the long-term clinical benefits of iron supplementation other than direct effects on the Hb concentration. There is similarly little information on the long-term adverse consequences of iron supplementation in excess of that necessary to provide adequate bone marrow iron stores.33, 34, 35 Since bone marrow aspiration for assessment of iron stores is rarely done in clinical practice, iron supplementation is typically assessed by blood-based iron status tests without knowledge of bone marrow iron stores.27, 28, 36, 37, 38

n in excess of that necessary to provide adequate bone marrow iron stores.33, 34, 35 Since bone marrow aspiration for assessment of iron stores is rarely done in clinical practice, iron supplementation is typically assessed by blood-based iron status tests without knowledge of bone marrow iron stores.27, 28, 36, 37, 38 The following statements provide recommendations for use of iron supplementation in patients with CKD. 2.1.1: When prescribing iron therapy, balance the potential benefits of avoiding or minimizing blood transfusions, ESA therapy, and anemia-related symptoms against the risks of harm in individual patients (e.g., anaphylactoid and other acute reactions, unknown long-term risks). (Not Graded) 2.1.2: For adult CKD patients with anemia not on iron or ESA therapy we suggest a trial of IV iron (or in CKD ND patients alternatively a 1–3 month trial of oral iron therapy) if (2C): an increase in Hb concentration without starting ESA treatment is desired* and TSAT is ≤30% and ferritin is ≤500 ng/ml (≤500 μg/l) 2.1.3: For adult CKD patients on ESA therapy who are not receiving iron supplementation, we suggest a trial of IV iron (or in CKD ND patients alternatively a 1–3 month trial of oral iron therapy) if (2C): an increase in Hb concentration** or a decrease in ESA dose is desired*** and TSAT is ≤30% and ferritin is ≤500 ng/ml (≤500 μg/l)

2.1.3: For adult CKD patients on ESA therapy who are not receiving iron supplementation, we suggest a trial of IV iron (or in CKD ND patients alternatively a 1–3 month trial of oral iron therapy) if (2C): an increase in Hb concentration** or a decrease in ESA dose is desired*** and TSAT is ≤30% and ferritin is ≤500 ng/ml (≤500 μg/l) 2.1.4: For CKD ND patients who require iron supplementation, select the route of iron administration based on the severity of iron deficiency, availability of venous access, response to prior oral iron therapy, side effects with prior oral or IV iron therapy, patient compliance, and cost. (Not Graded) 2.1.5: Guide subsequent iron administration in CKD patients based on Hb responses to recent iron therapy, as well as ongoing blood losses, iron status tests (TSAT and ferritin), Hb concentration, ESA responsiveness and ESA dose in ESA treated patients, trends in each parameter, and the patient's clinical status. (Not Graded) 2.1.6: For all pediatric CKD patients with anemia not on iron or ESA therapy, we recommend oral iron (or IV iron in CKD HD patients) administration when TSAT is ≤20% and ferritin is ≤100 ng/ml (≤100 μg/l). (1D) 2.1.7: For all pediatric CKD patients on ESA therapy who are not receiving iron supplementation, we recommend oral iron (or IV iron in CKD HD patients) administration to maintain TSAT >20% and ferritin >100 ng/ml (>100 μg/l). (1D)

2.1.6: For all pediatric CKD patients with anemia not on iron or ESA therapy, we recommend oral iron (or IV iron in CKD HD patients) administration when TSAT is ≤20% and ferritin is ≤100 ng/ml (≤100 μg/l). (1D) 2.1.7: For all pediatric CKD patients on ESA therapy who are not receiving iron supplementation, we recommend oral iron (or IV iron in CKD HD patients) administration to maintain TSAT >20% and ferritin >100 ng/ml (>100 μg/l). (1D) RATIONALE In patients with CKD-associated anemia, iron supplementation is intended to assure adequate iron stores for erythropoiesis, correct iron deficiency, and, in patients receiving ESA treatment, prevent iron deficiency from developing. Iron supplementation, particularly with intravenous iron, can enhance erythropoiesis and raise Hb levels in CKD patients with anemia even when TSAT and ferritin levels are not indicative of absolute iron deficiency, and even when bone marrow studies reveal adequate iron stores.38, 39, 40 Iron treatment, particularly when administered intravenously, has also been consistently demonstrated to improve the erythropoietic response to ESA treatment.27, 28, 32, 36, 37, 41, 42, 43 For any individual patient the optimal balance of Hb level, ESA dose, and iron dose at which clinical benefit is maximized and potential risk is minimized is not known. Prescribing iron therapy for CKD patients is complicated by the relatively poor diagnostic utility of serum ferritin and TSAT tests to estimate body iron stores or for predicting a Hb response to iron supplementation.23, 30 Even examination of bone marrow iron stores, considered the ‘gold standard' for assessment of iron stores, does not predict erythropoietic responsiveness to iron supplementation in patients with CKD with a high degree of accuracy.16, 23, 30, 40 It is important that the short- and long-term safety of oral and intravenous (IV) iron agents, when known, be carefully considered when iron therapy is prescribed, and that the potential for as yet undiscovered toxicities also be taken into account. In each patient there must be consideration of current and desired Hb level, ESA dose and trends in ESA dose over time, assessment of the Hb response to iron supplementation, ongoing blood loss, and changes in iron status tests. While observational studies have not for the most part produced strong evidence of significant toxicity of chronic IV iron administration, the clinical benefit of such treatment has also not been convincingly demonstrated, although a recent randomized controlled trial (RCT) in patients with heart failure (some of whom also had mild CKD) is encouraging.44

for the most part produced strong evidence of significant toxicity of chronic IV iron administration, the clinical benefit of such treatment has also not been convincingly demonstrated, although a recent randomized controlled trial (RCT) in patients with heart failure (some of whom also had mild CKD) is encouraging.44 TSAT and ferritin levels The two most widely available tests for assessing iron status are the TSAT and serum ferritin level. A very low serum ferritin (<30 ng/ml [<30 μg/l]) is indicative of iron deficiency.16 Except in this circumstance, the TSAT and serum ferritin level have only limited sensitivity and specificity in patients with CKD for prediction of bone marrow iron stores and erythropoietic response to iron supplementation16, 17, 18, 19, 20, 21, 40, 45 (Figures 1 and 2). Their utility is further compromised by substantial inter-patient variability unrelated to changes in iron store status.46 Evidence to support a recommendation for specific TSAT and ferritin levels at which iron therapy should be initiated or as ‘targets' for iron therapy is limited, with very few RCTs.16, 17, 18, 19, 20, 21 No iron intervention trials have been sufficiently powered or of long enough duration to assess long-term safety and no studies have addressed the clinical benefit, cost-effectiveness, and risk-benefit comparison of using different TSAT and ferritin levels for the diagnosis of iron deficiency or as a trigger for iron supplementation.

ntion trials have been sufficiently powered or of long enough duration to assess long-term safety and no studies have addressed the clinical benefit, cost-effectiveness, and risk-benefit comparison of using different TSAT and ferritin levels for the diagnosis of iron deficiency or as a trigger for iron supplementation. The Work Group sought to recommend iron targets that balance diagnostic sensitivity and specificity with assumptions regarding safety. Previous clinical practice recommendations (Kidney Diseae Outcomes Quality Initiative [KDOQI] 2006 and others), largely opinion-based, indicated that supplemental iron should be administered to maintain ferritin levels >200 ng/ml (>200 μg/l) in CKD 5HD patients and >100 ng/ml (>100 μg/l) in CKD ND and CKD 5PD with TSAT >20% in all CKD patients. These guidelines also indicated that there was insufficient evidence to recommend routine IV iron administration when the ferritin level was >500 ng/ml (>500 μg/l).

to maintain ferritin levels >200 ng/ml (>200 μg/l) in CKD 5HD patients and >100 ng/ml (>100 μg/l) in CKD ND and CKD 5PD with TSAT >20% in all CKD patients. These guidelines also indicated that there was insufficient evidence to recommend routine IV iron administration when the ferritin level was >500 ng/ml (>500 μg/l). Most CKD patients with serum ferritin levels >100 ng/ml (>100 μg/l) have normal bone marrow iron stores,16, 17, 18, 19, 20, 21 yet many such patients will also have an increase in Hb concentration and/or reduction in ESA dose if supplemental iron is provided.16, 23, 30, 31, 40, 45 A substantial fraction of CKD patients with anemia and TSAT >20% respond to iron supplementation with an increase in Hb concentration and/or reduction in ESA dose. Therefore, for patients who have not been receiving iron supplementation, we suggest iron administration in anemic CKD patients with TSAT <30% and serum ferritin <500 ng/ml (<500 μg/l) if an increase in Hb level is desired, particularly if intended to avoid transfusions and reduce anemia-related symptoms, and/or reduction in ESA dose, after consideration of the potential risks of iron administration. The safety of providing additional iron to intentionally maintain TSAT >30% and serum ferritin >500 ng/ml (>500 μg/l) has been studied in very few patients. We do not recommend routine use of iron supplementation in patients with TSAT >30% or serum ferritin >500 ng/ml (>500 μg/l) since, as stated above, the benefits and risks of doing so are inadequately studied. In all patients receiving iron, it is important to weigh both short-term and acute toxicities associated with iron therapy and exclude the presence of active infection (Recommendation 2.4) before embarking on a course of IV iron treatment.

e, as stated above, the benefits and risks of doing so are inadequately studied. In all patients receiving iron, it is important to weigh both short-term and acute toxicities associated with iron therapy and exclude the presence of active infection (Recommendation 2.4) before embarking on a course of IV iron treatment. There is only very limited evidence in patients with CKD that informs any decision about defining any specific upper limits for iron status targets in guiding iron treatment.47, 48 Previous guidelines, such as the 2006 KDOQI guidelines and others, have specified serum ferritin levels above which additional IV iron therapy was generally not recommended,8, 49, 50, 51, 52 usually citing limits of 500–800 ng/ml (500–800 μg/l). However, no RCTs and few other studies have examined the efficacy and safety of providing IV iron to maintain ferritin levels >500–800 ng/ml (>500–800 μg/l). Most studies are retrospective and the few prospective studies have had small numbers of patients and short follow up, using surrogate outcomes such as Hb and ESA dose rather than more meaningful patient outcomes such as infection risk and mortality. In most patients with TSAT >30% or serum ferritin >500 ng/ml (>500 μg/l), any erythropoietic responsive to iron supplementation alone (i.e., the incremental change in Hb and/or reduction in ESA dose) will be small. In one RCT conducted in CKD 5HD patients with anemia, serum ferritin 500–1200 ng/ml (500–1200 μg/l), and TSAT<25%, patients received a 25% increase in epoetin dose and were randomly assigned to receive either no iron (control) or 1000 mg IV iron. At 6 weeks, Hb increased to a greater extent in the IV iron group.53 This study was not considered in the choice of target levels for ferritin and TSAT in this guideline in part because it studied only a restricted group of patients, all of whom also received an increase in ESA dose. The number of patients was too small and the period of observation too short to assess either clinically important outcomes or toxicity in a meaningful way (Supplementary Tables 2–4 online).

this guideline in part because it studied only a restricted group of patients, all of whom also received an increase in ESA dose. The number of patients was too small and the period of observation too short to assess either clinically important outcomes or toxicity in a meaningful way (Supplementary Tables 2–4 online). High ferritin levels in some studies have been associated with higher death rates, but whether elevation of ferritin levels is a marker of excessive iron administration rather than a nonspecific acute phase reactant is not clear. At increasingly higher ferritin levels, there is some evidence to indicate that hepatic deposition of iron increases.54, 55 Clinical sequelae of this have not been documented although such hepatic iron deposition might be of particular concern in patients with hepatitis C virus (HCV) infection.56 While some data are available linking ferritin levels in patients with hemochromatosis and transfusional tissue iron deposition in patients without CKD,57 it is not clear to what extent these findings are relevant to CKD patients or should be used to guide clinical practice in CKD patients.

s C virus (HCV) infection.56 While some data are available linking ferritin levels in patients with hemochromatosis and transfusional tissue iron deposition in patients without CKD,57 it is not clear to what extent these findings are relevant to CKD patients or should be used to guide clinical practice in CKD patients. Rather than focusing on serum ferritin levels as a predictor of outcomes, some observational studies have examined associations between patient outcomes and amount of iron administered. One such study found no adverse association between 2-year survival when the IV iron dose over 6 months was ≤1000 mg, but a statistically significant higher mortality for iron doses >1000 mg (adjusted hazards ratio [HR] 1.09; 95% confidence interval [CI] 1.01–1.17 for > 1000 mg to 1800 mg and 1.18; 95% CI 1.09–1.27 for > 1800 mg).33 However, after using multivariable models accounting for time-varying measures of iron administration and other parameters, there was no statistically significant association between any level of iron administration and mortality. Another retrospective study using time-dependent and multivariate adjustment for case mix found that IV iron doses up to 400 mg/month were associated with lower death rates compared to doses >400 mg/month35 (Supplementary Table 5 online).

y significant association between any level of iron administration and mortality. Another retrospective study using time-dependent and multivariate adjustment for case mix found that IV iron doses up to 400 mg/month were associated with lower death rates compared to doses >400 mg/month35 (Supplementary Table 5 online). It is the consensus of the Work Group that additional IV iron should not routinely be administered in patients with serum ferritin levels that are consistently >500 ng/ml (>500 μg/l). In patients with Hb below the desired level who are receiving relatively high ESA doses, or in whom discontinuation of ESA therapy is preferred (for instance a CKD patient with malignancy), a therapeutic trial of additional IV iron (i.e., a single course of up to 1000 mg over a period of several weeks which can be repeated as needed) may be undertaken in patients with serum ferritin levels >500 ng/ml (>500 μg/l) after due consideration of potential acute toxicities and long-term risks. Subsequent treatment decisions should be based on the patient's clinical status, including trends in TSAT, ferritin, and Hb level, and ESA dose and responsiveness.

) may be undertaken in patients with serum ferritin levels >500 ng/ml (>500 μg/l) after due consideration of potential acute toxicities and long-term risks. Subsequent treatment decisions should be based on the patient's clinical status, including trends in TSAT, ferritin, and Hb level, and ESA dose and responsiveness. Ferritin levels need to be interpreted with caution in patients who may have an underlying inflammatory condition as they may not predict iron stores or responsiveness to iron therapy in a manner similar to that when inflammation is absent. In the absence of a clinically evident infectious or inflammatory process, assessment of CRP may suggest the presence of an occult inflammatory state that may be associated with an elevated ferritin level and ESA-hyporesponsiveness (Supplementary Table 6 online). Other iron status tests not as widely available as TSAT and ferritin such as percentage of hypochromic red cells, reticulocyte Hb content, zinc protoporphyrin, and soluble transferrin receptors may be used to assess iron status, but are less well studied.22, 23 There is no evidence that a higher ferritin target of 200ng/ml (200 μg/l) is the appropriate or inappropriate cutoff in CKD 5HD pediatric patients. Consequently no change has been made to the 2006 KDOQI guideline in children with CKD and anemia, which recommended a ferritin target greater than 100 ng/ml (100 μg/l) for CKD 5HD, as well as for CKD 5PD and CKD ND who are not on ESA therapy.58

) is the appropriate or inappropriate cutoff in CKD 5HD pediatric patients. Consequently no change has been made to the 2006 KDOQI guideline in children with CKD and anemia, which recommended a ferritin target greater than 100 ng/ml (100 μg/l) for CKD 5HD, as well as for CKD 5PD and CKD ND who are not on ESA therapy.58 Iron treatment A decision to provide an individual patient with iron therapy should be based on an assessment that an increase in Hb level is desirable, that is, to avoid transfusions or reduce anemia-related symptoms, and that the potential adverse effects of iron supplementation, either oral or IV, have been considered and are appropriately outweighed by the expected treatment benefit. Such supplementation could be in the form of oral or intravenous iron. Use of intramuscular iron has largely been abandoned. Each route has its own potential advantages and disadvantages. Oral iron is inexpensive, readily available, and does not require IV access, a particular concern in CKD patients not on HD. It is also not associated with severe adverse effects but gastrointestinal side effects are common and may limit adherence. This, along with variable gastrointestinal tract absorption, limits the efficacy of oral iron. IV iron avoids concerns about medication adherence and efficacy in treating iron deficiency, but requires IV access and has been associated with infrequent but severe adverse reactions. Decisions about the preferred route of iron supplementation should take into consideration severity of anemia and iron deficiency, the response, tolerance and adherence to prior oral iron administration, costs, and ease of obtaining venous access balanced against the desire to preserve venous access sites.

e reactions. Decisions about the preferred route of iron supplementation should take into consideration severity of anemia and iron deficiency, the response, tolerance and adherence to prior oral iron administration, costs, and ease of obtaining venous access balanced against the desire to preserve venous access sites. In patients with CKD ND, the available evidence supports an efficacy advantage of IV compared with oral administration of iron although the effect is rather small, with a weighted mean Hb difference of 0.31 g/dl (3.1 g/l).45, 59, 60, 61, 62, 63 Whether the small Hb benefit of IV iron in CKD ND patients is clinically meaningful or justifies the small risk of serious adverse events and unknown long-term risks is uncertain. The consensus of the Work Group is that a clearly defined advantage or preference for IV compared to oral iron was not supported by available evidence in CKD ND patients. Therefore, in such patients, the route of iron administration can be either IV or oral. In some patients the desire to avoid venipuncture (and preserve IV access) may favor in some patients, particularly those with mild iron deficiency, an initial trial of oral iron.

supported by available evidence in CKD ND patients. Therefore, in such patients, the route of iron administration can be either IV or oral. In some patients the desire to avoid venipuncture (and preserve IV access) may favor in some patients, particularly those with mild iron deficiency, an initial trial of oral iron. Oral iron is typically prescribed to provide approximately 200 mg of elemental iron daily (for instance ferrous sulfate 325 mg three times daily; each pill provides 65 mg elemental iron). Smaller daily doses may be useful and better tolerated in some patients. Although ferrous sulfate is commonly available and inexpensive, other oral iron preparations may also be used; there is not significant evidence to suggest that other oral iron formulations are more effective or associated with fewer adverse side effects than ferrous sulfate. If the goals of iron supplementation are not met with a 1–3 month course of oral iron, it is appropriate to consider IV iron supplementation in a manner consistent with the above recommendation statements and the discussion that follows.

more effective or associated with fewer adverse side effects than ferrous sulfate. If the goals of iron supplementation are not met with a 1–3 month course of oral iron, it is appropriate to consider IV iron supplementation in a manner consistent with the above recommendation statements and the discussion that follows. There is evidence supporting a preference for the IV route of iron administration in CKD 5HD patients derived from RCTs and other studies comparing IV iron with oral iron and placebo, with and without concomitant ESA treatment.27, 32, 62, 64, 65 In most of these studies, IV iron administration led to a greater increase in Hb concentration, a lower ESA dose, or both. In CKD 5HD patients, the ready IV access and convenience of being able to administer IV iron during HD treatments further supports the preference for the IV route for iron administration in these patients. In prior CKD anemia guidelines,50 CKD 5PD patients were considered more similar to CKD ND than CKD 5HD in their need for and likely responsiveness to iron, as well as in their absence of ready venous access for IV iron administration. Limited studies of iron administration in CKD 5PD patients indicate that oral iron is of limited efficacy and that IV iron is superior to oral iron in terms of achieved Hb level and ESA dose. Consequently, this route is preferred in these patients, although the desire to preserve potential future venous access sites must be considered in such patients.66, 67, 68, 69, 70

tients indicate that oral iron is of limited efficacy and that IV iron is superior to oral iron in terms of achieved Hb level and ESA dose. Consequently, this route is preferred in these patients, although the desire to preserve potential future venous access sites must be considered in such patients.66, 67, 68, 69, 70 IV iron may be provided as a single large dose or as repeated smaller doses depending on the specific IV iron preparation used (with the highest single dose varying by specific formulation). It is common practice to provide an initial course of IV iron amounting to approximately 1000 mg; this may be repeated if an initial dose fails to increase Hb level and/or allow a decrease in ESA dose and if the TSAT remains ≤30% and serum ferritin remains ≤500 ng/ml (≤500 μg/l).38

ose varying by specific formulation). It is common practice to provide an initial course of IV iron amounting to approximately 1000 mg; this may be repeated if an initial dose fails to increase Hb level and/or allow a decrease in ESA dose and if the TSAT remains ≤30% and serum ferritin remains ≤500 ng/ml (≤500 μg/l).38 Decisions regarding continued iron therapy should take into consideration recent patient responses to iron therapy, iron status tests (TSAT and ferritin), Hb concentration, ESA responsiveness and ESA dose in ESA-treated patients, ongoing blood losses, trends in each parameter, and the patient's clinical status. Serum ferritin and TSAT levels should not be measured until at least one week has elapsed since the most recent prior IV iron dose. Consideration of expected iron needs and evaluation for ongoing iron losses should precede further IV iron administration. Blood loss should be minimal in CKD ND and CKD 5PD patients, while CKD 5HD patients have reported to lose between 1–2 gm of iron per year related to the HD procedure and related circumstances.71, 72, 73 Thus, an apparent ongoing need for any iron supplementation in CKD ND and CKD 5PD patients or for more than 1–2 gm/yr in CKD 5HD patients should prompt assessment for a source of active blood loss. The need to consider trends in iron status tests are highlighted by consideration of a patient with decreasing TSAT and ferritin levels which may signify the presence of gastrointestinal bleeding or excessive dialysis-associated blood loss. As another example, an increasing TSAT and ferritin level may indicate excessive iron supplementation and a need to decrease or discontinue iron administration. Finally, an increase in ferritin level accompanied by a decrease in TSAT and Hb level suggests inflammation-mediated reticuloendothelial blockade.14

lood loss. As another example, an increasing TSAT and ferritin level may indicate excessive iron supplementation and a need to decrease or discontinue iron administration. Finally, an increase in ferritin level accompanied by a decrease in TSAT and Hb level suggests inflammation-mediated reticuloendothelial blockade.14 There are two commonly used approaches to ongoing or maintenance IV iron treatment in CKD 5HD patients: (1) periodic iron repletion, consisting of a series of IV iron doses administered episodically to replenish iron stores whenever iron status tests indicate the likelihood of iron deficiency or decrease below specific target levels; or (2) maintenance treatment, consisting of smaller doses administered at regular intervals to maintain iron status tests stable within specific limits with the intent of avoiding iron deficiency or decline of iron test parameters below specific levels. Limited evidence suggests that regular maintenance IV iron administration in CKD 5HD is associated with use of lower ESA doses and may result in lower cumulative iron doses41, 74, 75 but these data are insufficient to support a recommendation favoring any particular IV iron dosing strategy in this patient population. By nature of the clinical encounters with CKD 5PD patients, IV iron supplementation is often provided at periodic (e.g., monthly) visits.

esult in lower cumulative iron doses41, 74, 75 but these data are insufficient to support a recommendation favoring any particular IV iron dosing strategy in this patient population. By nature of the clinical encounters with CKD 5PD patients, IV iron supplementation is often provided at periodic (e.g., monthly) visits. Overall, the TSAT and ferritin recommendations above are applicable to children with CKD on ESA therapy. However, there is no evidence that a higher ferritin target of 200 ng/ml (200 μg/l) is the appropriate or inappropriate cutoff in pediatric CKD HD patients. Consequently no change has been made to the 2006 KDOQI guideline in CKD in children with anemia, which recommended a ferritin target greater than 100 ng/ml (100 μg/l) for CKD 5HD, as well as for CKD 5PD and CKD ND who are on ESA therapy.58 IRON STATUS EVALUATION 2.2.1: Evaluate iron status (TSAT and ferritin) at least every 3 months during ESA therapy, including the decision to start or continue iron therapy. (Not Graded) 2.2.2: Test iron status (TSAT and ferritin) more frequently when initiating or increasing ESA dose, when there is blood loss, when monitoring response after a course of IV iron, and in other circumstances where iron stores may become depleted. (Not Graded)

IRON STATUS EVALUATION 2.2.1: Evaluate iron status (TSAT and ferritin) at least every 3 months during ESA therapy, including the decision to start or continue iron therapy. (Not Graded) 2.2.2: Test iron status (TSAT and ferritin) more frequently when initiating or increasing ESA dose, when there is blood loss, when monitoring response after a course of IV iron, and in other circumstances where iron stores may become depleted. (Not Graded) RATIONALE In the absence of clinical trials that specifically inform the optimal frequency for testing of iron status, and consistent with prior guidelines,50 the consensus of the Work Group is that patients who are on ESA therapy, regardless of whether iron treatment is also being used, should have tests of iron status at least every 3 months. Falling TSAT and/or ferritin levels are likely to reflect ongoing blood loss or consumption of available iron stores, and can be used to anticipate the need for future or additional iron supplementation. In patients on oral iron treatment, iron status testing can also be used to assess adherence with iron treatment. Increasing TSAT and/or ferritin levels may indicate that iron treatment is excessive and can be stopped or reduced. Increasing ferritin levels in association with stable or declining TSAT levels may also indicate the presence of inflammation, infection, or other clinical situations inducing acute phase reactants during which time the appropriateness of continued iron administration may need to be reassessed.14

be stopped or reduced. Increasing ferritin levels in association with stable or declining TSAT levels may also indicate the presence of inflammation, infection, or other clinical situations inducing acute phase reactants during which time the appropriateness of continued iron administration may need to be reassessed.14 In some circumstances, more frequent iron status testing may be appropriate, including following initiation of ESA or iron therapy or when the ESA dose or dose frequency is increased. Iron status testing is also important in the assessment of patients who become less responsive to ESA treatment.

be stopped or reduced. Increasing ferritin levels in association with stable or declining TSAT levels may also indicate the presence of inflammation, infection, or other clinical situations inducing acute phase reactants during which time the appropriateness of continued iron administration may need to be reassessed.14 In some circumstances, more frequent iron status testing may be appropriate, including following initiation of ESA or iron therapy or when the ESA dose or dose frequency is increased. Iron status testing is also important in the assessment of patients who become less responsive to ESA treatment. Despite the absence of specific data in the pediatric CKD population, this recommendation is considered applicable to children since there are no reasons to suggest a different recommendation. Since the 2006 KDOQI guideline for anemia in pediatric CKD,58 no new evidence regarding iron therapy for children with CKD has been published. The suggestion for oral iron supplementation in children is 2–6 mg/kg/day of elemental iron in 2–3 divided doses.76, 77 An RCT of 35 iron replete pediatric CKD 5HD patients evaluated their response to either weekly IV iron dextran dosed by weight or oral iron 6 mg/kg/day. Only the IV iron dextran produced a significant increase in the serum ferritin levels and showed a significant decrease in ESA dose required to maintain target Hb levels.78 An international multicenter double-blind RCT investigated the safety and efficacy of two dosing regimens (1.5 mg/kg or 3 mg/kg) of ferric gluconate in iron-deficient pediatric hemodialysis patients receiving concomitant ESA therapy. Efficacy and safety profiles were comparable, with no unexpected adverse events with either dose.79 Based on this trial, the recommendation for initial ferric gluconate therapy is 1.5 mg/kg for eight doses for iron-deficient pediatric CKD 5HD patients and 1 mg/kg per week for iron-replete pediatric CKD 5HD patients, with subsequent dose adjustments made according to TSAT and/or ferritin levels.79, 80 Iron sucrose has also been used in children with CKD81 but, as of yet, no RCTs have been published in this population. Although it is not uncommon that pediatric CKD 5PD and CKD ND patients either do not respond to or tolerate oral iron therapy, the need for IV access for parenteral iron therapy often limits its utilization in children.

lso been used in children with CKD81 but, as of yet, no RCTs have been published in this population. Although it is not uncommon that pediatric CKD 5PD and CKD ND patients either do not respond to or tolerate oral iron therapy, the need for IV access for parenteral iron therapy often limits its utilization in children. CAUTIONS REGARDING IRON THERAPY 2.3: When the initial dose of IV iron dextran is administered, we recommend (1B) and when the initial dose of IV non-dextran iron is administered, we suggest (2C) that patients be monitored for 60 minutes after the infusion, and that resuscitative facilities (including medications) and personnel trained to evaluate and treat serious adverse reactions be available. RATIONALE Any form of IV iron may be associated with potentially severe acute reactions.82, 83, 84, 85, 86, 87, 88, 89, 90, 91 The symptoms of most concern are hypotension and dyspnea, which in the worst cases may be catastrophic with features of anaphylaxis. The cause of reactions has not been fully characterized, but may involve immune mechanisms and/or release of free, reactive iron into the circulation with induction of oxidative stress. The mechanisms of acute reactions may differ for different iron preparations. Certain iron dextrans in particular have been associated with reactions characteristic of anaphylaxis. The rate of such reactions is estimated to occur in 0.6–0.7% of patients treated. The serious adverse effect event rate may be lower with low molecular weight iron dextran compared to high molecular weight iron dextran.92, 93, 94, 95, 96

s in particular have been associated with reactions characteristic of anaphylaxis. The rate of such reactions is estimated to occur in 0.6–0.7% of patients treated. The serious adverse effect event rate may be lower with low molecular weight iron dextran compared to high molecular weight iron dextran.92, 93, 94, 95, 96 With non-dextran IV iron drugs, it is believed that anaphylactoid and other severe and potentially life-threatening reactions are less common, but this has not been well substantiated. Serious reactions including profound hypotension do occur, even if uncommonly, with all non-dextran IV iron preparations. Because all forms of IV iron drugs can be associated with serious immediate reactions, they should be used with vigilance. Since the rate of such reactions may be greater for iron dextran drugs we recommend that resuscitative medications and personnel trained to evaluate and treat serious adverse reactions be available when the initial dose of IV iron dextran is administered. The data to support such a recommendation for the initial dose of non-iron dextran compounds is not as strong. In the US, the Food and Drug Administration (FDA)-mandated labeling for ferumoxytol specifies that patients be observed for 60 minutes after administration. This may be reasonable advice for all IV iron drugs, including other new iron preparations such ferric carboxymaltose and iron isomaltoside. For each IV iron preparation prescribing physicians should be familiar with the drug's safety and toxicity profile and the product labeling warnings and recommendations for administration, as well as patient monitoring during and after treatment.

r new iron preparations such ferric carboxymaltose and iron isomaltoside. For each IV iron preparation prescribing physicians should be familiar with the drug's safety and toxicity profile and the product labeling warnings and recommendations for administration, as well as patient monitoring during and after treatment. Iron during infection 2.4: Avoid administering IV iron to patients with active systemic infections. (Not Graded)

r new iron preparations such ferric carboxymaltose and iron isomaltoside. For each IV iron preparation prescribing physicians should be familiar with the drug's safety and toxicity profile and the product labeling warnings and recommendations for administration, as well as patient monitoring during and after treatment. Iron during infection 2.4: Avoid administering IV iron to patients with active systemic infections. (Not Graded) RATIONALE Iron is essential for the growth and proliferation of most pathogens including many bacteria, viruses, fungi, parasites and helminthes, and also exerts subtle effects on immune function and host responses towards microbes.97 There is theoretical and experimental evidence to suggest that iron administration may worsen an existing infection but clinical evidence is lacking. In animal models, iron overload results in an impaired control of infections, specifically with intracellular bacteria or fungi.98, 99, 100, 101 In humans, tissue iron overload has been considered as a risk factor for the acquisition of certain infections and for an unfavorable clinical course of the infection. Data in CKD patients are conflicting.102, 103, 104 Since current evidence cannot provide a clear answer as to whether specific CKD patient groups are at increased risk for infection, or of having a poorer outcome with infection when anemia is treated with IV iron, the Work Group suggests that IV iron not be administered when patients have an active systemic infection. Clinical judgment is necessary in each individual patient to assess whether there is an immediate need for IV iron (as opposed to delaying treatment until resolution of an infection), likelihood of achieving benefit from a dose of IV iron in the setting of an active infection, and the severity of an infection.

fection. Clinical judgment is necessary in each individual patient to assess whether there is an immediate need for IV iron (as opposed to delaying treatment until resolution of an infection), likelihood of achieving benefit from a dose of IV iron in the setting of an active infection, and the severity of an infection. RESEARCH RECOMMENDATIONS Much regarding the testing of iron status and use of iron supplementation, particularly IV, in CKD patients of all stages remains unknown. There is a serious lack of large, prospective clinical trials with assessment of clinically meaningful outcomes and toxicities; rather, most have been small, short-term studies focusing primarily on surrogate outcomes such as increase in Hb level and reduction in ESA dose. Some important questions that should be addressed in future studies might include: What is the comparative risk-benefit balance of various treatment strategies that include differing ratios of ESA dosing and iron supplementation to achieve a particular Hb level? Is there a role, and if so under what circumstances, for anemia management in CKD patients with iron alone, without ESA treatment (or with only ESA ‘rescue therapy' for particularly low Hb levels)? Is there important long-term toxicity of IV iron supplementation and if so, under what circumstances and in what CKD patient groups? Is IV iron administration, with or without concomitant ESA dose increases, safe and of clinical benefit, in patients with ferritin levels >500–800 ng/ml (>500–800 μg/l)?

Is there a role, and if so under what circumstances, for anemia management in CKD patients with iron alone, without ESA treatment (or with only ESA ‘rescue therapy' for particularly low Hb levels)? Is there important long-term toxicity of IV iron supplementation and if so, under what circumstances and in what CKD patient groups? Is IV iron administration, with or without concomitant ESA dose increases, safe and of clinical benefit, in patients with ferritin levels >500–800 ng/ml (>500–800 μg/l)? What are the best laboratory tests to guide decisions regarding initiation, ongoing treatment, and discontinuation of iron supplementation? Is current iron and anemia management in pediatric CKD patients appropriate?

Is IV iron administration, with or without concomitant ESA dose increases, safe and of clinical benefit, in patients with ferritin levels >500–800 ng/ml (>500–800 μg/l)? What are the best laboratory tests to guide decisions regarding initiation, ongoing treatment, and discontinuation of iron supplementation? Is current iron and anemia management in pediatric CKD patients appropriate? DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. SUPPLEMENTARY MATERIAL Supplemental Table 2: Summary table of RCT examining the effect of IV iron + EPO vs. EPO only in patients with HD-CKD (categorical outcomes). Supplemental Table 3: Summary table of RCT examining the effect of IV iron + EPO vs. EPO only in patients with HD-CKD (continuous outcomes).

DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. SUPPLEMENTARY MATERIAL Supplemental Table 2: Summary table of RCT examining the effect of IV iron + EPO vs. EPO only in patients with HD-CKD (categorical outcomes). Supplemental Table 3: Summary table of RCT examining the effect of IV iron + EPO vs. EPO only in patients with HD-CKD (continuous outcomes). Supplemental Table 4: Summary table of adverse events in RCT examining the effect of IV iron + EPO vs. EPO only in patients with HD-CKD (continuous outcomes). Supplemental Table 5: Association between cumulative iron dose and clinical outcome in multivariable analyses. Supplemental Table 6: Association between iron status and clinical outcome in multivariable analyses.

Supplemental Table 4: Summary table of adverse events in RCT examining the effect of IV iron + EPO vs. EPO only in patients with HD-CKD (continuous outcomes). Supplemental Table 5: Association between cumulative iron dose and clinical outcome in multivariable analyses. Supplemental Table 6: Association between iron status and clinical outcome in multivariable analyses. Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php * Based on patient symptoms and overall clinical goals, including avoidance of transfusion, improvement in anemia-related symptoms, and after exclusion of active infection. ** Consistent with Recommendations #3.4.2 and 3.4.3. *** Based on patient symptoms and overall clinical goals including avoidance of transfusion and improvement in anemia-related symptoms, and after exclusion of active infection and other causes of ESA hyporesponsiveness. Figure 1 Receiver operating characteristic (ROC) curves, examining the utility of iron status tests to distinguish iron deficient from nondeficient study patients. Reprinted with permission from Macmillan Publishers Ltd: Kidney International. Van Wyck DB, Roppolo M, Martinez CO et al. A randomized, controlled trial comparing IV iron sucrose to oral iron in anemic patients with nondialysis-dependent CKD. Kidney Int 2005; 68: 2846–2856;45 accessed http://www.nature.com/ki/journal/v68/n6/full/4495631a.html.

from Macmillan Publishers Ltd: Kidney International. Van Wyck DB, Roppolo M, Martinez CO et al. A randomized, controlled trial comparing IV iron sucrose to oral iron in anemic patients with nondialysis-dependent CKD. Kidney Int 2005; 68: 2846–2856;45 accessed http://www.nature.com/ki/journal/v68/n6/full/4495631a.html. Figure 2 Sensitivity and specificity of TSAT and serum ferritin (ferritin) and their combination (TSAT + ferritin) and bone marrow iron (BM iron) to identify correctly a positive erythropoietic response (≥1-g/dl [≥10-g/l] increase in Hb [ΔHb]) to intravenous iron in 100 nondialysis patients with CKD (areas under the ROCs). Reproduced with permission from American Society of Nephrology40 from Stancu S, Barsan L, Stanciu A et al. Can the response to iron therapy be predicted in anemic nondialysis patients with chronic kidney disease? Clin J Am Soc Nephrol 2010; 5: 409–416; permission conveyed through Copyright Clearance Center; accessed http: http://cjasn.asnjournals.org/content/5/3/409.long

ESA INITIATION BACKGROUND The introduction of recombinant human erythropoietin (rHuEPO) into clinical practice in the 1980 s was a major breakthrough in the treatment of the anemia of patients with CKD. The development of rHuEPO was aimed at replacing the insufficient endogenous erythropoietin (EPO) production related to CKD progression. It remains unclear whether the main cause of anemia is a loss of kidney EPO production capacity or a derangement in oxygen sensing, as proposed more recently.105 In the early years, rHuEPO administration was regarded by the nephrology community as a beneficial therapy for long-term dialysis patients whose Hb values fell to extremely low levels, making them transfusion-dependent. The immediate benefit of rHuEPO in CKD patients with severe anemia and anemia-related signs and symptoms was clear. In addition, the reduction in the need for regular blood transfusions was another major benefit, resulting in less frequent transmission of blood-borne viral diseases, such as hepatitis B and C, less allosensitization, predisposing to prolonged wait times or failure to receive a kidney transplant, transplant rejection, and less transfusional hemosiderosis.106, 107, 108, 109

transfusions was another major benefit, resulting in less frequent transmission of blood-borne viral diseases, such as hepatitis B and C, less allosensitization, predisposing to prolonged wait times or failure to receive a kidney transplant, transplant rejection, and less transfusional hemosiderosis.106, 107, 108, 109 After introduction of rHuEPO into clinical practice its administration was limited to dialysis patients with the most severe forms of anemia. Progressively, its use was extended to the majority of dialysis patients with renal anemia, and subsequently also to anemic patients with CKD 4–5 in countries in which the high cost of rHuEPO did not limit the number of patients eligible for this treatment.

ted to dialysis patients with the most severe forms of anemia. Progressively, its use was extended to the majority of dialysis patients with renal anemia, and subsequently also to anemic patients with CKD 4–5 in countries in which the high cost of rHuEPO did not limit the number of patients eligible for this treatment. Hb targets also increased progressively, often into the range of normal values. The idea that anemia should be corrected completely was based on pathophysiologic considerations and the demonstration by numerous observational studies of an inverse association between Hb concentrations up into the normal range and intermediate outcomes such as left ventricular hypertrophy,110 as well as hard patient outcomes such as cardiovascular events,111, 112, 113 hospital admission,114 and death.115, 116 Of note, a recent study also showed that CKD 5D patients with naturally occurring Hb concentrations greater than 12 g/dl (120 g/l) were not at increased mortality risk.117 However, the suggestion drawn from epidemiological studies that anemia should be completely corrected in patients with CKD was not supported by the Normal Hematocrit Study in CKD 5D patients118 and several recent randomized controlled trials (RCTs) performed in large CKD patient cohorts (Supplementary Table 7 online).

However, the suggestion drawn from epidemiological studies that anemia should be completely corrected in patients with CKD was not supported by the Normal Hematocrit Study in CKD 5D patients118 and several recent randomized controlled trials (RCTs) performed in large CKD patient cohorts (Supplementary Table 7 online). In CKD 5D patients Hb concentrations often fall below 8 g/dl (80 g/l) if anemia is untreated, whereas in CKD ND patients higher Hb concentrations are usual, unless patients are close to dialysis or have another contributing cause. The decision to prescribe ESAs should be based on evidence accrued from RCTs. However substantial heterogeneity exists in RCTs performed to evaluate ESA therapy, particularly in relation to classification of patients, research design, baseline Hb, target Hb, clinical outcome measures, and definitions of clinically meaningful improvements.

cribe ESAs should be based on evidence accrued from RCTs. However substantial heterogeneity exists in RCTs performed to evaluate ESA therapy, particularly in relation to classification of patients, research design, baseline Hb, target Hb, clinical outcome measures, and definitions of clinically meaningful improvements. Outcomes of interest in RCTs of ESAs include mortality, cardiovascular and kidney endpoints, safety, quality of life (QoL), blood transfusions and cost. QoL outcomes are particularly important for CKD 5D patients and for some may be more important than cardiovascular events or mortality, since they have relatively short life expectancy and the symptoms attributable to anemia (e.g., low energy, fatigue, decreased physical function, and low exercise capacity) occur frequently and can be disabling.119 However, QoL is extremely difficult to quantify as is the clinical importance of changes measured. Furthermore, unless assessed under rigorous double-blind conditions, the validity of QoL measurements is questionable. Avoidance of transfusions is important, as mentioned above.

occur frequently and can be disabling.119 However, QoL is extremely difficult to quantify as is the clinical importance of changes measured. Furthermore, unless assessed under rigorous double-blind conditions, the validity of QoL measurements is questionable. Avoidance of transfusions is important, as mentioned above. The guidelines to treat or not to treat the anemia of CKD are also valid for CKD 4–5T patients. Of note, blood transfusions may increase the risk of alloreactivity and rejection episodes after kidney transplantation.120 In addition a recent randomized trial has shown that early post-kidney transplant anemia correction by ESAs reduces the progression of allograft nephropathy, although its effect on hard outcomes in this patient population remains unknown.121 3.1: Address all correctable causes of anemia (including iron deficiency and inflammatory states) prior to initiation of ESA therapy. (Not Graded) RATIONALE After diagnosing anemia in a patient with CKD all correctable causes should be treated before considering ESA therapy. Above all, this recommendation is based on the observation that iron supplementation given to CKD patients with proven iron deficiency or impaired iron availability (‘functional iron deficiency') generally leads to an increase in Hb (See Chapter 2). However, the correction of other deficiency states also may ameliorate anemia. In patients with inflammatory diseases, including bacterial and viral infections, the attenuation of the inflammatory status is often followed by an improvement of Hb.

l iron deficiency') generally leads to an increase in Hb (See Chapter 2). However, the correction of other deficiency states also may ameliorate anemia. In patients with inflammatory diseases, including bacterial and viral infections, the attenuation of the inflammatory status is often followed by an improvement of Hb. There are several reasons why correctable causes other than erythropoietin deficiency should be actively sought. As in any disease state, pathological conditions which can be cured should be corrected first. As examples, ESA treatment is unlikely to be fully effective in raising Hb concentrations until either severe systemic bacterial infections or severe secondary hyperparathyroidism are appropriately treated (Supplementary Table 8 online). When several different factors are thought to contribute to the anemia of CKD, even though the main underlying cause is impaired kidney EPO synthesis, appropriate medical care dictates treating all underlying causes. 3.2: In initiating and maintaining ESA therapy, we recommend balancing the potential benefits of reducing blood transfusions and anemia-related symptoms against the risks of harm in individual patients (e.g., stroke, vascular access loss, hypertension). (1B) RATIONALE Treatment of severe anemia Objective evidence to support treatment of Hb concentrations below 9 g/dl (90 g/l) is quite strong because the transfusion benefits are substantial and the QoL improvements are clinically important. However the safety of ESAs in treating severe anemia has not been evaluated in large placebo controlled trials.

re anemia Objective evidence to support treatment of Hb concentrations below 9 g/dl (90 g/l) is quite strong because the transfusion benefits are substantial and the QoL improvements are clinically important. However the safety of ESAs in treating severe anemia has not been evaluated in large placebo controlled trials. The Canadian Erythropoietin Study Group reported a double-blind RCT of 118 CKD 5HD patients in 1990. ESA was utilized in patients with Hb concentrations <9 g/dl (<90 g/l), and three randomly allocated groups were followed (placebo, target Hb 9.5–11 g/dl [95–110 g/l], high target Hb >11 g/dl [>110 g/l]).122 Baseline Hb was 7.0 g/dl (70 g/l) and the mean transfusion requirement was 7 transfusions per year. After 8 weeks, 58% (N=23/40) in the placebo group were transfused and only 2.5% (N=1/40) was transfused in the group with target Hb of 9.5–11g/dl (95–110 g/l) and 2.6% (N=1/38) in the group with target Hb>11g/dl (>110 g/l). After 6 months, significant improvements in fatigue, physical function, and 6 minute walking tests were reported for the low Hb group compared to placebo, but no improvement was observed comparing low vs high Hb group. In an open-label RCT of only 83 CKD ND patients with Hb <10 g/dl (<100 g/l), significant improvements in energy and physical function were also reported.123

ysical function, and 6 minute walking tests were reported for the low Hb group compared to placebo, but no improvement was observed comparing low vs high Hb group. In an open-label RCT of only 83 CKD ND patients with Hb <10 g/dl (<100 g/l), significant improvements in energy and physical function were also reported.123 Treatment of moderate anemia There are several large RCTs of ESA therapy where baseline Hb is >10 g/dl (>100 g/l). 118, 124, 125, 126, 127, 128 The intervention being tested in these trials is complete correction of anemia with ESAs, compared to partial correction with ESAs in five RCTs118, 124, 125, 126, 128 and to placebo in one.127 A double-blind design is necessary to accurately assess subjective or clinician-driven endpoints particularly QoL, starting dialysis, and giving transfusions. Notably, only 3 of the 6 trials were double-blind – the Normal Hematocrit Study reported in 1998,118 the Canada-Europe Study reported in 2005,126 and TREAT reported in 2009.127 The Scandinavian Study,125 CREATE124 and CHOIR128 trials were open label.

dpoints particularly QoL, starting dialysis, and giving transfusions. Notably, only 3 of the 6 trials were double-blind – the Normal Hematocrit Study reported in 1998,118 the Canada-Europe Study reported in 2005,126 and TREAT reported in 2009.127 The Scandinavian Study,125 CREATE124 and CHOIR128 trials were open label. The US Normal Hematocrit Trial by Besarab et al.118 was the first of a series of RCTs which cast serious doubt on the assumption that full anemia correction should be achieved in the majority of dialysis patients. A cohort of 1233 prevalent CKD 5HD patients with symptomatic heart failure or ischemic heart disease were allocated to either partial treatment of anemia or full anemia correction, using epoetin-alfa. The eventually achieved hematocrit values were 31% and 40%, respectively. In the normal hematocrit group treated with epoetin there were 183 deaths and 19 myocardial infarcts, producing 202 primary events, compared to 164 events (150 deaths, 14 myocardial infarcts) in the group in which anemia was partially corrected with epoetin. The risk ratio for the primary endpoint was 1.3 (95% CI 0.9–1.9) which did not satisfy the pre-specified criterion for statistical significance (even though the nominal p value was 0.03) after adjusting for interim analyses. The trial was stopped early in a situation where the primary hypothesis was unlikely to be proven and the intervention being tested caused harm: 39% had vascular access clotting in the intervention arm and 29% in the control arm (P=0.001).

(even though the nominal p value was 0.03) after adjusting for interim analyses. The trial was stopped early in a situation where the primary hypothesis was unlikely to be proven and the intervention being tested caused harm: 39% had vascular access clotting in the intervention arm and 29% in the control arm (P=0.001). The double-blind Canada-Europe trial by Parfrey et al.126 of 596 incident CKD 5HD patients without symptomatic heart disease (18% with diabetic nephropathy) examined the question whether full anemia correction by epoetin-alfa in the group randomized to a Hb target of 13.5–14.5 g/dl (135–145 g/l), as compared to partial treatment of anemia in the group randomized to a Hb target of 9.5–11.5 g/dl (95–115 g/l), had a beneficial effect on left ventricular volume and mass index. The eventually achieved Hb values were 13.1 and 10.8 g/dl (131 and 108 g/l), respectively. There was no difference in left ventricular volume index or mass index between the two groups during this 96-week study. Of note, patients in the full anemia correction group had a significantly higher stroke incidence (secondary endpoint) than patients in the partial treatment correction group. However, the absolute numbers of patients with stroke were very small. As one might expect, the high Hb group received significantly fewer transfusions than the low Hb group, but extent of the benefit was modest: although 9% in the high Hb arm received at least one transfusion compared to 19% in the low Hb arm (P=0.004) during the 96-week study, the transfusions per patient per year was 0.3 in the high Hb arm and 0.7 in the low Hb arm (P<0.0001).129 In addition significant improvements in QoL were reported for the a priori selected domains of vitality and of fatigue.126, 130

one transfusion compared to 19% in the low Hb arm (P=0.004) during the 96-week study, the transfusions per patient per year was 0.3 in the high Hb arm and 0.7 in the low Hb arm (P<0.0001).129 In addition significant improvements in QoL were reported for the a priori selected domains of vitality and of fatigue.126, 130 The goal of the CREATE study by Drueke et al.124 was to show superiority of full anemia correction in terms of cardiovascular events, as compared to partial correction of anemia, when starting ESA therapy at an earlier stage than end-stage renal disease (ESRD). In this trial, 603 CKD 3–5 patients (26% with diabetes) were randomly allocated to either a Hb target of 13.0–15.0 g/dl (130–150 g/l) or a Hb target of 10.5–11.5 g/dl (105–115 g/l) using epoetin-beta. The eventually achieved Hb values were 13.5 and 11.6 g/dl (135 and 116 g/l), respectively. Dialysis was required in significantly more patients in the high Hb group than in the low Hb group. However the rate of fall of GFR in the two groups during the 3 year study was similar. Statistically significant improvements in some domains of QoL, including physical function and vitality, were observed in the high Hb group, although these must be interpreted cautiously because the study was open-label.

ow Hb group. However the rate of fall of GFR in the two groups during the 3 year study was similar. Statistically significant improvements in some domains of QoL, including physical function and vitality, were observed in the high Hb group, although these must be interpreted cautiously because the study was open-label. The US CHOIR study by Singh et al.128 similarly aimed to show superiority of full anemia correction by ESA administration in terms of cardiovascular events and death, as compared to partial treatment of anemia, in patients with CKD not yet on dialysis. In this trial, 1432 CKD 3–4 patients (49% with diabetes) were randomized to Hb targets of 13.5 g/dl (135 g/l) and 11.3 g/dl (113 g/l) using epoetin-alfa. Withdrawal rate was high: 17% due to renal replacement therapy and 21% for other reasons. The study was prematurely stopped after an interim analysis with a median study duration of 16 months. The achieved Hb values were 12.6 and 11.3 g/dl (126 and 113 g/l), respectively. At this time point, 125 patients in the complete anemia correction group but only 97 patients in the standard correction group had reached the primary combined cardiovascular endpoint (P=0.03). No differences in QoL were observed comparing the two groups although, again, this finding must be interpreted cautiously because the study was open-label.

nts in the complete anemia correction group but only 97 patients in the standard correction group had reached the primary combined cardiovascular endpoint (P=0.03). No differences in QoL were observed comparing the two groups although, again, this finding must be interpreted cautiously because the study was open-label. Finally, the international trial of darbepoetin-alfa in type 2 diabetes and CKD (TREAT) by Pfeffer et al.127 examined cardiovascular and kidney outcomes in 4038 CKD 3–4 patients. Of note, this is by far the largest ESA trial, and has the best research design, as it was placebo controlled and double-blinded. Patients received either darbepoetin-alfa to achieve a Hb target of 13.0 g/dl (130 g/l) or placebo with rescue darbepoetin-alfa when the Hb concentration was <9.0 g/dl (<90 g/l). The achieved Hb values were 12.5 and 10.6 g/dl (125 and 106 g/l), respectively. The median follow-up duration of the study was 29 months. There were no differences in the two primary endpoints, which were the composite outcomes of death or a cardiovascular event (first primary endpoint) and death or ESRD (second primary endpoint). The hazard ratio for death/composite cardiovascular event was 1.05 (95% CI 0.94–1.17), and for death or ESRD it was 1.06 (95% CI 0.96–1.19). However there was a substantial increased risk of stroke (HR 1.92; 95% CI 1.38–2.68), although the absolute risk of stroke overall was modest: 5.0% of the high Hb group had a stroke compared to 2.6% in the placebo group (P<0.001). The relative increase in risk of stroke was similar in patients with and without a past history of stroke. As a result, the absolute risk of stroke was substantial in the 11% of subjects with a prior history of stroke; 12% in the darbepoetin group compared to 4% in the placebo group. Venous thrombo-embolic events occurred significantly more frequently in the high Hb arm (2.0%) compared to the placebo arm (1.1%, P=0.02). A signal that normalization of Hb with darbepoetin may be harmful in patients with a history of malignancy was reported following a post-hoc analysis: 14/188 (7.4%) of those with a history of malignancy at baseline died from cancer in the darbepoetin arm compared to 1/160 (0.6%) (P=0.002) in the placebo arm.

, P=0.02). A signal that normalization of Hb with darbepoetin may be harmful in patients with a history of malignancy was reported following a post-hoc analysis: 14/188 (7.4%) of those with a history of malignancy at baseline died from cancer in the darbepoetin arm compared to 1/160 (0.6%) (P=0.002) in the placebo arm. A statistically significant improvement in Functional Assessment of Cancer Therapy-Fatigue (FACT-fatigue) scores was reported at week 26 favoring the darbepoetin group, but the clinical significance of this was modest, as 55% of the high Hb group had a clinically important improvement in fatigue score compared to 50% of the placebo group. Transfusions were prescribed relatively frequently, and more often in the placebo arm (25%) compared to the high Hb arm (15%). The harm:benefit trade-off in TREAT was 1 stroke for 5 transfusions prevented by the high Hb target131 (Supplementary Tables 9–19 online). In a large subset of the TREAT patients QoL was assessed using FACT-fatigue, SF-36, and EQ-5D through 97 weeks. Compared to placebo, darbepoetin conferred a consistent, but small improvement over 97 weeks in fatigue and overall QoL, but none in energy and physical function. Interim stroke had a substantial negative impact on fatigue and physical function.132

QoL was assessed using FACT-fatigue, SF-36, and EQ-5D through 97 weeks. Compared to placebo, darbepoetin conferred a consistent, but small improvement over 97 weeks in fatigue and overall QoL, but none in energy and physical function. Interim stroke had a substantial negative impact on fatigue and physical function.132 Meta-analyses Assessment of ESAs in CKD using meta-analysis is problematic because of the heterogeneity of patients entered, the different quality and research designs of the RCTs performed, and differences in definitions of endpoints. In addition abstraction of aggregate data from the reports of RCTs to populate the meta-analysis data base is also a limitation, as individual patient data would be preferable. The most recent meta-analysis133 concluded that higher Hb concentrations in CKD increases risk for stroke (relative risk [RR] 1.51, 95% CI 1.03–2.21), hypertension (RR 1.67, 95% CI 1.31–2.12), and vascular access thrombosis (RR 1.33; 95% CI 1.16–1.53), and may perhaps increase risk for death (RR 1.09; 95% CI 0.99–1.20), serious cardiovascular events (RR 1.15, 95% CI 0.98–1.33) or ESRD (RR 1.08; 95% CI 0.97–1.20). In our opinion, because of the heterogeneity of patients and interventions across studies in the meta-analysis greater credence should be given to the results of the very large, placebo controlled, double-blind trial, TREAT, than to the meta-analyses, in areas where the results differ: TREAT found no difference between the higher Hb, darbepoetin, group and the lower Hb, placebo, group for the two primary composite outcomes (either death or a cardiovascular event, or death or a renal event).127

placebo controlled, double-blind trial, TREAT, than to the meta-analyses, in areas where the results differ: TREAT found no difference between the higher Hb, darbepoetin, group and the lower Hb, placebo, group for the two primary composite outcomes (either death or a cardiovascular event, or death or a renal event).127 The existing meta-analyses of QoL outcomes are further complicated by inclusion of data from open label studies, different instruments to measure QoL, differences in research design across RCTs, incomplete reporting as some trials chose (a priori) specific domains as trial outcomes, and differences in the definition of clinically meaningful improvement in QoL domains.119 Results from two systematic reviews published recently134, 135 suggest that improvements in QoL are maximized in the 10–12 g/dl (100–120 g/l) range. In CKD ND patients the review focused on energy and physical function134 and in CKD 5D patients the review focused on physical function and the meta-analysis on exercise tolerance.135 3.3: We recommend using ESA therapy with great caution, if at all, in CKD patients with active malignancy—in particular when cure is the anticipated outcome—(1B), a history of stroke (1B), or a history of malignancy (2C).

D patients the review focused on physical function and the meta-analysis on exercise tolerance.135 3.3: We recommend using ESA therapy with great caution, if at all, in CKD patients with active malignancy—in particular when cure is the anticipated outcome—(1B), a history of stroke (1B), or a history of malignancy (2C). RATIONALE The joint guideline from the American Society of Clinical Oncology136 and the American Society of Hematology137 recommend using ESA therapy with great caution in patients with active malignancy, particularly when cure is the anticipated outcome. This advice is supported in CKD patients by the post-hoc analysis in TREAT which demonstrated a significantly higher death rate from cancer in the darbepoetin arm in patients with a history of a malignant condition at baseline as compared with the placebo arm.127 The relative risk of stroke in patients in the darbepoetin arm of TREAT was the same in those with and without a history of stroke (i.e., approximately doubled). However the absolute risk of stroke was much higher in subjects with a history of stroke (in both study arms) and the absolute risk of stroke attributable to high Hb/darbepoetin was particularly high, 8% in those with a history of stroke vs 1% in those without a history of stroke over 29 months.138 Consequently the Work Group concluded that ESAs should be used with great caution in those with a prior history of stroke. 3.4.1: For adult CKD ND patients with Hb concentration ≥10.0 g/dl (≥100 g/l), we suggest that ESA therapy not be initiated. (2D)

stroke vs 1% in those without a history of stroke over 29 months.138 Consequently the Work Group concluded that ESAs should be used with great caution in those with a prior history of stroke. 3.4.1: For adult CKD ND patients with Hb concentration ≥10.0 g/dl (≥100 g/l), we suggest that ESA therapy not be initiated. (2D) 3.4.2: For adult CKD ND patients with Hb concentration <10.0 g/dl (<100 g/l) we suggest that the decision whether to initiate ESA therapy be individualized based on the rate of fall of Hb concentration, prior response to iron therapy, the risk of needing a transfusion, the risks related to ESA therapy and the presence of symptoms attributable to anemia. (2C) 3.4.3: For adult CKD 5D patients, we suggest that ESA therapy be used to avoid having the Hb concentration fall below 9.0 g/dl (90 g/l) by starting ESA therapy when the hemoglobin is between 9.0–10.0 g/dl (90–100 g/l). (2B) 3.4.4: Individualization of therapy is reasonable as some patients may have improvements in quality of life at higher Hb concentration and ESA therapy may be started above 10.0 g/dl (100 g/l). (Not Graded) 3.4.5: For all pediatric CKD patients, we suggest that the selection of Hb concentration at which ESA therapy is initiated in the individual patient includes consideration of potential benefits (e.g., improvement in quality of life, school attendance/performance, and avoidance of transfusion) and potential harms. (2D)

3.4.4: Individualization of therapy is reasonable as some patients may have improvements in quality of life at higher Hb concentration and ESA therapy may be started above 10.0 g/dl (100 g/l). (Not Graded) 3.4.5: For all pediatric CKD patients, we suggest that the selection of Hb concentration at which ESA therapy is initiated in the individual patient includes consideration of potential benefits (e.g., improvement in quality of life, school attendance/performance, and avoidance of transfusion) and potential harms. (2D) RATIONALE In adult CKD-ND patients TREAT demonstrated that the high Hb darbepoetin arm was associated with harm. In the patients on placebo with rescue treatment allowed when Hb fell to below 9.0 g/dl (90 g/l) the achieved median Hb value was as high as 10.6 g/dl (106 g/l), despite the majority of patients receiving no or little darbepoetin127 (Supplementary Tables 15–19 online).

high Hb darbepoetin arm was associated with harm. In the patients on placebo with rescue treatment allowed when Hb fell to below 9.0 g/dl (90 g/l) the achieved median Hb value was as high as 10.6 g/dl (106 g/l), despite the majority of patients receiving no or little darbepoetin127 (Supplementary Tables 15–19 online). There is no convincing evidence that the active increase of Hb towards concentrations in the normal range leads to demonstrable benefit in adult patients with CKD stages 3–5. Moreover, when Hb falls below 10 g/dl (100 g/l) in these patients the Work Group were unconvinced that all patients should have an ESA initiated, particularly as the rate of Hb fall may be slow. It was suggested that the decision to initiate ESA therapy in CKD-ND when Hb is >9.0 and <10.0 g/dl (>90 and <100 g/l) should be individualized based on risk of requiring transfusions and on the presence of symptoms attributable to anemia, particularly as some patients may be at higher risk of requiring red-cell transfusions, and some patients are more prone to developing symptoms and signs associated with anemia (Supplementary Tables 15–19 online).

vidualized based on risk of requiring transfusions and on the presence of symptoms attributable to anemia, particularly as some patients may be at higher risk of requiring red-cell transfusions, and some patients are more prone to developing symptoms and signs associated with anemia (Supplementary Tables 15–19 online). In adult hemodialysis patients the rate of fall of Hb is faster than in ND patients, and if untreated Hb will frequently fall below 8 g/dl (80 g/l).122 As the risk of transfusions is high in those HD patients whose Hb falls below 9 g/dl (90 g/l) the Work Group suggested that ESA therapy should be used to prevent the Hb concentration from falling below 9.0 g/dl (90 g/l), which in practice means that the Hb concentration at which ESA should be initiated should be between 9.0 and 10.0 g/dl [90 and 100 g/l] (Supplementary Tables 9–14 online). However, there may be subgroups of adult CKD stage 3–5 and 5D patients in whom it may not be wise to let Hb values descend below 10 g/dl (100 g/l), particularly in elderly patients who are more prone to developing symptoms and signs associated with anemia, and those who are prone to requiring red-cell transfusions.

In adult hemodialysis patients the rate of fall of Hb is faster than in ND patients, and if untreated Hb will frequently fall below 8 g/dl (80 g/l).122 As the risk of transfusions is high in those HD patients whose Hb falls below 9 g/dl (90 g/l) the Work Group suggested that ESA therapy should be used to prevent the Hb concentration from falling below 9.0 g/dl (90 g/l), which in practice means that the Hb concentration at which ESA should be initiated should be between 9.0 and 10.0 g/dl [90 and 100 g/l] (Supplementary Tables 9–14 online). However, there may be subgroups of adult CKD stage 3–5 and 5D patients in whom it may not be wise to let Hb values descend below 10 g/dl (100 g/l), particularly in elderly patients who are more prone to developing symptoms and signs associated with anemia, and those who are prone to requiring red-cell transfusions. Moreover, physical and mental performances and QoL may be seriously compromised in adult CKD patients with severe anemia. RCTs supporting registration of epoetin-alfa for the treatment of anemia in dialysis patients demonstrated that ESA treatment of subjects with a Hb of < 10 g/dl (<100 g/l) to a Hb target of approximately 10–12 g/dl (100–120 g/l) improved patient-reported physical functioning.134, 135 The question of the Hb value above which there is no further improvement in these parameters remains unsolved, especially for CKD-ND patients without diabetes and CKD-5D patients with or without diabetes.

l) to a Hb target of approximately 10–12 g/dl (100–120 g/l) improved patient-reported physical functioning.134, 135 The question of the Hb value above which there is no further improvement in these parameters remains unsolved, especially for CKD-ND patients without diabetes and CKD-5D patients with or without diabetes. In anemic children with CKD there are no RCTs examining the effects of ESA administration on hard outcomes. Therefore, any suggestion for Hb targets in this subgroup of CKD patients has to rely on results obtained in the adult CKD patient population and on clinical experience in the pediatric setting. The upper and lower Hb targets are opinion-based, in keeping with the lack of pediatric specific evidence. There are a number of factors unique to children that make exclusive reliance on evidence in adults inappropriate such as age-specific variation of normal Hb concentrations as well as QoL, growth, developmental, and psychological differences between children and adults.58 Limited data suggest that children with CKD and a Hb less than 9.9 g/dl (99 g/l) are at increased risk for mortality,139 left ventricular hypertrophy,140, 141 and/or decreased exercise capacity142 compared to those with a Hb greater than 9.9 g/dl (99 g/l). When evaluated as a continuous variable, hematocrit (Hct) was linked directly to measures of improved health and physical functioning in a health based QoL questionnaire administered to a pediatric CKD population.143 ESA MAINTENANCE THERAPY

In anemic children with CKD there are no RCTs examining the effects of ESA administration on hard outcomes. Therefore, any suggestion for Hb targets in this subgroup of CKD patients has to rely on results obtained in the adult CKD patient population and on clinical experience in the pediatric setting. The upper and lower Hb targets are opinion-based, in keeping with the lack of pediatric specific evidence. There are a number of factors unique to children that make exclusive reliance on evidence in adults inappropriate such as age-specific variation of normal Hb concentrations as well as QoL, growth, developmental, and psychological differences between children and adults.58 Limited data suggest that children with CKD and a Hb less than 9.9 g/dl (99 g/l) are at increased risk for mortality,139 left ventricular hypertrophy,140, 141 and/or decreased exercise capacity142 compared to those with a Hb greater than 9.9 g/dl (99 g/l). When evaluated as a continuous variable, hematocrit (Hct) was linked directly to measures of improved health and physical functioning in a health based QoL questionnaire administered to a pediatric CKD population.143 ESA MAINTENANCE THERAPY 3.5.1: In general, we suggest that ESAs not be used to maintain Hb concentration above 11.5 g/dl (115 g/l) in adult patients with CKD. (2C) 3.5.2: Individualization of therapy will be necessary as some patients may have improvements in quality of life at Hb concentration above 11.5 g/dl (115 g/l) and will be prepared to accept the risks. (Not Graded)

3.5.1: In general, we suggest that ESAs not be used to maintain Hb concentration above 11.5 g/dl (115 g/l) in adult patients with CKD. (2C) 3.5.2: Individualization of therapy will be necessary as some patients may have improvements in quality of life at Hb concentration above 11.5 g/dl (115 g/l) and will be prepared to accept the risks. (Not Graded) RATIONALE The suggestion to set the upper Hb target in general to values ≤11.5 g/dl (≤115 g/l) in adult CKD patients is based on the interpretation of the combined results of the recent major RCTs that there may be more harm than benefit at higher Hb concentrations. Of note, the update of the 2006 KDOQI anemia guideline in 2007 had already led to the recommendation to limit the upper Hb target to 12 g/dl (120 g/l), not to exceed 13 g/dl (130 g/l).51 The present suggestion not to exceed in general a Hb limit of 11.5 g/dl (115 g/l) has been influenced by the fact that the upper boundary of the Hb concentration in the control group of the major ESA RCTs usually did not exceed 11.5 g/dl (115 g/l); no data exist on the benefits of Hb targets between 11.5 and 13.0 g/dl (115 and 130 g/l); and high Hb targets are associated with adverse outcomes.

g/l) has been influenced by the fact that the upper boundary of the Hb concentration in the control group of the major ESA RCTs usually did not exceed 11.5 g/dl (115 g/l); no data exist on the benefits of Hb targets between 11.5 and 13.0 g/dl (115 and 130 g/l); and high Hb targets are associated with adverse outcomes. The Work Group recognized that some patients experience an improvement in QoL when the Hb value is above 11.5 g/dl (115 g/l). This opinion is supported by the heterogeneity of QoL outcomes in the major RCTs: in the double-blind Canada-Europe Study and in open label CREATE study statistically significant improvements in some QoL domains that may be clinically important were reported with higher Hb values.124, 126, 130 In the double-blind TREAT study the QoL benefits of higher Hb were modest127, 132 and in open label CHOIR study no benefits were observed128 (Supplementary Tables 9–19 online). As all CKD patients in TREAT study also had type 2 diabetes, it is possible that improvements in QoL may be more difficult to achieve in this subgroup of patients than in those not suffering from diabetes. An increase of Hb above 11.5 g/dl (115 g/l) towards 13 g/dl (130 g/l) may also be justified in individual patients with a high bleeding tendency since this results in lower transfusion needs, as shown by 8 RCTs.133

As all CKD patients in TREAT study also had type 2 diabetes, it is possible that improvements in QoL may be more difficult to achieve in this subgroup of patients than in those not suffering from diabetes. An increase of Hb above 11.5 g/dl (115 g/l) towards 13 g/dl (130 g/l) may also be justified in individual patients with a high bleeding tendency since this results in lower transfusion needs, as shown by 8 RCTs.133 Obviously, increasing Hb above 11.5 g/dl (115 g/l) up to 13 g/dl (130 g/l) has to be weighed against the probability of increased harm. This perspective needs to be clearly explained to each patient who wishes to examine the possible benefits of more complete anemia correction. 3.6: In all adult patients, we recommend that ESAs not be used to intentionally increase the Hb concentration above 13 g/dl (130 g/l). (1A)

st the probability of increased harm. This perspective needs to be clearly explained to each patient who wishes to examine the possible benefits of more complete anemia correction. 3.6: In all adult patients, we recommend that ESAs not be used to intentionally increase the Hb concentration above 13 g/dl (130 g/l). (1A) RATIONALE The strong recommendation not to aim for Hb increases to concentrations >13 g/dl (>130 g/l) is based on the interpretation of the combined results of the recent major RCTs showing more harm than benefit with higher Hb targets, as compared to lower Hb targets, including increased risks for stroke,126, 127 hypertension,133 and vascular access thrombosis (in hemodialysis patients).118 TREAT did not demonstrate significant differences for serious cardiovascular or kidney events comparing correction of anemia with darbepoetin to the placebo group.127 Thus the increased risk of kidney events reported in CREATE124 and of cardiovascular events reported in CHOIR128 were not substantiated in the much larger TREAT trial.127 However, a recent meta-analysis point estimate indicated increased mortality at higher Hb target133 (Supplementary Tables 9–19 online). An exception to the recommendation to avoid Hb increases to concentrations >13 g/dl (>130 g/l) might however be made for patients with comorbidities that are normally associated with elevated Hb levels (e.g., cyanotic heart disease). 3.7: In all pediatric CKD patients receiving ESA therapy, we suggest that the selected Hb concentration be in the range of 11.0 to 12.0 g/dl (110 to 120 g/l). (2D)

3 g/dl (>130 g/l) might however be made for patients with comorbidities that are normally associated with elevated Hb levels (e.g., cyanotic heart disease). 3.7: In all pediatric CKD patients receiving ESA therapy, we suggest that the selected Hb concentration be in the range of 11.0 to 12.0 g/dl (110 to 120 g/l). (2D) RATIONALE As mentioned above, in children with CKD observational data associates high Hb with better survival139 and/or increased exercise capacity.142 Moreover, a recent North American Pediatric Renal Trials and Collaborative Studies (NAPRTCS) retrospective analysis done on pediatric CKD patients found an increased risk of hospitalization in children with low Hb compared to those with normal Hb.144 However, based on recent experience with the adult CKD patient population, caution is warranted with any extrapolation from observational treatment studies to conclusions on hard outcomes. This being said, direct extrapolation of the results from adult trials to pediatric patients is not appropriate given the differences in causes of CKD, contributions of age to growth and development, and impact of comorbidities on outcomes. ESA DOSING 3.8.1: We recommend determining the initial ESA dose using the patient's Hb concentration, body weight, and clinical circumstances. (1D) 3.8.2: We recommend that ESA dose adjustments be made based on the patient's Hb concentration, rate of change in Hb concentration, current ESA dose and clinical circumstances. (1B)

ESA DOSING 3.8.1: We recommend determining the initial ESA dose using the patient's Hb concentration, body weight, and clinical circumstances. (1D) 3.8.2: We recommend that ESA dose adjustments be made based on the patient's Hb concentration, rate of change in Hb concentration, current ESA dose and clinical circumstances. (1B) 3.8.3: We suggest decreasing ESA dose in preference to withholding ESA when a downward adjustment of Hb concentration is needed. (2C) 3.8.4: Re-evaluate ESA dose if (Not Graded): The patient suffers an ESA-related adverse event The patient has an acute or progressive illness that may cause ESA hyporesponsiveness (see Recommendations 3.13.1–3.13.2) RATIONALE The initiation of ESA therapy, ESA dose adjustments and rates of changes have remained similar to those outlined in the 2006 KDOQI Anemia Guideline.50 In general, the objective of initial ESA therapy is a rate of increase in Hb concentrations of 1.0 to 2.0 g/dl (10 to 20 g/l) per month. This is consistent with the findings in ESA trials of CKD-associated anemia where the mean initial rates of Hb concentration increase were of 0.7 to 2.5 g/dl (7 to 25 g/l) in the first 4 weeks. However, a rise in Hb of greater than 2.0 g/dl (20 g/l) over a 4-week period should be avoided.

g/dl (10 to 20 g/l) per month. This is consistent with the findings in ESA trials of CKD-associated anemia where the mean initial rates of Hb concentration increase were of 0.7 to 2.5 g/dl (7 to 25 g/l) in the first 4 weeks. However, a rise in Hb of greater than 2.0 g/dl (20 g/l) over a 4-week period should be avoided. The rate of increase varies greatly as a function of individual ESA responsiveness. Poor responders are more likely to be female, to have a history of cardiovascular disease (CVD), to have signs of iron deficiency and inflammation, and to be overweight.145 The response also depends on initial dose, dosing frequency, and route of administration. The dependence on dosing frequency and route of administration concerns epoetin-alfa, epoetin-beta, and darbepoetin but not CERA (continuous erythropoietin receptor activator [methoxy polyethylene glycol-epoetin-beta]). When ESAs were introduced into clinical practice over 20 years ago, hypertension was frequently noted in the first 3 months after initiating therapy in severely anemic patients, and seizures in rare instances. It is possible, although not proven, that these events were related to a too rapid rate of increase in Hb concentrations.

ere introduced into clinical practice over 20 years ago, hypertension was frequently noted in the first 3 months after initiating therapy in severely anemic patients, and seizures in rare instances. It is possible, although not proven, that these events were related to a too rapid rate of increase in Hb concentrations. Epoetin-alfa or epoetin-beta dosing usually starts at 20 to 50 IU/kg body weight three times a week. Darbepoetin-alfa dosing usually starts at 0.45 μg/kg body weight once weekly by subcutaneous (SC) or IV administration, or 0.75 μg/kg body weight once every 2 weeks by SC administration. CERA dosing starts at 0.6 μg/kg body weight once every 2 weeks by SC or IV administration for CKD ND and CKD 5D patients, respectively, or 1.2 μg/kg body weight once every 4 weeks by SC administration for CKD ND patients. Higher baseline Hb concentrations require lower initial ESA doses, except for CERA for which there is no initial dose change. In patients with a history of CVD, thrombo-embolism or seizures, or in those with high blood pressure, the initial doses should be in the lower range. Epoetin-alfa or epoetin-beta dosage may subsequently be increased every 4 weeks by a weekly dose of 3 × 20 IU/kg if the increase of Hb is not adequate. Increases in dose should not be made more frequently than once a month. If the Hb is increasing and approaching 11.5 g/dl (115 g/l), the dose should be reduced by approximately 25%. If the Hb continues to increase, doses should be temporarily withheld until the Hb begins to decrease, at which point therapy should be reinitiated at a dose approximately 25% below the previous dose. Alternatively, one could simply repeat the Hb determination again in a shorter interval (e.g., weekly) and interpret any further rise, in particular in light of reticulocyte counts and their direction, before considering holding the dose. If the Hb increases by more than 1.0 g/dl (10 g/l) in any 2-week period, the dose should be decreased by approximately 25%. See Recommendations 3.13.1 to 3.15.2 regarding ESA hyporesponsiveness and loss of ESA response (Supplementary Table 20 online).

yte counts and their direction, before considering holding the dose. If the Hb increases by more than 1.0 g/dl (10 g/l) in any 2-week period, the dose should be decreased by approximately 25%. See Recommendations 3.13.1 to 3.15.2 regarding ESA hyporesponsiveness and loss of ESA response (Supplementary Table 20 online). Dose adjustments may be necessary once the Hb target range has been reached. Note that in clinical practice, achieved Hb values may easily rise above or fall below the optimal Hb limits. Therefore, cautious dose adaptations are required. In general, ESA dose adjustments are made only after the first 4 weeks after ESA initiation. The frequency of ESA dose adjustment should be determined by the rate of increase in Hb concentrations during initial ESA therapy, the stability of Hb concentrations during maintenance ESA therapy, and the frequency of Hb testing. The minimum interval between ESA dose adjustments in the outpatient setting generally is 2 weeks because the effect of most dose changes will not be seen within a shorter interval. ESA doses should be decreased, but not necessarily held, when a downward adjustment of Hb concentration is needed. Withholding ESA doses, particularly for long periods, may lead to a delayed decrease in Hb concentrations to less than target range. Such a decrease may initiate periodic cycling of Hb concentrations at greater than and less than the target Hb range.146 Hb variability has been found to be an independent predictor of mortality in a large US CKD 5HD patient population147 although this observation could not be confirmed in a large European CKD 5HD patient cohort.148

e may initiate periodic cycling of Hb concentrations at greater than and less than the target Hb range.146 Hb variability has been found to be an independent predictor of mortality in a large US CKD 5HD patient population147 although this observation could not be confirmed in a large European CKD 5HD patient cohort.148 Each time a patient with CKD is hospitalized the treating clinician should evaluate or reevaluate the patient's ESA requirements. Disease states such as severe infections or post-surgery may modify the ESA responsiveness profoundly. In case of profound anemia and markedly impaired ESA response a red cell transfusion may be preferred to administering ESAs or increasing ESA dose. ESA ADMINISTRATION 3.9.1: For CKD 5HD patients and those on hemofiltration or hemodiafiltration therapy, we suggest either intravenous or subcutaneous administration of ESA. (2C) 3.9.2: For CKD ND and CKD 5PD patients, we suggest subcutaneous administration of ESA. (2C)

Each time a patient with CKD is hospitalized the treating clinician should evaluate or reevaluate the patient's ESA requirements. Disease states such as severe infections or post-surgery may modify the ESA responsiveness profoundly. In case of profound anemia and markedly impaired ESA response a red cell transfusion may be preferred to administering ESAs or increasing ESA dose. ESA ADMINISTRATION 3.9.1: For CKD 5HD patients and those on hemofiltration or hemodiafiltration therapy, we suggest either intravenous or subcutaneous administration of ESA. (2C) 3.9.2: For CKD ND and CKD 5PD patients, we suggest subcutaneous administration of ESA. (2C) RATIONALE As outlined in the 2006 KDOQI guideline,50 the route of administration should be determined by the CKD stage, treatment setting, efficacy considerations, and the class of ESA used. Among CKD 5D patients undergoing intermittent hemodialysis or hemofiltration therapy, either SC or IV administration is possible. In the outpatient setting, SC administration is the only routinely feasible route of administration for patients with CKD 3–5 or on peritoneal dialysis treatment. Among short-acting ESAs, efficacy of SC administration in patients with CKD 5HD may be superior to that of IV administration, as shown by a large multicenter RCT in hemodialysis patients.149 However, another RCT of much smaller sample size did not find an advantage of SC over IV administration in CKD 5HD patients.150 Among long-acting ESAs, efficacy of SC compared with IV administration appears to be equivalent at examined dosing frequencies.151, 152, 153 Furthermore, CKD 5HD patients in general prefer IV to SC administration of ESAs because SC administration may be painful (Supplementary Tables 21–24 online).

CKD 5HD patients.150 Among long-acting ESAs, efficacy of SC compared with IV administration appears to be equivalent at examined dosing frequencies.151, 152, 153 Furthermore, CKD 5HD patients in general prefer IV to SC administration of ESAs because SC administration may be painful (Supplementary Tables 21–24 online). Frequency of administration 3.10: We suggest determining the frequency of ESA administration based on CKD stage, treatment setting, efficacy considerations, patient tolerance and preference, and type of ESA. (2C) RATIONALE The frequency of ESA administration depends on considerations of efficacy, convenience and comfort. Maximum efficacy occurs within dosing intervals that are ESA class specific. For example, in patients on hemodialysis treatment receiving SC or IV short-acting ESA therapy, epoetin-alfa efficacy decreases when the dosing is extended from 3 times weekly to once-weekly administration,154 and even more so when the dosing intervals are extended to every other week administration.155 Among long-acting ESAs, darbepoetin-alfa appears to have maximum efficacy when administered every 2 weeks, and methoxy polyethylene glycol-epoetin-beta (CERA) every 4 weeks.156 When converting short-acting ESAs to long-acting ESAs, differences in drug half-life need to be considered. For the sake of comparison, 3 times weekly administered epoetin-alfa to darbepoetin-alfa given only once monthly resulted in a decreased frequency of injections needed to maintain Hb concentrations of CKD patients within an accepted target range157 (Supplementary Tables 25–28 online).

g half-life need to be considered. For the sake of comparison, 3 times weekly administered epoetin-alfa to darbepoetin-alfa given only once monthly resulted in a decreased frequency of injections needed to maintain Hb concentrations of CKD patients within an accepted target range157 (Supplementary Tables 25–28 online). When converting a patient from one ESA to another the pharmacokinetic and pharmacodynamic characteristics of the new ESA need to be taken into consideration. The manufacturers have provided conversions from epoetin-alfa or epoetin-beta to darbepoetin-alfa or CERA. Note that the conversion ratios from epoetin to darbepoetin are non-linear. When using different types of approved ESAs (biosimilars that have received approval by official regulatory bodies such as FDA and European Medicines Agency [EMA]), license information provided by companies should also be taken into account. TYPE OF ESA 3.11.1: We recommend choosing an ESA based on the balance of pharmacodynamics, safety information, clinical outcome data, costs, and availability. (1D) 3.11.2: We suggest using only ESAs that have been approved by an independent regulatory agency. Specifically for ‘copy' versions of ESAs, true biosimilar products should be used. (2D)

3.11.1: We recommend choosing an ESA based on the balance of pharmacodynamics, safety information, clinical outcome data, costs, and availability. (1D) 3.11.2: We suggest using only ESAs that have been approved by an independent regulatory agency. Specifically for ‘copy' versions of ESAs, true biosimilar products should be used. (2D) RATIONALE As outlined above, the choice of short-acting or long-acting ESAs needs to take into account a number of different aspects, encompassing patient-oriented issues and country-specific considerations. At present, there is no evidence that any given ESA brand is superior to another in terms of patient outcomes, with the historical exception of the temporary increase in the incidence of antibody-mediated pure red cell aplasia (PRCA) about 10–20 years ago, which was associated with SC administration of an epoetin-alfa formulation available in Europe, but not in the United States.158, 159 It is the considered opinion of the Work Group that the likelihood of differences in clinical outcomes among ESA brands is low, although there is no robust evidence supporting this assumption (Supplementary Tables 29–32 online).

n of an epoetin-alfa formulation available in Europe, but not in the United States.158, 159 It is the considered opinion of the Work Group that the likelihood of differences in clinical outcomes among ESA brands is low, although there is no robust evidence supporting this assumption (Supplementary Tables 29–32 online). At present, a number of different types of short-acting or long-acting ESAs are available worldwide, including original formulations, biosimilars, and ‘copy' ESAs which have not been exposed to the rigor of scientific evaluation as mandated by the regulatory agencies prior to approval. Their accessibility and costs vary from country to country. True biosimilars, as defined by the EMA, are not identical to the originator products, but they have undergone a minimum number of regulatory ‘equivalence' or ‘non-inferiority' studies to gain marketing authorization in Europe. In other countries outside Europe, some ‘copy' ESA products have been marketed that may not have undergone the same rigorous testing.160 Since patient safety is one of the most important drug treatment issues, only biosimilars approved by an independent regulatory agency should be used. EVALUATING AND CORRECTING PERSISTENT FAILURE TO REACH OR MAINTAIN INTENDED HEMOGLOBIN CONCENTRATION Frequency of monitoring 3.12.1: During the initiation phase of ESA therapy, measure Hb concentration at least monthly. (Not Graded) 3.12.2: For CKD ND patients, during the maintenance phase of ESA therapy measure Hb concentration at least every 3 months. (Not Graded)

EVALUATING AND CORRECTING PERSISTENT FAILURE TO REACH OR MAINTAIN INTENDED HEMOGLOBIN CONCENTRATION Frequency of monitoring 3.12.1: During the initiation phase of ESA therapy, measure Hb concentration at least monthly. (Not Graded) 3.12.2: For CKD ND patients, during the maintenance phase of ESA therapy measure Hb concentration at least every 3 months. (Not Graded) 3.12.3: For CKD 5D patients, during the maintenance phase of ESA therapy measure Hb concentration at least monthly. (Not Graded)

3.12.1: During the initiation phase of ESA therapy, measure Hb concentration at least monthly. (Not Graded) 3.12.2: For CKD ND patients, during the maintenance phase of ESA therapy measure Hb concentration at least every 3 months. (Not Graded) 3.12.3: For CKD 5D patients, during the maintenance phase of ESA therapy measure Hb concentration at least monthly. (Not Graded) RATIONALE ESA initiation phase. The suggestion to monitor Hb values at least monthly in patients in whom ESA therapy is started is intended to provide sufficient surveillance information to assist in achieving and maintaining desired Hb concentrations safely and follows common practice.50 The minimum interval between ESA dose adjustments is 2 weeks because the effect of most dose changes will not be seen within a shorter interval. Consideration of an ESA dose adjustment is based on the next projected Hb concentration. Because the accuracy of projection (extrapolation) increases with the number of contributing data points, the frequency of Hb monitoring is likely to be an important determinant of the accuracy of ESA dose adjustment. However, evidence to support this line of reasoning is indirect. Several RCTs have randomized CKD 5HD patients with target-range Hb concentrations to a change in frequency of ESA administration, a change in ESA class, or both. RCTs that have monitored Hb values weekly and adjusted ESA doses as frequently as every 2 weeks have achieved stable Hb concentrations early after randomization.152, 161, 162 In contrast, an RCT that monitored Hb concentrations and considered ESA dose adjustment monthly required 6 to 9 months to stabilize Hb concentrations after randomization,163 but mean Hb concentration remained within the target range for that trial.

hieved stable Hb concentrations early after randomization.152, 161, 162 In contrast, an RCT that monitored Hb concentrations and considered ESA dose adjustment monthly required 6 to 9 months to stabilize Hb concentrations after randomization,163 but mean Hb concentration remained within the target range for that trial. ESA maintenance phase. Within the recommended ranges for monitoring and dose adjustment, unstable Hb concentration, inappropriate high or low Hb concentration, and hemodialysis favor shorter intervals of ESA administration, whereas stable Hb concentration, within target Hb concentration, peritoneal dialysis, CKD 3–5, and minimizing laboratory resource utilization favor longer intervals for long-acting ESAs such as darbepoetin. The frequency of ESA dose adjustment is unaffected by length of action: during an 8-week period with weekly Hb monitoring, about equal numbers of patients receiving either short-acting ESA thrice weekly or darbepoetin once weekly required dose adjustments (44% and 49%, respectively).162 Initial ESA hyporesponsiveness 3.13.1: Classify patients as having ESA hyporesponsiveness if they have no increase in Hb concentration from baseline after the first month of ESA treatment on appropriate weight-based dosing. (Not Graded) 3.13.2: In patients with ESA hyporesponsiveness, we suggest avoiding repeated escalations in ESA dose beyond double the initial weight-based dose. (2D) Subsequent ESA hyporesponsiveness

3.13.1: Classify patients as having ESA hyporesponsiveness if they have no increase in Hb concentration from baseline after the first month of ESA treatment on appropriate weight-based dosing. (Not Graded) 3.13.2: In patients with ESA hyporesponsiveness, we suggest avoiding repeated escalations in ESA dose beyond double the initial weight-based dose. (2D) Subsequent ESA hyporesponsiveness 3.14.1: Classify patients as having acquired ESA hyporesponsiveness if after treatment with stable doses of ESA, they require 2 increases in ESA doses up to 50% beyond the dose at which they had been stable in an effort to maintain a stable Hb concentration. (Not Graded) 3.14.2: In patients with acquired ESA hyporesponsiveness, we suggest avoiding repeated escalations in ESA dose beyond double the dose at which they had been stable. (2D) Management of poor ESA responsiveness 3.15.1: Evaluate patients with either initial or acquired ESA hyporesponsiveness and treat for specific causes of poor ESA response. (Not Graded) 3.15.2: For patients who remain hyporesponsive despite correcting treatable causes, we suggest individualization of therapy, accounting for relative risks and benefits of (2D): decline in Hb concentration continuing ESA, if needed to maintain Hb concentration, with due consideration of the doses required, and blood transfusions

3.15.2: For patients who remain hyporesponsive despite correcting treatable causes, we suggest individualization of therapy, accounting for relative risks and benefits of (2D): decline in Hb concentration continuing ESA, if needed to maintain Hb concentration, with due consideration of the doses required, and blood transfusions RATIONALE Relative resistance to the effect of ESAs is a common problem in managing the anemia of patients with CKD and remains the subject of intense interest, all the more since ESA hyporesponsiveness has been found to be among the most powerful predictors of the risk of cardiovascular events and mortality.164 Recently a report from TREAT assessed the initial Hb response to darbepoetin after two weight-based doses at 2 weekly intervals, in 1872 patients with CKD and diabetes.145 Patients with a poor response, (the lowest quartile, who had <2% change in Hb concentration after 1 month), had higher rates of the composite cardiovascular events (adjusted HR 1.31, 95% CI 1.09–1.59), compared to those with a better response. Although this differential effect may be related to comorbidity in hyporesponsive patients, nonetheless it is possible that the high ESA doses used in hyporesponsive patients may be toxic. Though not empirically tested, per se, the definition of initial hyporesponsiveness agreed upon by the Work Group is derived from the secondary analysis of the TREAT study.145 Since a <2% increase in the Hb concentration is likely to be within the variability range of Hb values in individual patients, this value is considered as ″no increase.″ The definition of initial hyporesponsiveness relies on presently accepted ESA starting doses, as indicated in the Rationale under 3.8.1–3.8.4. Of note, weight-based doses for darbepoetin do not differ for IV or SC routes, but do differ for epoetin-alfa.

in individual patients, this value is considered as ″no increase.″ The definition of initial hyporesponsiveness relies on presently accepted ESA starting doses, as indicated in the Rationale under 3.8.1–3.8.4. Of note, weight-based doses for darbepoetin do not differ for IV or SC routes, but do differ for epoetin-alfa. If lower initial dosages than those used in TREAT are chosen, the diagnosis of hyporesponsiveness must take this into account. For example, in the USA the label for darbepoetin now recommends a starting dose of 0.45 μg per kg per four weeks, much lower than the dose used in TREAT or in Europe (i.e., 0.45 μg per kg per week or 0.75 μg per kg per two weeks). If such lower starting doses are used, repeated escalations in ESA dose should be allowed to reach double the weight-based dose used in TREAT. Although the distinction between initial ESA hyporesponsiveness and acquired partial or complete loss of ESA responsiveness in a patient with already treated, stable anemia is somewhat artificial, it is useful in our opinion for clinical practice.

If lower initial dosages than those used in TREAT are chosen, the diagnosis of hyporesponsiveness must take this into account. For example, in the USA the label for darbepoetin now recommends a starting dose of 0.45 μg per kg per four weeks, much lower than the dose used in TREAT or in Europe (i.e., 0.45 μg per kg per week or 0.75 μg per kg per two weeks). If such lower starting doses are used, repeated escalations in ESA dose should be allowed to reach double the weight-based dose used in TREAT. Although the distinction between initial ESA hyporesponsiveness and acquired partial or complete loss of ESA responsiveness in a patient with already treated, stable anemia is somewhat artificial, it is useful in our opinion for clinical practice. In the Normal Hematocrit Study both the high Hb and the low Hb groups revealed an inverse relationship between achieved Hb and the primary outcome (death or myocardial infarction).118 This is consistent with the idea that those patients who failed to achieve the target Hb were unable to do so because comorbid condition(s) existed that prevented achievement of this target. Thus, hyporesponsiveness may just have been a marker for adverse outcomes, although the possibility that high ESA doses used in hyporesponsive patients are toxic in themselves cannot be excluded. Dose-targeting bias has been reported by the Kidney Disease Clinical Studies Initiative Hemodialysis Study (HEMO) investigators.165 In this RCT ESRD patients, randomly allocated to either high or low quantity of dialysis, as measured by Kt/V, demonstrated an inverse relationship between achieved Kt/V and mortality. The interpretation was that patients with comorbid conditions were unable to achieve higher Kt/V and that comorbidity predisposed these patients to earlier death.

randomly allocated to either high or low quantity of dialysis, as measured by Kt/V, demonstrated an inverse relationship between achieved Kt/V and mortality. The interpretation was that patients with comorbid conditions were unable to achieve higher Kt/V and that comorbidity predisposed these patients to earlier death. The same principle as used with defining hyporesponsiveness to darbepoetin could be applied to the early response to other short-acting ESAs but cannot be applied to longer acting ESAs such as CERA. In that case, evaluating the Hb response after a time period of 2 months appears to be appropriate. Early ESA hyporesponsiveness or the subsequent occurrence of hyporesponsiveness in CKD patients with previously stable Hb values should lead to an intensive search for potentially correctable factors which might be causally involved. Unfortunately, besides iron deficiency, there are only few other easily reversible factors that contribute to ESA hyporesponsiveness, as shown in Table 3. If other such factors are identified they should be treated as well. Although most disorders associated with hyporesponsiveness are readily apparent, hyporesponsive patients should be evaluated for coexisting oncological or hematologic disorders. They include hematological and non-hematological malignancies as well as such diverse hematological conditions as thalassemia, sickle cell disease or the anemia associated with other chronic diseases. Myelodysplastic syndromes are a particular case. If at all ESA responsive, the anemia in patients with myelodysplastic syndrome responds more slowly. Therefore, 1 month may be too short to define hyporesponsiveness in this and several other conditions. Moreover, patients with myelodysplastic syndromes may need higher ESA doses. Finally, a rare disorder, PRCA, deserves special consideration (see 3.17.1–3.17.3). The estimation of loss of ESA response also may require a longer observation time in some patients. Note that poor ESA response, either in the initial correction phase or subsequently, is most often a transient condition. Complete loss of response is exceptional. Poor responders should periodically be re-tested for responsiveness, including after the correction of treatable causes of hyporesponsiveness.

ome patients. Note that poor ESA response, either in the initial correction phase or subsequently, is most often a transient condition. Complete loss of response is exceptional. Poor responders should periodically be re-tested for responsiveness, including after the correction of treatable causes of hyporesponsiveness. It is important to note that the dosing requirements may differ substantially between children and adults. Registry data from NAPRTCS showed that young children require higher doses of ESA than adults, ranging from 275 U/kg/week to 350 U/kg/week for infants and 200–250 U/kg/week for older children.166 Another retrospective analysis among patients on chronic hemodialysis found that children and adolescents required higher absolute doses of ESA than adults to maintain target hemoglobin levels, despite the lower mean body weight of the children.167 Unfortunately, there are no RCTs that establish the appropriate dosing of ESA in children. Future research to establish pediatric ESA dosing guidelines is needed, especially for infants and younger children. There may be toxicity from high doses of ESA, as suggested, though not proven, by recent post-hoc analyses of major ESA RCTs,145, 168 especially in conjunction with the achievement of high Hb levels.169 Therefore, in general ESA dose escalation should be avoided. The Work Group suggestions for initial and acquired hyopresponsiveness imply that maximal doses should be no greater than four times initial weight-based appropriate doses.

ESA RCTs,145, 168 especially in conjunction with the achievement of high Hb levels.169 Therefore, in general ESA dose escalation should be avoided. The Work Group suggestions for initial and acquired hyopresponsiveness imply that maximal doses should be no greater than four times initial weight-based appropriate doses. In practice, Tables 3 and 4 can guide to diagnose and correct ESA hyporesponsiveness. In patients in whom all correctable causes have been maximally treated but who remain hyporesponsive, ESA therapy may be continued cautiously at doses up to 4 times the initial dose to prevent a further decline in Hb concentration. Red cell transfusions can be used to prevent or treat anemia-related symptoms and signs. The treatment strategy needs to take into account each patient's anemia tolerance and potential benefits and risks linked to increases in Hb values solely obtained by high ESA dosing. Given the disproportionate burden of morbidity and mortality that the hyporesponsive patient population bears and the ESA expense that hyporesponsiveness engenders, further research is necessary on the causes and management of hyporesponsiveness. ADJUVANT THERAPIES 3.16.1: We recommend not using androgens as an adjuvant to ESA treatment. (1B) 3.16.2: We suggest not using adjuvants to ESA treatment including vitamin C, vitamin D, vitamin E, folic acid, L-carnitine, and pentoxifylline. (2D) RATIONALE Several adjuvant treatments have been proposed, either with the goal of limiting the use of more expensive ESA therapy or to improve ESA responsiveness.

3.16.1: We recommend not using androgens as an adjuvant to ESA treatment. (1B) 3.16.2: We suggest not using adjuvants to ESA treatment including vitamin C, vitamin D, vitamin E, folic acid, L-carnitine, and pentoxifylline. (2D) RATIONALE Several adjuvant treatments have been proposed, either with the goal of limiting the use of more expensive ESA therapy or to improve ESA responsiveness. Androgens. The use of androgens for treatment of anemia was suggested long before rHuEPO became available in clinical practice. Androgens were used regularly in many centers in the treatment of anemia in dialysis patients despite the need for intramuscular (IM) injection and a variety of adverse events, including acne, virilization, priapism, liver dysfunction, injection-site pain, and risk for peliosis hepatis and hepatocellular carcinoma. The three RCTs that tested androgens in combination with ESA therapy in CKD 5HD patients were all small short-term studies. Currently recommended Hb concentrations were not achieved, and in two of them the ESA doses used were lower than current practice.170, 171, 172 The studies did not enroll patients with ESA hyporesponsiveness, so the effect of androgens on hyporesponsiveness is unknown. The risks of androgen therapy and their uncertain benefit on Hb concentration or clinical outcomes argue against their use as an ESA adjuvant.

used were lower than current practice.170, 171, 172 The studies did not enroll patients with ESA hyporesponsiveness, so the effect of androgens on hyporesponsiveness is unknown. The risks of androgen therapy and their uncertain benefit on Hb concentration or clinical outcomes argue against their use as an ESA adjuvant. Vitamin C. Vitamin C has been reported to increase the release of iron from ferritin and the reticuloendothelial system and increase iron utilization during heme synthesis.173, 174 A recent meta-analysis of vitamin C use in CKD 5HD175 and a more recent small RCT176 concluded that vitamin C may result in larger increases in Hb and may limit the use of ESAs. In seven trials, patients generally had functional iron deficiency and in three studies they had EPO hyporesponsiveness (variously defined).176, 177, 178 However, the number of patients studied was insufficient to address the safety of this intervention. Thus the long-term safety of IV ascorbic acid in HD patients remains undefined, and whether secondary oxalosis should be a concern. Convincing data do not exist for other potential adjuvants including vitamin D, vitamin E, folic acid, L-carnitine and pentoxifylline. Several anecdotal reports, small case series, and nonrandomized studies, primarily in CKD 5HD patients, have been published, but do not provide sufficient evidence upon which to base a recommendation. Future RCTs are clearly needed for ESA adjuvants. EVALUATION FOR PURE RED CELL APLASIA (PRCA)

Convincing data do not exist for other potential adjuvants including vitamin D, vitamin E, folic acid, L-carnitine and pentoxifylline. Several anecdotal reports, small case series, and nonrandomized studies, primarily in CKD 5HD patients, have been published, but do not provide sufficient evidence upon which to base a recommendation. Future RCTs are clearly needed for ESA adjuvants. EVALUATION FOR PURE RED CELL APLASIA (PRCA) 3.17.1: Investigate for possible antibody-mediated PRCA when a patient receiving ESA therapy for more than 8 weeks develops the following (Not Graded): Sudden rapid decrease in Hb concentration at the rate of 0.5 to 1.0 g/dl (5 to 10 g/l) per week OR requirement of transfusions at the rate of approximately 1 to 2 per week, AND Normal platelet and white cell counts, AND Absolute reticulocyte count less than 10,000/μl 3.17.2: We recommend that ESA therapy be stopped in patients who develop antibody-mediated PRCA. (1A) 3.17.3: We recommend peginesatide be used to treat patients with antibody-mediated PRCA. (1B)

3.17.1: Investigate for possible antibody-mediated PRCA when a patient receiving ESA therapy for more than 8 weeks develops the following (Not Graded): Sudden rapid decrease in Hb concentration at the rate of 0.5 to 1.0 g/dl (5 to 10 g/l) per week OR requirement of transfusions at the rate of approximately 1 to 2 per week, AND Normal platelet and white cell counts, AND Absolute reticulocyte count less than 10,000/μl 3.17.2: We recommend that ESA therapy be stopped in patients who develop antibody-mediated PRCA. (1A) 3.17.3: We recommend peginesatide be used to treat patients with antibody-mediated PRCA. (1B) RATIONALE Rarely, patients undergoing ESA therapy develop antibodies that neutralize both ESA and endogenous erythropoietin. The resulting syndrome, antibody-mediated PRCA, is characterized by the sudden development of severe transfusion-dependent anemia. Rapid recognition, appropriate evaluation, and prompt intervention can be effective in limiting the consequences of this life-threatening condition. Antibody-mediated PRCA, although rare in patients administered ESAs, received urgent attention after 1998. Between 1989 and 1998, three reports described the development of PRCA in only a small number of patients with CKD administered ESAs. Reports of PRCA increased sharply in 1998 and reached a peak in 2002.159, 179 These reports were associated with SC administration of an epoetin-alfa formulation not available in the United States. After removal of this formulation from the market, by 2004, the incidence of new antibody-mediated PRCA had decreased to pre-1998 levels. Isolated cases of PRCA have been observed in association with the use of other ESAs.159, 179, 180 Outside this historical episode the incidence rate of PRCA with SC use of all other forms of SC-administered ESA is estimated to be 0.5 cases/10,000 patient-years.158 Antibody-associated PRCA stemming from IV administration of ESAs is rare and has only been reported anecdotally.181

h the use of other ESAs.159, 179, 180 Outside this historical episode the incidence rate of PRCA with SC use of all other forms of SC-administered ESA is estimated to be 0.5 cases/10,000 patient-years.158 Antibody-associated PRCA stemming from IV administration of ESAs is rare and has only been reported anecdotally.181 Recommendations based on expert opinions have been published to guide the workup and therapy of patients suspected to have antibody-mediated PRCA.179, 182, 183, 184 The two main distinguishing features of antibody-mediated PRCA are the associated decline in blood Hb concentration of approximately 4 g/dl (40 g/l) per month, and a decrease in the number of circulating reticulocytes to <10,000/μl of blood.185 Bone marrow biopsy characteristically shows reduced numbers or absence of erythroblasts. The definitive diagnosis is dependent upon demonstration of the presence of neutralizing antibodies against erythropoietin. Evidence for parvovirus infection as an alternative cause of PRCA should be sought and excluded.

blood.185 Bone marrow biopsy characteristically shows reduced numbers or absence of erythroblasts. The definitive diagnosis is dependent upon demonstration of the presence of neutralizing antibodies against erythropoietin. Evidence for parvovirus infection as an alternative cause of PRCA should be sought and excluded. Following a diagnosis of antibody-mediated PRCA, patients should stop treatment with the incriminated ESA immediately and not resume treatment with the same or another EPO-derived ESA.184 Immunosuppressive therapy may hasten the disappearance of circulating antibodies in patients with EPO-induced PRCA, and allow endogenous erythropoiesis to recover to pre-treatment levels. In a retrospective study of 47 patients who developed PRCA during EPO therapy (primarily epoetin brand ‘Eprex®' in Europe), 29 of 37 patients (78%) who received immunosuppressive therapy recovered, whereas none of the nine patients who did not receive immunosuppressive therapy recovered.185 Red cell production recovered only when patients received immunosuppressive treatment. Re-exposure to epoetins or darbepoetin-alfa can re-induce the formation of antibodies.186 Anaphylactoid reactions after repeated injections of epoetin- or darbepoetin-alfa have been reported in a patient with pure red-cell aplasia.187 A novel approach to the treatment of this condition using a synthetic, peptide-based erythropoietin-receptor agonist (peginesatide) has generated optimistic results,188 and has the advantage of avoiding immunosuppressive therapy.

poetin- or darbepoetin-alfa have been reported in a patient with pure red-cell aplasia.187 A novel approach to the treatment of this condition using a synthetic, peptide-based erythropoietin-receptor agonist (peginesatide) has generated optimistic results,188 and has the advantage of avoiding immunosuppressive therapy. The recognition of antibody-mediated PRCA in patients treated with recombinant epoetins has underscored the need for full clinical documentation and post-marketing surveillance with newer ESAs and biosimilar products, as well as therapeutic recombinant proteins in general.189 If a decision to treat with peginesatide is taken, it can be initiated at a dose of 0.05 to 0.075 mg/kg body weight by subcutaneous injection every 4 weeks. Subsequently, the dose needs to be adjusted to reach the desired target Hb value. RESEARCH RECOMMENDATIONS The following research questions have arisen during the deliberations of the Work Group, and further research will be necessary to answer them. In cohort studies moderate anemia is associated with an increased incidence of cardiovascular events. Is anemia really a risk factor for these events or is it a marker for some other cardiovascular risk factor(s)? There is uncertainty about optimal Hb targets for ESA therapy. What is the risk-benefit ratio of low Hb targets <10.0 g/dl (<100 g/l) or high targets of 11.5–13.0 g/dl (115–130 g/l), compared to conventional targets of 10.0–11.5 g/dl (100–115 g/l)?

In cohort studies moderate anemia is associated with an increased incidence of cardiovascular events. Is anemia really a risk factor for these events or is it a marker for some other cardiovascular risk factor(s)? There is uncertainty about optimal Hb targets for ESA therapy. What is the risk-benefit ratio of low Hb targets <10.0 g/dl (<100 g/l) or high targets of 11.5–13.0 g/dl (115–130 g/l), compared to conventional targets of 10.0–11.5 g/dl (100–115 g/l)? These guidelines have stressed individualization of anemia therapy. Should the objective of anemia therapy be improvement in clinical outcomes (provided Hb concentration is <13.0 g/dl [<130 g/l]) rather than achievement of a specified Hb target range? Should these outcomes include improvements in QoL, and if so, what defines clinically important improvements? As the relationship between ESA responsiveness and hard patient outcomes may be the result of co-morbidity or of high ESA dose, what is the impact of high vs low dose on clinical outcomes in ESA hyporesponsive patients? Is the risk-benefit ratio of anemia correction similar in non-diabetic and diabetic CKD patients? Is there a difference in adverse clinical outcomes comparing IV and SC routes of administration? Are the risk-benefit ratios for biosimilars comparable to current ESAs? What is the pathogenesis of cerebrovascular and vascular toxicity associated with normalization of Hb using ESAs? Are CKD patients with cancer or a cancer history who are receiving ESA therapy at higher cardiovascular risk than non-CKD patients with cancer or a cancer history?

Are the risk-benefit ratios for biosimilars comparable to current ESAs? What is the pathogenesis of cerebrovascular and vascular toxicity associated with normalization of Hb using ESAs? Are CKD patients with cancer or a cancer history who are receiving ESA therapy at higher cardiovascular risk than non-CKD patients with cancer or a cancer history? What is the effect of vitamin C administration in functional iron deficiency and what is the clinical impact of increased oxalate levels? There appears to be differences in anemia treatment outcomes between different geographic regions. What are the reasons for this? What are the risks and benefits of ESA administration on outcomes in anemic children with CKD? What are the appropriate, weight-based, dosing regimens for the younger pediatric patients, especially those under the age of two years?

There appears to be differences in anemia treatment outcomes between different geographic regions. What are the reasons for this? What are the risks and benefits of ESA administration on outcomes in anemic children with CKD? What are the appropriate, weight-based, dosing regimens for the younger pediatric patients, especially those under the age of two years? DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. SUPPLEMENTARY MATERIAL Supplemental Table 7: Association between anemia severity (prior to erythropoietin use) and clinical outcome in multivariable analyses. Supplemental Table 8: Association between hyperparathyroidism and ESA responsiveness in multivariable analyses.

DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. SUPPLEMENTARY MATERIAL Supplemental Table 7: Association between anemia severity (prior to erythropoietin use) and clinical outcome in multivariable analyses. Supplemental Table 8: Association between hyperparathyroidism and ESA responsiveness in multivariable analyses. Supplemental Table 9: Evidence profile of RCTs comparing higher vs. lower Hb targets/ESA doses in the HD-CKD and PD-CKD populations. Supplemental Table 10: Summary table of RCTs comparing different Hb targets/ESA doses on key clinical outcomes in the HD-CKD and PD-CKD populations. Supplemental Table 11: Summary table of RCTs comparing different Hb targets/ESA doses on quality of life in the HD-CKD and PD-CKD populations.

Supplemental Table 9: Evidence profile of RCTs comparing higher vs. lower Hb targets/ESA doses in the HD-CKD and PD-CKD populations. Supplemental Table 10: Summary table of RCTs comparing different Hb targets/ESA doses on key clinical outcomes in the HD-CKD and PD-CKD populations. Supplemental Table 11: Summary table of RCTs comparing different Hb targets/ESA doses on quality of life in the HD-CKD and PD-CKD populations. Supplemental Table 12: Summary table of RCTs comparing different Hb targets/ESA doses on Fatigue, Vitality/Energy, and Physical function in the HD-CKD and PD-CKD populations. Supplemental Table 13: Summary table of RCTs comparing different Hb targets/ESA doses on non-CVD/mortality adverse event rates in the HD-CKD and PD-CKD populations. Supplemental Table 14: Summary table of RCTs comparing different Hb targets/ESA doses on exercise capacity in the HD-CKD and PD-CKD populations. Supplemental Table 15: Evidence profile of RCTs comparing different higher vs. lower Hb targets/ESA doses in the ND-CKD populations. Supplemental Table 16: Summary table of RCTs comparing different Hb targets/ESA doses on key clinical outcomes in the ND-CKD population. Supplemental Table 17: Summary table of RCTs comparing different Hb targets/ESA doses on quality of life in the ND-CKD population. Supplemental Table 18: Summary table of RCTs comparing different Hb targets/ESA doses on Fatigue, Vitality/Energy, and Physical function in the ND-CKD population.

Supplemental Table 16: Summary table of RCTs comparing different Hb targets/ESA doses on key clinical outcomes in the ND-CKD population. Supplemental Table 17: Summary table of RCTs comparing different Hb targets/ESA doses on quality of life in the ND-CKD population. Supplemental Table 18: Summary table of RCTs comparing different Hb targets/ESA doses on Fatigue, Vitality/Energy, and Physical function in the ND-CKD population. Supplemental Table 19: Summary table of RCTs comparing different Hb targets/ESA doses on non-CVD/mortality adverse event rates in the ND-CKD population. Supplemental Table 20: ESA protocols from the major trials in CKD populations. Supplemental Table 21: Evidence profile of RCTs examining IV vs. SC EPO in CKD patients with anemia. Supplemental Table 22: Summary table of RCTs examining IV vs. SC ESA in CKD patients with anemia (categorical outcomes). Supplemental Table 23: Summary table of RCTs examining IV vs. SC ESA in CKD patients with anemia (continuous outcomes). Supplemental Table 24: Summary table of adverse events in RCTs examining IV vs. SC EPO in CKD patients with anemia. Supplemental Table 25: Evidence profile of RCTs examining different dosing schedules in CKD patients with anemia. Supplemental Table 26: Summary table of RCTs examining different dosing schedules in CKD patients with anemia (categorical outcomes). Supplemental Table 27: Summary table of RCTs examining different dosing schedules in CKD patients with anemia (continuous outcomes).

Supplemental Table 25: Evidence profile of RCTs examining different dosing schedules in CKD patients with anemia. Supplemental Table 26: Summary table of RCTs examining different dosing schedules in CKD patients with anemia (categorical outcomes). Supplemental Table 27: Summary table of RCTs examining different dosing schedules in CKD patients with anemia (continuous outcomes). Supplemental Table 28: Summary table of adverse events in RCTs examining different dosing schedules in CKD patients with anemia. Supplemental Table 29: Evidence profile of RCTs examining ESA vs. ESA in CKD patients with anemia. Supplemental Table 30: Summary table of RCTs examining ESA vs. ESA in CKD patients with anemia (categorical outcomes). Supplemental Table 31: Summary table of RCTs examining ESA vs. ESA in CKD patients with anemia (continuous outcomes). Supplemental Table 32: Summary table of adverse events in RCTs examining ESA vs. ESA in CKD patients with anemia (categorical outcomes). Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php *Excluding iron which is discussed in Chapter 2.

Supplemental Table 31: Summary table of RCTs examining ESA vs. ESA in CKD patients with anemia (continuous outcomes). Supplemental Table 32: Summary table of adverse events in RCTs examining ESA vs. ESA in CKD patients with anemia (categorical outcomes). Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php *Excluding iron which is discussed in Chapter 2. Table 3 Potentially correctable versus non correctable factors involved in the anemia of CKD, in addition to ESA deficiency Easily correctable Potentially correctable Impossible to correct Absolute iron deficiency Vitamin B12/folate deficiency Hypothyroidism ACEi/ARB Non-adherence Infection/ inflammation Underdialysis Hemolysis Bleeding Hyperparathyroidism PRCA Malignancy Malnutrition Hemoglobinopathies Bone marrow disorders ACEi, angiotensin-converting enzyme inhibitor; ARB, angiotensin-receptor blocker; PRCA, pure red cell aplasia.

in B12/folate deficiency Hypothyroidism ACEi/ARB Non-adherence Infection/ inflammation Underdialysis Hemolysis Bleeding Hyperparathyroidism PRCA Malignancy Malnutrition Hemoglobinopathies Bone marrow disorders ACEi, angiotensin-converting enzyme inhibitor; ARB, angiotensin-receptor blocker; PRCA, pure red cell aplasia. Table 4 Practical approach in presence of ESA hyporesponsiveness Tests Finding and action 1. Check adherence If poor, attempt to improve (if self-injection) 2. Reticulocyte count If >130,000/μl, look for blood loss or hemolysis: endoscopy, colonoscopy, hemolysis screen Serum vitamin B12, folate If low, replenish Iron status If low, replenish iron Serum PTH If elevated, manage hyperparathyroidism Serum CRP If elevated, check for and treat infection or inflammation Underdialysis If underdialyzed, improve dialysis efficiency ACEi/ARB use If yes, consider reducing dose or discontinuing drug 3. Bone marrow biopsy Manage condition diagnosed e.g., dyscrasia, infiltration, fibrosis ACEi, angiotensin-converting enzyme inhibitor; ARB, angiotensin-receptor blocker; CRP, C-reactive protein; PTH, parathyroid hormone.

USE OF RED CELL TRANSFUSION IN CHRONIC ANEMIA Repeated transfusions or use of an erythropoiesis-stimulating agent (ESA) are treatment options for chronic anemia in CKD. The choice between these depends on their relative benefits and harms, which vary among patients. For example, patients with a previous stroke have the greatest absolute risk of ESA-related stroke,127 whereas multiparous women have the highest risk of allosensitization with transfusion.190, 191 Although the clinical importance of allosensitization is disputed, it may delay or reduce the possibility of future kidney transplantation. 4.1.1: When managing chronic anemia, we recommend avoiding, when possible, red cell transfusions to minimize the general risks related to their use. (1B) 4.1.2: In patients eligible for organ transplantation, we specifically recommend avoiding, when possible, red cell transfusions to minimize the risk of allosensitization. (1C) 4.1.3: When managing chronic anemia, we suggest that the benefits of red cell transfusions may outweigh the risks in patients in whom (2C): ESA therapy is ineffective (e.g., hemoglobinopathies, bone marrow failure, ESA resistance) The risks of ESA therapy may outweigh its benefits (e.g., previous or current malignancy, previous stroke) 4.1.4: We suggest that the decision to transfuse a CKD patient with non-acute anemia should not be based on any arbitrary Hb threshold, but should be determined by the occurrence of symptoms caused by anemia. (2C)

4.1.3: When managing chronic anemia, we suggest that the benefits of red cell transfusions may outweigh the risks in patients in whom (2C): ESA therapy is ineffective (e.g., hemoglobinopathies, bone marrow failure, ESA resistance) The risks of ESA therapy may outweigh its benefits (e.g., previous or current malignancy, previous stroke) 4.1.4: We suggest that the decision to transfuse a CKD patient with non-acute anemia should not be based on any arbitrary Hb threshold, but should be determined by the occurrence of symptoms caused by anemia. (2C) RATIONALE As with any treatment, the use of red cell transfusions should be considered in terms of the balance of benefit and harms. The primary benefit is in maintaining sufficient oxygen-carrying capacity and improvement in anemia-related symptoms.192 The harms are summarized in Tables 5 and 6 and discussed further below. This balance must also be considered alongside the balance between the benefits and harms of ESA therapy which is an alternative treatment for the anemia of CKD. The benefits and harms of ESA therapy are discussed in detail in Chapter 3, but, in summary, the benefits include improvement in anemia-related symptoms and reduced need for transfusion, and the most important harms are increased risk of stroke, thromboembolic events, and cancer progression or recurrence. When choosing between these two treatments for anemia in an individual, patient characteristics which influence the balance between benefits and harms for each treatment should be considered. These include history of stroke and previous or current cancer which place patients receiving ESA therapy at much higher absolute risk of these two problems. Conversely, patients potentially eligible for kidney transplantation have the greatest potential harm from transfusion, in terms of allosensitization,191, 193, 194 although the clinical importance of allosensitization is disputed. Previously transplanted patients and multiparous women seem to have the greatest absolute risk of allosensitization.190, 191

igible for kidney transplantation have the greatest potential harm from transfusion, in terms of allosensitization,191, 193, 194 although the clinical importance of allosensitization is disputed. Previously transplanted patients and multiparous women seem to have the greatest absolute risk of allosensitization.190, 191 A related issue is when should the decision to treat a patient with either an ESA or a transfusion be made? This decision is subtly different for the two types of treatment as ESAs may be used to avoid transfusion and therefore before the need for transfusion has arisen i.e., in a prophylactic sense. Furthermore, the magnitude of the potential harms of transfusion (e.g., from infection) and some of the benefits from ESAs (e.g., transfusion avoidance) is dependent on the threshold for transfusion. If that threshold is high (i.e., transfusion is reserved until symptoms become severe or the Hb reaches a very low level) the risks related to transfusion will be low and the benefit of ESA therapy in avoiding transfusions will be small. Unfortunately, there is no consensus about when transfusion is indicated although we do know that the rate of transfusion increases markedly when the Hb falls below 10 g/dl (100 g/l);122, 127 whether that simply reflects practice-patterns or represents clear clinical need is uncertain. The following trials give examples of transfusion rates in CKD 5D and CKD ND patients. The trial conducted by the Canadian Erythropoietin Study Group, published in 1990, enrolled 118 CKD 5HD patients Hb <9.0 g/dl (<90 g/l), 49 (42%) of whom were described as ‘transfusion-dependent'.122 The patients averaged approximately 7 transfusions each in the previous 12 months. These patients were randomized, equally, to 6 months treatment with placebo, erythropoietin with a target Hb 9.5–11.0 g/dl (95–110 g/l), or erythropoietin with a target Hb 11.5–13.0 g/dl (115–130 g/l). After 8 weeks, 23 patients in the placebo group received a blood-transfusion, compared with one in each of the two erythropoietin groups (for a gastrointestinal hemorrhage and following surgery).

erythropoietin with a target Hb 9.5–11.0 g/dl (95–110 g/l), or erythropoietin with a target Hb 11.5–13.0 g/dl (115–130 g/l). After 8 weeks, 23 patients in the placebo group received a blood-transfusion, compared with one in each of the two erythropoietin groups (for a gastrointestinal hemorrhage and following surgery). More recently, in the Trial to Reduce Cardiovascular Events with Aranesp Therapy (TREAT), published in 2009, 4038 patients with diabetes, CKD ND and anemia (Hb≤11.0 g/dl [≤110 g/l]), were randomized, equally, to darbepoetin-alfa with target Hb 13 g/dl (130 g/l) or to placebo, with ‘rescue' darbepoetin-alfa when Hb fell below 9.0 g/dl (90 g/l).127 Over a median follow-up of 29 months, 297/2012 (15%) patients randomized to darbepoetin-alfa and 496/2026 (25%) assigned to placebo received red cell transfusions (HR 0.56, 95% CI 0.49–0.65, P<0.001). We suggest that the decision to transfuse in the patient with non-acute anemia related to CKD should not be based upon any arbitrary Hb threshold and should, instead, be determined by the occurrence of symptoms and signs caused by anemia. We recognize that symptoms such as dyspnea and fatigue are non-specific, and that anemia-related symptoms may occur at different Hb levels in different patients.

elated to CKD should not be based upon any arbitrary Hb threshold and should, instead, be determined by the occurrence of symptoms and signs caused by anemia. We recognize that symptoms such as dyspnea and fatigue are non-specific, and that anemia-related symptoms may occur at different Hb levels in different patients. Risks of blood transfusion Risks associated with blood transfusion include transfusion errors, volume overload, hyperkalemia, citrate toxicity (leading to metabolic alkalosis and hypocalcemia), hypothermia, coagulopathy, immunologically-mediated transfusion reactions, including transfusion-related acute lung injury (TRALI), and iron overload, all of which are uncommon (Table 5).190, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207 Transmission of infections, although rare, is a major concern and this risk varies between countries (Table 6).208, 209, 210, 211 These complications are reviewed extensively elsewhere. The importance of human leukocyte antigen (HLA) sensitization is disputed and discussed in more detail below. HLA sensitization The risk of sensitization after blood transfusion has changed over time probably, at least in part, due to changes in blood transfusion practices and the use of more precise methods to measure allosensitization.

nt strip readings) were used in RCTs, these measures were translated to albumin excretion rates (AERs) per 24 h, recognizing that these converted values are approximations at best. Recommendations and suggestions for interventions based on albumin levels expressed in milligrams per 24 h can also be converted (Table 1). BP thresholds and targets Perhaps the most important questions for health care professionals are first, at what BP level should BP-lowering strategies be introduced in CKD patients (i.e., what is the BP treatment threshold?), and second, what BP levels should be aimed for (i.e., what is the BP treatment target?). Although the evidence base for the BP treatment threshold differs from the evidence base for the BP target, we could not find a robust justification to recommend different BP levels for these two parameters. Doing so might also lead to confusion, since we would be recommending two different BP levels possibly with two evidence ratings and would not be able to provide coherent advice for managing patients between the recommended threshold and target BPs.

Risks of blood transfusion Risks associated with blood transfusion include transfusion errors, volume overload, hyperkalemia, citrate toxicity (leading to metabolic alkalosis and hypocalcemia), hypothermia, coagulopathy, immunologically-mediated transfusion reactions, including transfusion-related acute lung injury (TRALI), and iron overload, all of which are uncommon (Table 5).190, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207 Transmission of infections, although rare, is a major concern and this risk varies between countries (Table 6).208, 209, 210, 211 These complications are reviewed extensively elsewhere. The importance of human leukocyte antigen (HLA) sensitization is disputed and discussed in more detail below. HLA sensitization The risk of sensitization after blood transfusion has changed over time probably, at least in part, due to changes in blood transfusion practices and the use of more precise methods to measure allosensitization. In the early 1980s, Opelz et al. examined the risk of sensitization in 737 CKD 5HD patients (Figures 3A and 3B), of whom 331 were followed prospectively (Figure 3C).190 Approximately 90% of all transfusions were given in the form of ‘packed cells' and antibodies were measured by the lymphocyte cytotoxicity test. Overall, 28% of patients followed prospectively developed HLA antibodies. Of these, 18% developed reactivity to 10–50% of the panel, 7% to 50–90%, and <3% to >90% of the panel after up to 20 transfusions (Figure 3C). Among men, 90% remained ‘unresponsive' (<10% antibody reactivity against the panel) and 10% developed reactivity to 10–50% of the panel (Figure 3C). In contrast, after 10 transfusions, only 60% of the women were ‘unresponsive,' 11% demonstrated 10–50% reactivity, 23% 51–90% reactivity, and 6% >90% reactivity (Figure 3C). These data suggested that the main drivers of HLA sensitization following red cell transfusion are previous pregnancies and previous transplantation. The data also suggested that men have a much lower risk of HLA sensitization following transfusion than women, and women with multiple pregnancies have a much greater risk of HLA sensitization than nulliparous women. However, more recent data from the US Renal Data System (USRDS) 2010 Annual Report,191 have challenged this assumption, suggesting that males receiving previous blood transfusions may also be at increased risk.

en, and women with multiple pregnancies have a much greater risk of HLA sensitization than nulliparous women. However, more recent data from the US Renal Data System (USRDS) 2010 Annual Report,191 have challenged this assumption, suggesting that males receiving previous blood transfusions may also be at increased risk. Studies performed in the last two decades showed that the risk of sensitization with blood transfusion is apparently lower than previously reported, with an overall response rate ranging from 2 to 21%.216, 217, 218 A possible, albeit controversial, explanation for this lower sensitization rate is that red cell transfusions in recent years are less immunogenic because they contain fewer leukocytes due to widespread use of blood filters. Other tentative conclusions from previous studies include the following: a) washed-red cells do not appear to be less immunogenic than non-washed red cells;190 b) no consistent reduction in sensitization has been demonstrated with donor-specific217 and HLA-DR matched transfusions;219 c) higher numbers of blood transfusions have been associated with an increased risk of sensitization in some studies220, 221 but not in others.190, 222

s immunogenic than non-washed red cells;190 b) no consistent reduction in sensitization has been demonstrated with donor-specific217 and HLA-DR matched transfusions;219 c) higher numbers of blood transfusions have been associated with an increased risk of sensitization in some studies220, 221 but not in others.190, 222 However, more recent data from the USRDS indicates that risk of sensitization with blood transfusions is substantial. For example, compared with patients who have never received a blood transfusion, patients who received transfusions have an odds ratio of having panel reactive antibody (PRA) >80% of 2.38.191 Interestingly, in this analysis the risk of being highly sensitized at the time of transplantation was higher for men than for women.

xample, compared with patients who have never received a blood transfusion, patients who received transfusions have an odds ratio of having panel reactive antibody (PRA) >80% of 2.38.191 Interestingly, in this analysis the risk of being highly sensitized at the time of transplantation was higher for men than for women. Effect of leukocyte-reduced blood transfusions on sensitization Although, leukocytes may be a contributor to, if not the cause of, a number of adverse consequences of blood transfusion, including immunologically-mediated effects, infectious disease transmission, and reperfusion injury, leukoreduction of blood products does not decrease sensitization in previously transplanted or in potential future kidney transplant candidates.223, 224, 225 One recent study reported that male patients awaiting their first organ transplant had a fourfold increased risk of developing HLA antibody if they had been previously transfused when compared with those who did not have a history of a transfusion.226 Thus, transfusion in the post-leukodepletion era still continues to pose a significant risk of sensitization. A possible reason for this finding is that the number of HLA molecules contributed by the red cells is comparable to that of leukocytes.227

en compared with those who did not have a history of a transfusion.226 Thus, transfusion in the post-leukodepletion era still continues to pose a significant risk of sensitization. A possible reason for this finding is that the number of HLA molecules contributed by the red cells is comparable to that of leukocytes.227 Association between sensitization and delay in organ transplantation According to USRDS data reported in 2010, the mean wait-time to transplant for patients listed between 1991 and 2008 was an average of 2 months longer for transfused than non-transfused patients in the United States.191 Increased PRA titers, whether due to blood transfusions or other factors, were associated with a longer wait to find a compatible donor and may have completely precluded transplantation in some patients. Non-sensitized patients (0% PRA at the time of listing) had the shortest wait-time (median of 2.5 years in 2005) while those with a PRA of 1–19% and 20–79% had median wait-times of 2.9 and 4.3 years, respectively. Highly sensitized patients (≥80% PRA) waited the longest and in these patients a median wait-time could not be calculated for patients listed in 2005. As a result of the delay in finding compatible donors in patients with PRA ≥80%, the percentage of these patients increased from 7.5% at listing to 13.3% five years after listing.

sitized patients (≥80% PRA) waited the longest and in these patients a median wait-time could not be calculated for patients listed in 2005. As a result of the delay in finding compatible donors in patients with PRA ≥80%, the percentage of these patients increased from 7.5% at listing to 13.3% five years after listing. Not being transplanted, or having to wait longer for transplantation, is associated with lower survival.228, 229 Receiving a transfusion while on the transplant wait list is associated with a nearly 5-fold higher risk of dying while on the wait list in the first five years, and an 11% reduction in the likelihood of receiving a transplant within the first five years.191, 230 In transplanted patients, the presence of preformed HLA antibodies is associated with an increased risk of early and late graft loss.193, 194, 231, 232 Recent data also suggest that pre-existing donor-specific HLA antibodies identified by a Luminex single-antigen assay at the time of transplantation are associated with a higher incidence of antibody-mediated rejection and inferior graft survival.233 URGENT TREATMENT OF ANEMIA 4.2: In certain acute clinical situations, we suggest patients are transfused when the benefits of red cell transfusions outweigh the risks; these include (2C): When rapid correction of anemia is required to stabilize the patient's condition (e.g., acute hemorrhage, unstable coronary artery disease) When rapid pre-operative Hb correction is required

URGENT TREATMENT OF ANEMIA 4.2: In certain acute clinical situations, we suggest patients are transfused when the benefits of red cell transfusions outweigh the risks; these include (2C): When rapid correction of anemia is required to stabilize the patient's condition (e.g., acute hemorrhage, unstable coronary artery disease) When rapid pre-operative Hb correction is required RATIONALE In certain urgent clinical situations, red cell transfusion may be needed for the immediate correction of anemia. These include acute severe hemorrhage and other clinical problems caused by, or exacerbated by, anemia, such as acute myocardial ischemia. When urgent surgery is required, transfusion may also be given to achieve rapid preoperative correction of Hb. The Hb threshold for transfusion in this situation is uncertain but we suggest that this treatment be considered if the Hb is <7 g/dl (<70 g/l). Table 7 and Figure 4 summarize the approaches to the use of red cell transfusions in patients with CKD. RESEARCH RECOMMENDATIONS There is a lack of randomized controlled trials on the use of blood transfusions as a primary intervention in patients with anemia and CKD. Given the logistical difficulties in conducting such trials, it is likely that observational data will continue to predominate in this therapeutic area.

Table 7 and Figure 4 summarize the approaches to the use of red cell transfusions in patients with CKD. RESEARCH RECOMMENDATIONS There is a lack of randomized controlled trials on the use of blood transfusions as a primary intervention in patients with anemia and CKD. Given the logistical difficulties in conducting such trials, it is likely that observational data will continue to predominate in this therapeutic area. Future research should include: Prospective observational data collection on the use of red cell transfusions in CKD patients, particularly dialysis patients, including the reason(s) for transfusion, intent to list for future kidney transplantation, likelihood of receiving a kidney transplant, and graft outcomes. Prospective observational evaluation of the impact of red cell transfusions on the level of HLA sensitization. Given a striking disparity in the use of blood transfusions between the US and Europe, Canada and Australia in the TREAT study, and between the US and Europe in the Phase 3 peginesatide clinical trial program, further research is needed to ascertain the ‘drivers' for transfusion in CKD patients. Is this related to practice patterns or a real higher clinical need for transfusions in the US?

US and Europe, Canada and Australia in the TREAT study, and between the US and Europe in the Phase 3 peginesatide clinical trial program, further research is needed to ascertain the ‘drivers' for transfusion in CKD patients. Is this related to practice patterns or a real higher clinical need for transfusions in the US? DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature.

ion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. Figure 3 Lymphocytotoxic antibody reactivity against random donor test panel in relation to the number of blood transfusions. Fractions of patients reacting against <10%, 10 to 50%, 51 to 90% and >90% of the panel donors are plotted. All 737 patients were on chronic hemodialysis, waiting for a first kidney transplant. Numbers of patients after 2, 5, 10, 15, and 20 transfusions are indicated at top of graphs. (A) Male and female patients. (B) Females patients separated by the number of previous pregnancies. (C) Lymphocytotoxic antibodies in patients who were studied prospectively throughout the course of treatment. Reprinted from Opelz G, Graver B, Mickey MR et al. Lymphocytotoxic antibody responses to transfusions in potential kidney transplant recipients. Transplantation 1981; 32(3): 177–183 (ref. 190) with permission from Lippincott Williams & Wilkins; accessed http://journals.lww.com/transp lantjournal/Abstract/1981/0900 0/Lymphocytoxic_Antibody_Respo nses_to_Transfusions.2.aspx Figure 4 Algorithms for red cell transfusion use in CKD patients. ESA, erythropoiesis-stimulating agent; Hb, hemoglobin.

Figure 3 Lymphocytotoxic antibody reactivity against random donor test panel in relation to the number of blood transfusions. Fractions of patients reacting against <10%, 10 to 50%, 51 to 90% and >90% of the panel donors are plotted. All 737 patients were on chronic hemodialysis, waiting for a first kidney transplant. Numbers of patients after 2, 5, 10, 15, and 20 transfusions are indicated at top of graphs. (A) Male and female patients. (B) Females patients separated by the number of previous pregnancies. (C) Lymphocytotoxic antibodies in patients who were studied prospectively throughout the course of treatment. Reprinted from Opelz G, Graver B, Mickey MR et al. Lymphocytotoxic antibody responses to transfusions in potential kidney transplant recipients. Transplantation 1981; 32(3): 177–183 (ref. 190) with permission from Lippincott Williams & Wilkins; accessed http://journals.lww.com/transp lantjournal/Abstract/1981/0900 0/Lymphocytoxic_Antibody_Respo nses_to_Transfusions.2.aspx Figure 4 Algorithms for red cell transfusion use in CKD patients. ESA, erythropoiesis-stimulating agent; Hb, hemoglobin. Table 5 Estimated risk associated with blood transfusions per unit transfused Adverse event Estimated risk* Immunological Fever/allergic reactions 1 in 100–200a,b Hemolytic reaction 1 in 6000b Transfusion-related acute lung injury (TRALI) 1 in 12,350a Anaphylaxis 1 in 50,000b Fatal hemolysis 1 in 1,250,000a Graft versus host disease (GVHD) Rare Other Mistransfusion 1 in 14,000–19,000c *United States data. a Data from Carson JL et al.212 b Data from Klein.213 c Data from Klein HG et al.214

Table 5 Estimated risk associated with blood transfusions per unit transfused Adverse event Estimated risk* Immunological Fever/allergic reactions 1 in 100–200a,b Hemolytic reaction 1 in 6000b Transfusion-related acute lung injury (TRALI) 1 in 12,350a Anaphylaxis 1 in 50,000b Fatal hemolysis 1 in 1,250,000a Graft versus host disease (GVHD) Rare Other Mistransfusion 1 in 14,000–19,000c *United States data. a Data from Carson JL et al.212 b Data from Klein.213 c Data from Klein HG et al.214 Table 6 Estimated risk of transfusion-related infections per unit transfused Potential transfusion-related risks Estimated risk* Hepatitis B 1 in 282,000–1 in 357,000a West Nile virus 1 in 350,000b Death from bacterial sepsis 1 in 1,000,000b Hepatitis C 1 in 1,149,000a Human immunodeficiency virus (HIV) 1 in 1,467,000a *United States data. a Data from Carson JL et al.212 b Data from Rawn J.215

Table 6 Estimated risk of transfusion-related infections per unit transfused Potential transfusion-related risks Estimated risk* Hepatitis B 1 in 282,000–1 in 357,000a West Nile virus 1 in 350,000b Death from bacterial sepsis 1 in 1,000,000b Hepatitis C 1 in 1,149,000a Human immunodeficiency virus (HIV) 1 in 1,467,000a *United States data. a Data from Carson JL et al.212 b Data from Rawn J.215 Table 7 Indications for blood transfusions Indication Comments When rapid correction of anemia is required to stabilize the patient's condition (e.g., acute hemorrhage, unstable myocardial ischemia) • Red cell transfusion in patients with acute hemorrhage is indicated in the following situations: a) rapid acute hemorrhage without immediate control of bleeding; b) estimated blood loss >30–40% of blood volume (1500–2000 ml) with symptoms of severe blood loss; c) estimated blood loss <25–30% blood volume with no evidence of uncontrolled hemorrhage, if signs of hypovolemia recur despite colloid/crystalloid resuscitation; d) in patients with co-morbid factors, transfusions may be necessary with lesser degrees of blood loss.234 • Studies evaluating the importance of anemia and the role of transfusion in the setting of an acute coronary syndrome (i.e., unstable angina, myocardial infarction) have reached differing conclusions. • The American College of Cardiology/American Heart Association and American College of Chest Physicians guidelines do not make any recommendations concerning the potential benefit or risk of blood transfusion in the setting of an acute coronary syndrome.235, 236 However, in a review of clinical trials of patients with a non-ST elevation acute coronary syndrome, the risk of cardiovascular mortality, nonfatal myocardial infarction, or recurrent ischemia at 30 days was significantly higher in patients with a Hb concentration below 11 g/dl (110 g/l) than those with a Hb ≥11 g/dl (≥110 g/l).237 • Although anemia occurs frequently in patients with heart failure, limited data are available on treatment of anemia in this population. • Correction of anemia is not an evidence-based therapy in heart failure as noted in the 2006 Heart Failure Society of America guidelines, 2012 European Society of Cardiology (ESC) guidelines, and 2009 American College of Cardiology/American Heart Association guidelines.238, 239, 240 • Therefore, the general indications for red cell transfusion apply to patients with heart failure; however, careful attention must be paid to volume status.

rica guidelines, 2012 European Society of Cardiology (ESC) guidelines, and 2009 American College of Cardiology/American Heart Association guidelines.238, 239, 240 • Therefore, the general indications for red cell transfusion apply to patients with heart failure; however, careful attention must be paid to volume status. When rapid pre-operative Hb correction is required • Criteria have been proposed for perioperative transfusions.234 These are generally not recommended when the Hb is ≥10 g/dl (≥100 g/l) in otherwise healthy subjects, but should be given when the Hb is less than 7 g/dl (70 g/l). • When Hb concentration is less than 7 g/dl (70 g/l) and the patient is otherwise stable, 2 units of red cells should be transfused and the patient's clinical status and circulating Hb should be reassessed. • High-risk patients (>65 years and/or those with cardiovascular or respiratory disease) may tolerate anemia poorly, and may be transfused when Hb concentration is less than 8 g/dl (80 g/l). • For Hb concentration between 7 and 10 g/dl (70 and 100 g/l), the correct strategy is unclear. When symptoms and signs related to anemia are present in patients in whom ESA therapy is ineffective (e.g., bone marrow failure, hemoglobinopathies, ESA resistance) • Patients with chronic anemia (e.g., bone marrow failure syndromes) may be dependent upon red cell replacement over a period of months or years, which can lead to iron overload. • Approximately 200 mg of iron are delivered per unit of red cells; this iron is released when Hb from the transfused red cells is metabolized after red cell death. • Given the progressive loss of red cell viability which occurs during storage, the “freshest-available” units should be selected in order to maximize post-transfusion survival. • Hemosiderosis can produce organ damage when the total iron delivered approaches 15 to 20 grams, the amount of iron in 75 to 100 units of red cells. • The issue of red cell transfusion in patients with acquired or congenital hemolytic anemia is more complex. When symptoms and signs related to anemia are present in patients in whom the risks of ESA therapy may outweigh the benefits • ESAs should be used with great caution, if at all, in CKD patients with active malignancy, a history of malignancy, or prior history of stroke. CKD, chronic kidney disease; ESA, erythropoiesis-stimulating agent; Hb, hemoglobin.

AIM The overall aim of the project was to develop an evidence-based clinical practice guideline for management of anemia and chronic kidney disease (CKD). The guideline consists of recommendations, rationale statements and a summary of systematically generated evidence on relevant predefined clinical topics. OVERVIEW PROCESS Guideline development process included the following sequential and concurrent steps: Appointing Work Group members and Evidence Review Team (ERT). Discussing process, methods, and results. Developing and refining topics. Identifying populations, interventions or predictors, and outcomes of interest. Selecting topics for systematic evidence review. Standardizing quality assessment methodology. Developing and implementing literature search strategies. Screening abstracts and retrieving full text articles based on predefined eligibility criteria. Creating data extraction forms. Data extracting and performing critical appraisal of the literature. Grading the methodology and outcomes in individual studies. Tabulating data from individual studies into summary tables. Grading quality of evidence for each outcome across studies, and assessing the overall quality of evidence across outcomes with the aid of evidence profiles. Grading the strength of recommendations based on the quality of evidence and other considerations. Finalizing guideline recommendations and supporting rationale statements. Sending the guideline draft for peer review to the KDIGO Board of Directors in June 2011, and for public review in September 2011. Publishing the final version of the guideline.

Grading the strength of recommendations based on the quality of evidence and other considerations. Finalizing guideline recommendations and supporting rationale statements. Sending the guideline draft for peer review to the KDIGO Board of Directors in June 2011, and for public review in September 2011. Publishing the final version of the guideline. The Work Group, KDIGO Co-Chairs, ERT, and KDIGO support staff met for two 2-day meetings for training in the guideline development process, topic discussion, and consensus development. Commissioning of work group and evidence review team KDIGO Co-Chairs appointed the Work Group Co-chairs. Work Group Co-Chairs then assembled the Work Group consisting of domain experts, including individuals with expertise in internal medicine, adult and pediatric nephrology, cardiology, hematology, oncology, hypertension, pathology, pharmacology, epidemiology and endocrinology. Tufts Center for Kidney Disease Guideline Development and Implementation at Tufts Medical Center in Boston, Massachusetts, USA was contracted to conduct systematic evidence review and provide expertise in guideline development methodology. The ERT consisted of physician-methodologists with expertise in nephrology, a project coordinator and manager, and a research assistant. The ERT instructed and advised Work Group members in all steps of literature review, critical literature appraisal, and guideline development. The Work Group and the ERT collaborated closely throughout the project.

methodologists with expertise in nephrology, a project coordinator and manager, and a research assistant. The ERT instructed and advised Work Group members in all steps of literature review, critical literature appraisal, and guideline development. The Work Group and the ERT collaborated closely throughout the project. Defining scope and topics Work Group Co-Chairs first defined the overall scope and goals of the guideline. Work Group Co-Chairs then drafted a preliminary list of topics and key clinical questions. In light of new evidence, it was decided that an update of the topics presented in the 2006 and 2007 KDOQI guidelines would be the best approach. The Work Group and ERT further developed and refined each topic, specified screening criteria, literature search strategies, and data extraction forms (Table 8).

l questions. In light of new evidence, it was decided that an update of the topics presented in the 2006 and 2007 KDOQI guidelines would be the best approach. The Work Group and ERT further developed and refined each topic, specified screening criteria, literature search strategies, and data extraction forms (Table 8). Establishing the process for guideline development The ERT performed literature searches, organized abstract and article screening. The ERT also coordinated the methodological and analytic process of the report, defined and standardized the methodology of performing literature searches, data extraction, and summarizing the evidence. Throughout the project, the ERT offered suggestions for guideline development, led discussions on systematic review, literature searches, data extraction, assessment of quality and applicability of articles, evidence synthesis, grading of evidence and guideline recommendations, and consensus development. The Work Group took the primary role of writing the guidelines and rationale statements and retained final responsibility for the content of the guideline statements and the accompanying narrative.

plicability of articles, evidence synthesis, grading of evidence and guideline recommendations, and consensus development. The Work Group took the primary role of writing the guidelines and rationale statements and retained final responsibility for the content of the guideline statements and the accompanying narrative. The Work Group Co-Chairs prepared the first draft of the scope of work document as a series of topics to be considered by Work Group members. The scope of work document was based primarily on the existing KDOQI guidelines on anemia. At their first two-day meeting, Work Group members revised the initial working document to include all topics of interest to the Work Group. The inclusive, combined set of questions formed the basis for the deliberation and discussion that followed. The Work Group strove to ensure that all topics deemed clinically relevant and worthy of review were identified and addressed. Formulating questions of interest Questions of interest were formulated according to the PICODD (Population, Intervention, Comparator, Outcome, study Design and Duration of follow up) criteria. Details of the PICODD criteria are presented in Table 8. Ranking of outcomes The Work Group ranked outcomes of interest based on their importance for informing clinical decision making (Table 9). Mortality, cardiovascular mortality, cardiovascular events and ESRD outcomes were graded as ‘critical,' transfusion and QoL outcomes were graded as ‘high,' and all other outcomes were graded as ‘moderate.'

Work Group ranked outcomes of interest based on their importance for informing clinical decision making (Table 9). Mortality, cardiovascular mortality, cardiovascular events and ESRD outcomes were graded as ‘critical,' transfusion and QoL outcomes were graded as ‘high,' and all other outcomes were graded as ‘moderate.' Literature searches and article selection The Work Group sought to build on the evidence base and topics addressed in the previous Kidney Disease Outcomes Quality Initiative (KDOQI) clinical practice guidelines and clinical practice recommendations for anemia in chronic kidney disease in 2006 as well as the KDOQI clinical practice guidelines and clinical practice recommendations for anemia in chronic kidney disease 2007 update of hemoglobin target. Modules were created for randomized controlled trials (RCTs), kidney disease, anemia, and erythropoietin, transfusion, iron deficiency, and adjuvant search terms. The search terms were then limited to years 2006–2010 for studies related to anemia interventions. For transfusion the literature search was conducted from 1989–2010. A separate search was run for observational studies on iron overload and hemoglobin status as predictors for clinical outcomes (See Appendix 1 online).

. The search terms were then limited to years 2006–2010 for studies related to anemia interventions. For transfusion the literature search was conducted from 1989–2010. A separate search was run for observational studies on iron overload and hemoglobin status as predictors for clinical outcomes (See Appendix 1 online). The searches were run in MEDLINE, Cochrane Central Register of Controlled Clinical Trials and Cochrane Database of Systematic Reviews. The initial search for RCTs was conducted in April 2010 and subsequently updated in October of 2010. The search for observational studies was later conducted in September 2010. The search yield was also supplemented by articles provided by Work Group members through March 2012. MEDLINE search results were screened by members of the ERT for relevance using pre-defined eligibility criteria.

y updated in October of 2010. The search for observational studies was later conducted in September 2010. The search yield was also supplemented by articles provided by Work Group members through March 2012. MEDLINE search results were screened by members of the ERT for relevance using pre-defined eligibility criteria. The total yield from the search was 4,334 abstracts for RCTs and 3,717 abstracts for observational studies. Fifty-six abstracts and 53 full texts from RCTs were accepted and 97 abstracts and 21 full texts from observational studies were accepted. Journal articles reporting original data, meta-analyses or systematic reviews were selected for evidence review. Editorials, letters, abstracts, unpublished reports and articles published in non-peer reviewed journals were not included. The Work Group also decided to exclude publications from journal supplements because of potential differences in the process of how they get solicited, selected, reviewed and edited compared to peer-reviewed publications. The overall search yield along with the number of abstracts identified and articles reviewed is presented in Table 10. Data extraction Fifty-three full text articles from RCTs were extracted by the ERT. The ERT, in consultation with the Work Group, designed forms to capture data on design, methodology, sample characteristics, interventions, comparators, outcomes, results and limitations of individual studies. Methodology and outcomes were also systematically graded (see the section on grading below) and recorded during the data extraction process.

the Work Group, designed forms to capture data on design, methodology, sample characteristics, interventions, comparators, outcomes, results and limitations of individual studies. Methodology and outcomes were also systematically graded (see the section on grading below) and recorded during the data extraction process. Summary tables Summary tables were developed for each comparison of interest. Studies included in the evidence base for the KDOQI clinical practice guidelines on Anemia in CKD and update of hemoglobin target were also incorporated if they fulfilled the inclusion criteria for the current guideline. Summary tables contain outcomes of interest, relevant population characteristics, description of intervention and comparator, results, and quality grading for each outcome. Categorical and continuous outcomes were summarized separately. Work Group members proofed all summary table data and quality assessments. Summary tables will be available at www.kdigo.org/clinical_practice_guidelines/anemia.php.

description of intervention and comparator, results, and quality grading for each outcome. Categorical and continuous outcomes were summarized separately. Work Group members proofed all summary table data and quality assessments. Summary tables will be available at www.kdigo.org/clinical_practice_guidelines/anemia.php. Evidence profiles Evidence profiles were constructed to assess and record quality grading and description of effect for each outcome across studies, and quality of overall evidence and description of net benefits or harms of intervention or comparator across all outcomes. These profiles aim to make the evidence synthesis process transparent. Decisions in the evidence profiles were based on data from the primary studies listed in corresponding summary tables, and on judgments of the ERT and the Work Group. When the body of evidence for a particular comparison of interest consisted of only one study, the summary table provided the final level of synthesis and evidence profile was not generated. Each evidence profile was initially constructed by the ERT and then reviewed, edited and approved by the Work Group.

Work Group. When the body of evidence for a particular comparison of interest consisted of only one study, the summary table provided the final level of synthesis and evidence profile was not generated. Each evidence profile was initially constructed by the ERT and then reviewed, edited and approved by the Work Group. Grading of quality of evidence for outcomes of individual studies Methodological quality Methodological quality (internal validity) refers to the design, conduct, and reporting of outcomes of a clinical study. Previously devised three-level classification system for quality assessment was used to grade the overall study quality and quality for all relevant outcomes in the study (Table 11). Variations of this system have been used in most KDOQI and all KDIGO guidelines and have been recommended for the US Agency for Healthcare Research and Quality Evidence-based Practice Center program (http://effectivehealthcare.ahrq.gov/repFiles/2007_10DraftMethodsGuide.pdf). Each study was given an overall quality grade based on its design, methodology (randomization, allocation, blinding, definition of outcomes, appropriate use of statistical methods etc), conduct (drop-out percentage, outcome assessment methodologies, etc) and reporting (internal consistency, clarity, thoroughness/precision, etc). Each reported outcome was then evaluated and given an individual grade depending on the quality of reporting and methodological issues specific to that outcome. However, the quality grade of an individual outcome could not exceed the quality grade for the overall study.

nsistency, clarity, thoroughness/precision, etc). Each reported outcome was then evaluated and given an individual grade depending on the quality of reporting and methodological issues specific to that outcome. However, the quality grade of an individual outcome could not exceed the quality grade for the overall study. Rating the quality of evidence and the strength of guideline recommendations A structured approach, based on GRADE241, 242, 243 and facilitated by the use of evidence profiles was used in order to grade the quality of the overall evidence and the strength of recommendations. For each topic, the discussion on grading of the quality of the evidence was led by the ERT, and the discussion regarding the strength of the recommendations was led by the Work Group Chairs. The ‘strength of a recommendation' indicates the extent to which one can be confident that adherence to the recommendation will do more good than harm. The ‘quality of a body of evidence' refers to the extent to which our confidence in an estimate of effect is sufficient to support a particular recommendation.242

airs. The ‘strength of a recommendation' indicates the extent to which one can be confident that adherence to the recommendation will do more good than harm. The ‘quality of a body of evidence' refers to the extent to which our confidence in an estimate of effect is sufficient to support a particular recommendation.242 Grading the quality of evidence for each outcome Following GRADE, the quality of a body of evidence pertaining to a particular outcome of interest was initially categorized based on study design. For questions of interventions, the initial quality grade was ‘High' when the body of evidence consisted of randomized controlled trials; ‘Low', if it consisted of observational studies; or ‘Very Low', if it consisted of studies of other study designs. For questions of interventions, the Work Group decided to use only randomized controlled trials. The grade for the quality of evidence for each intervention/outcome pair was then lowered if there were serious limitations to the methodological quality of the aggregate of studies, if there were important inconsistencies in the results across studies, if there was uncertainty about the directness of evidence including limited applicability of the findings to the population of interest, if the data were imprecise (a low event rate [0 or 1 event] in either arm or confidence interval spanning a range <0.5 to >2.0) or sparse (only 1 study or total N<100), or if there was thought to be a high likelihood of bias. The final grade for the quality of the evidence for an intervention/outcome pair could be one of the following four grades: ‘High', ‘Moderate', ‘Low' or ‘Very Low' (Table 12).

idence interval spanning a range <0.5 to >2.0) or sparse (only 1 study or total N<100), or if there was thought to be a high likelihood of bias. The final grade for the quality of the evidence for an intervention/outcome pair could be one of the following four grades: ‘High', ‘Moderate', ‘Low' or ‘Very Low' (Table 12). Grading the overall quality of evidence The quality of the overall body of evidence was then determined based on the quality grades for all outcomes of interest, taking into account explicit judgments about the relative importance of each outcome. The resulting four final categories for the quality of overall evidence were: ‘A', ‘B', ‘C' or ‘D' (Table 13). Assessment of the net health benefit across all important clinical outcomes The net health benefit was determined based on the anticipated balance of benefits and harms across all clinically important outcomes (Table 14). The assessment of net benefit was affected by the judgment of the Work Group and the ERT.

Grading the overall quality of evidence The quality of the overall body of evidence was then determined based on the quality grades for all outcomes of interest, taking into account explicit judgments about the relative importance of each outcome. The resulting four final categories for the quality of overall evidence were: ‘A', ‘B', ‘C' or ‘D' (Table 13). Assessment of the net health benefit across all important clinical outcomes The net health benefit was determined based on the anticipated balance of benefits and harms across all clinically important outcomes (Table 14). The assessment of net benefit was affected by the judgment of the Work Group and the ERT. Grading the strength of the recommendations The strength of a recommendation is graded as Level 1 or Level 2. Table 15 shows the KDIGO nomenclature for grading the strength of a recommendation and the implications of each level for patients, clinicians and policy makers. Recommendations can be for or against doing something. Table 16 shows that the strength of a recommendation is determined not just by the quality of the evidence, but also by other, often complex judgments regarding the size of the net medical benefit, values and preferences, and costs. Formal decision analyses including cost analysis were not conducted.

t doing something. Table 16 shows that the strength of a recommendation is determined not just by the quality of the evidence, but also by other, often complex judgments regarding the size of the net medical benefit, values and preferences, and costs. Formal decision analyses including cost analysis were not conducted. Ungraded statements This category was designed to allow the Work Group to issue general advice. Typically an ungraded statement meets the following criteria: it provides guidance based on common sense; it provides reminders of the obvious; it is not sufficiently specific to allow application of evidence to the issue and therefore it is not based on systematic evidence review. Common examples include recommendations about frequency of testing, referral to specialists, and routine medical care. We strove to minimize the use of ungraded recommendations. This grading scheme with two levels for the strength of a recommendation together with four levels of grading the quality of the evidence, and the option of an ungraded statement for general guidance was adopted by the KDIGO Board in December 2008. The Work Group took the primary role of writing the recommendations and rationale statements and retained final responsibility for the content of the guideline statements and the accompanying narrative. The ERT reviewed draft recommendations and grades for consistency with the conclusions of the evidence review.

2008. The Work Group took the primary role of writing the recommendations and rationale statements and retained final responsibility for the content of the guideline statements and the accompanying narrative. The ERT reviewed draft recommendations and grades for consistency with the conclusions of the evidence review. Format for guideline recommendations Each chapter contains one or more specific recommendations. Within each recommendation, the strength of recommendation is indicated as level 1 or level 2 and the quality of the supporting evidence is shown as A, B, C or D. These are followed by a brief background with relevant definitions of terms and the rationale summarizing the key points of the evidence base and narrative supporting the recommendation. Where appropriate, research recommendations are suggested for future research to resolve current uncertainties. Limitations of approach While the literature searches were intended to be comprehensive, they were not exhaustive. MEDLINE was the only database searched. Hand searches of journals were not performed, and review articles and textbook chapters were not systematically searched. However, important studies known to domain experts that were missed by the electronic literature searches were added to retrieved articles and reviewed by the Work Group.

the only database searched. Hand searches of journals were not performed, and review articles and textbook chapters were not systematically searched. However, important studies known to domain experts that were missed by the electronic literature searches were added to retrieved articles and reviewed by the Work Group. Summary of the methodological review process Several tools and checklists have been developed to assess the quality of the methodological process for systematic review and guideline development. These include the Appraisal of Guidelines for Research and Evaluation (AGREE) criteria,244 the Conference on Guideline Standardization (COGS) checklist,245 and the Institute of Medicine's recent Standards for Systematic Reviews246 and Clinical Practice Guidelines We Can Trust.247 Table 17 and Appendix 2 online show, respectively, the COGS criteria which correspond to the AGREE checklist and the Institute of Medicine standards, and how each one of them is addressed in this guideline. SUPPLEMENTARY MATERIAL Appendix 1: Online search strategies. Appendix 2: Concurrence with Institute of Medicine standards for systematic reviews and for guidelines. Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php

Summary of the methodological review process Several tools and checklists have been developed to assess the quality of the methodological process for systematic review and guideline development. These include the Appraisal of Guidelines for Research and Evaluation (AGREE) criteria,244 the Conference on Guideline Standardization (COGS) checklist,245 and the Institute of Medicine's recent Standards for Systematic Reviews246 and Clinical Practice Guidelines We Can Trust.247 Table 17 and Appendix 2 online show, respectively, the COGS criteria which correspond to the AGREE checklist and the Institute of Medicine standards, and how each one of them is addressed in this guideline. SUPPLEMENTARY MATERIAL Appendix 1: Online search strategies. Appendix 2: Concurrence with Institute of Medicine standards for systematic reviews and for guidelines. Supplementary material is linked to the online version of the paper at http://www.kdigo.org/clinical_practice_guidelines/anemia.php Table 8 Systematic review topics and screening criteria Identifying why, when and which patients to treat for anemia and iron deficiency Population All CKD stages for longitudinal, cross-sectional or RCTs. Any population for systematic reviews Intervention RBC transfusion, Iron (all forms, routes of administration, dosages), ESA (all forms, dosages, targets, protocols, schedules, etc), pharmacological and non-pharmacological adjuvants to ESA, Hb or iron status Comparator Other interventions, “no” interventions, different forms, routes of administration, dosages, targets, protocols, schedules, etc. Outcomes All-cause mortality, Cardiovascular events, ESRD, Quality of life, Progression of kidney disease, Transfusions, Major symptoms Study design RCTs, Large longitudinal (prospective or retrospective) observational studies or cross sectional studies with multivariate analyses N≥50 per arm

protocols, schedules, etc. Outcomes All-cause mortality, Cardiovascular events, ESRD, Quality of life, Progression of kidney disease, Transfusions, Major symptoms Study design RCTs, Large longitudinal (prospective or retrospective) observational studies or cross sectional studies with multivariate analyses N≥50 per arm Evaluating anemia treatment, including treatment resistance Population Adults and children with CKD, any stage and any comorbidity (including cancer, CVD, etc.) Intervention RBC transfusions; Iron (all forms, routes of administration, dosages), ESA (all forms, dosages, targets, protocols, etc), pharmacological and non-pharmacological adjuvants to ESA including L-carnitine, vitamin C, androgens, pentoxifylline; other interventions used to treat or enhance the treatment of anemia or anemia-related symptoms Comparator Other interventions, “no” interventions, different forms, routes of administration, dosages, targets, protocols, schedules, etc. Outcomes Death, Cardiac events, Stroke, CKD progression, Quality of life, Thromboembolic events, Pulmonary embolism, Symptomatic deep vein thrombosis, Loss of vascular access, Transfusion requirements, Cognitive function, Sexual function, Other similar quality of life measures, Objective physical function tests, Infections, Loss of transplant eligibility due to antibody sensitization, Antibody sensitization, New cancer or progression of existing cancer, Seizure, Other clinically important adverse events, ESA dose: for comparisons of different ESA regimens and for iron and adjuvant interventions, Achieved Hb/Hb variability for comparisons of different ESA regimens and for iron and adjuvant interventions Study Design RCTs N≥50 per arm Minimum follow-up duration: 6 months CKD, chronic kidney disease; CVD, cardiovascular disease; ESA, erythropoiesis-stimulating agent; ESRD, end-stage renal disease; Hb, hemoglobin; RBC, red blood cell; RCT, randomized controlled trial.

ESA regimens and for iron and adjuvant interventions Study Design RCTs N≥50 per arm Minimum follow-up duration: 6 months CKD, chronic kidney disease; CVD, cardiovascular disease; ESA, erythropoiesis-stimulating agent; ESRD, end-stage renal disease; Hb, hemoglobin; RBC, red blood cell; RCT, randomized controlled trial. Table 9 Hierarchy of importance of outcomes Hierarchya Outcomesb Critical importance Mortality, Cardiovascular mortality, Cardiovascular events, ESRD High importance Transfusion, Quality of life Moderate importance Hb (categorical and continuous), ESA dose (categorical and continuous), adverse events ESA, erythropoiesis-stimulating agent; ESRD, end-stage renal disease; Hb, hemoglobin. a Outcomes of lesser importance are excluded from review. b This categorization was the consensus of the Work Group for the purposes of this guideline only. The lists are not meant to reflect outcome ranking for other areas of kidney disease management. The Work Group acknowledges that not all clinicians, patients or families, or societies would rank all outcomes the same. Table 10 Literature search yield of primary articles for systematic review topics Total abstracts from updated search Abstracts accepted Full text accepted Full text extracted Articles in summary tables 4,334 RCT 56 53 53 31 3,717 Observational 97 21 21 21 RCT, randomized controlled trial.

b This categorization was the consensus of the Work Group for the purposes of this guideline only. The lists are not meant to reflect outcome ranking for other areas of kidney disease management. The Work Group acknowledges that not all clinicians, patients or families, or societies would rank all outcomes the same. Table 10 Literature search yield of primary articles for systematic review topics Total abstracts from updated search Abstracts accepted Full text accepted Full text extracted Articles in summary tables 4,334 RCT 56 53 53 31 3,717 Observational 97 21 21 21 RCT, randomized controlled trial. Table 11 Classification of study quality Good quality Low risk of bias and no obvious reporting errors, complete reporting of data. Must be prospective. If study of intervention, must be randomized controlled study (RCT). Fair quality Moderate risk of bias, but problems with study/paper are unlikely to cause major bias. If study of intervention, must be prospective. Poor quality High risk of bias or cannot exclude possible significant biases. Poor methods, incomplete data, reporting errors. Prospective or retrospective. Table 12 GRADE system for grading quality of evidence Step 1: Starting grade for quality of evidence based on study design Step 2: Reduce grade Step 3: Raise grade Final grade for quality of evidence and definition Randomized trials = High Observational study = Low

Table 11 Classification of study quality Good quality Low risk of bias and no obvious reporting errors, complete reporting of data. Must be prospective. If study of intervention, must be randomized controlled study (RCT). Fair quality Moderate risk of bias, but problems with study/paper are unlikely to cause major bias. If study of intervention, must be prospective. Poor quality High risk of bias or cannot exclude possible significant biases. Poor methods, incomplete data, reporting errors. Prospective or retrospective. Table 12 GRADE system for grading quality of evidence Step 1: Starting grade for quality of evidence based on study design Step 2: Reduce grade Step 3: Raise grade Final grade for quality of evidence and definition Randomized trials = High Observational study = Low Any other evidence = Very low Study quality −1 level if serious limitations −2 levels if very serious limitations Consistency −1 level if important inconsistency Directness −1 level if some uncertainty −2 levels if major uncertainty Other −1 level if sparse or imprecise datac −1 level if high probability of reporting bias Strength of association +1 level is stronga, no plausible confounders +2 levels if very strongb, no major threats to validity Other +1 level if evidence of a dose-response gradient +1 level if all residual plausible confounders would have reduced the observed effect High = Further research is unlikely to change confidence in the estimate of the effect Moderate = Further research is likely to have an important impact on confidence in the estimate of effect, and may change the estimate Low = Further research is very likely to have an important impact on confidence in the estimate, and may change the estimate Very low = Any estimate of effect is very uncertain GRADE, Grading of Recommendations Assessment, Development, and Evaluation.

on confidence in the estimate of effect, and may change the estimate Low = Further research is very likely to have an important impact on confidence in the estimate, and may change the estimate Very low = Any estimate of effect is very uncertain GRADE, Grading of Recommendations Assessment, Development, and Evaluation. a Strong evidence of association is defined as ‘significant relative risk of >2 (<0.5)' based on consistent evidence from two or more observational studies, with no plausible confounders. b Very strong evidence of association is defined as ‘significant relative risk of >5 (<0.2)' based on direct evidence with no major threats to validity. c Sparse if there is only one study or if total N <100. Imprecise if there is a low event rate (0 or 1 event) in either arm or confidence interval spanning a range <0.5 to >2.0. Adapted by permission from Macmillan Publishers Ltd: Kidney International. Uhlig K, Macleod A, Craig J et al. Grading evidence and recommendations for clinical practice guidelines in nephrology. A position statement from Kidney Disease: Improving Global Outcomes (KDIGO). Kidney Int 2006; 70: 2058–2065;243 accessed http://www.nature.com/ki/journal/v70/n12/pdf/5001875a.pdf

Kidney International. Uhlig K, Macleod A, Craig J et al. Grading evidence and recommendations for clinical practice guidelines in nephrology. A position statement from Kidney Disease: Improving Global Outcomes (KDIGO). Kidney Int 2006; 70: 2058–2065;243 accessed http://www.nature.com/ki/journal/v70/n12/pdf/5001875a.pdf Table 13 Final grade for overall quality of evidence Grade Quality of evidence Meaning A High We are confident that the true effect lies close to that of the estimate of the effect. B Moderate The true effect is likely to be close to the estimate of the effect, but there is a possibility that it is substantially different. C Low The true effect may be substantially different from the estimate of the effect. D Very low The estimate of effect is very uncertain, and often will be far from the truth. Table 14 Balance of benefits and harm When there was evidence to determine the balance of medical benefits and harm of an intervention to a patient, conclusions were categorized as follows: • For statistically significant benefit/harm report as ‘Benefit/Harm of Drug X'. • For non-statistically significant benefit/harm, report as ‘Possible benefit/harm of Drug X'. • In instances where studies are inconsistent, report as ‘Possible benefit/harm of Drug X'. • ‘No difference' can only be reported if a study is not imprecise. • ‘Insufficient evidence' if imprecision is a factor. Table 15 KDIGO nomenclature and description for grading recommendations Grade* Implications Patients Clinicians Policy Level 1 ‘We recommend' Most people in your situation would want the recommended course of action and only a small proportion would not. Most patients should receive the recommended course of action. The recommendation can be evaluated as a candidate for developing a policy or a performance measure. Level 2 ‘We suggest' The majority of people in your situation would want the recommended course of action, but many would not. Different choices will be appropriate for different patients. Each patient needs help to arrive at a management decision consistent with her or his values and preferences. The recommendation is likely to require substantial debate and involvement of stakeholders before policy can be determined. *The additional category ‘Not Graded' was used, typically, to provide guidance based on common sense or where the topic does not allow adequate application of evidence.

his values and preferences. The recommendation is likely to require substantial debate and involvement of stakeholders before policy can be determined. *The additional category ‘Not Graded' was used, typically, to provide guidance based on common sense or where the topic does not allow adequate application of evidence. The most common examples include recommendations regarding monitoring intervals, counseling, and referral to other clinical specialists. The ungraded recommendations are generally written as simple declarative statements, but are not meant to be interpreted as being stronger recommendations than Level 1 or 2 recommendations.

st common examples include recommendations regarding monitoring intervals, counseling, and referral to other clinical specialists. The ungraded recommendations are generally written as simple declarative statements, but are not meant to be interpreted as being stronger recommendations than Level 1 or 2 recommendations. Table 16 Determinants of strength of recommendation Factor Comment Balance between desirable and undesirable effects The larger the difference between the desirable and undesirable effects, the more likely a strong recommendation is warranted. The narrower the gradient, the more likely a weak recommendation is warranted. Quality of the evidence The higher the quality of evidence, the more likely a strong recommendation is warranted. Values and preferences The more variability in values and preferences, or more uncertainty in values and preferences, the more likely a weak recommendation is warranted. Costs (resource allocation) The higher the costs of an intervention—that is, the more resources consumed—the less likely a strong recommendation is warranted. Table 17 The Conference on Guideline Standardization (COGS) checklist245 for reporting clinical practice guidelines Topic Description Discussed in KDIGO Anemia Guideline 1. Overview material Provide a structured abstract that includes the guideline's release date, status (original, revised, updated), and print and electronic sources. Abstract and Methods for Guideline Development. 2. Focus Describe the primary disease/condition and intervention/service/technology that the guideline addresses. Indicate any alternative preventative, diagnostic or therapeutic interventions that were considered during development. Management of adults and children with CKD and kidney transplant recipients at risk for or with anemia. 3. Goal Describe the goal that following the guideline is expected to achieve, including the rationale for development of a guideline on this topic. This clinical practice guideline is intended to assist the practitioner caring for patients with CKD and anemia and to prevent deaths, cardiovascular disease events and progression to kidney failure while optimizing patients' quality of life. 4. User/setting Describe the intended users of the guideline (e.g. provider types, patients) and the settings in which the guideline is intended to be used. Providers: Nephrologists (adult and pediatric), Dialysis providers (including nurses), Internists, and Pediatricians.

e while optimizing patients' quality of life. 4. User/setting Describe the intended users of the guideline (e.g. provider types, patients) and the settings in which the guideline is intended to be used. Providers: Nephrologists (adult and pediatric), Dialysis providers (including nurses), Internists, and Pediatricians. Patients: Adult and children with CKD at risk for or with anemia. Policy Makers: Those in related health fields. 5. Target population Describe the patient population eligible for guideline recommendations and list any exclusion criteria. CKD individuals at risk for or with anemia, adult and children. 6. Developer Identify the organization(s) responsible for guideline development and the names/credentials/potential conflicts of interest of individuals involved in the guideline's development. Organization: KDIGO. 7. Funding source/sponsor Identify the funding source/sponsor and describe its role in developing and/or reporting the guideline. Disclose potential conflict of interest. KDIGO is supported by the following consortium of sponsors: Abbott, Amgen, Bayer Schering Pharma, Belo Foundation, Bristol-Myers Squibb, Chugai Pharmaceutical, Coca-Cola Company, Dole Food Company, Fresenius Medical Care, Genzyme, Hoffmann-LaRoche, JC Penney, Kyowa Hakko Kirin, NATCO—The Organization for Transplant Professionals, NKF-Board of Directors, Novartis, Pharmacosmos, PUMC Pharmaceutical, Robert and Jane Cizik Foundation, Shire, Takeda Pharmaceutical, Transwestern Commercial Services, Vifor Pharma, and Wyeth. No funding is accepted for the development or reporting of specific guidelines. All stakeholders could participate in open review. Refer to Work Group Financial Disclosures. 8. Evidence collection Describe the methods used to search the scientific literature, including the range of dates and databases searched, and criteria applied to filter the retrieved evidence. Modules were created for randomized controlled trials (RCTs), kidney disease, anemia, and erythropoietin, transfusion, iron deficiency, and adjuvant search terms. The search terms were then limited to years 2006–2010 for studies related to anemia interventions. For transfusion the literature search was conducted from 1989–2010. A separate search was run for observational studies on iron overload and hemoglobin status as predictors for clinical outcomes. See Table 8 for screening criteria.

terms were then limited to years 2006–2010 for studies related to anemia interventions. For transfusion the literature search was conducted from 1989–2010. A separate search was run for observational studies on iron overload and hemoglobin status as predictors for clinical outcomes. See Table 8 for screening criteria. Searches were run in MEDLINE, Cochrane Central Register of Controlled Clinical Trials and Cochrane Database of Systematic Reviews. The initial search for RCTs was conducted in April 2010 and subsequently updated in October of 2010. The search for observational studies was later conducted in September 2010. The search yield was also supplemented by articles provided by Work Group members through March 2012. 9. Recommendation grading criteria Describe the criteria used to rate the quality of evidence that supports the recommendations and the system for describing the strength of the recommendations. Recommendation strength communicates the importance of adherence to a recommendation and is based on both the quality of the evidence and the magnitude of anticipated benefits and harms. Quality of individual studies was graded in a three-tiered grading system (see Table 11). Quality of evidence (Table 12) was graded following the GRADE approach. Strength of the recommendation was graded in a two-level grading system which was adapted from GRADE for KDIGO with the quality of overall evidence graded on a four-tiered system (Tables 13 and 15). The Work Group could provide general guidance in ungraded statements. 10. Method for synthesizing evidence Describe how evidence was used to create recommendations, e.g., evidence tables, meta-analysis, decision analysis. For systematic review topics, summary tables and evidence profiles were generated. For recommendations on treatment interventions, the steps outlined by GRADE were followed. 11. Prerelease review Describe how the guideline developer reviewed and/or tested the guidelines prior to release. The guideline has undergone internal review by the KDIGO Board of Directors in June 2011 and external review in September 2011. Public review comments were compiled and fed back to the Work Group, which considered comments in its revision of the guideline. 12. Update plan State whether or not there is a plan to update the guideline and, if applicable, expiration date for this version of the guideline. There is no date set for updating.

er 2011. Public review comments were compiled and fed back to the Work Group, which considered comments in its revision of the guideline. 12. Update plan State whether or not there is a plan to update the guideline and, if applicable, expiration date for this version of the guideline. There is no date set for updating. The need for updating of the guideline will depend on the publication of new evidence that would change the quality of the evidence or the estimates for the benefits and harms. Results from registered ongoing studies and other publications will be reviewed periodically to evaluate their potential to impact on the recommendations in this guideline. 13. Definitions Define unfamiliar terms and those critical to correct application of the guideline that might be subject to misinterpretation. Abbreviations and Acronyms. 14. Recommendations and rationale State the recommended action precisely and the specific circumstances under which to perform it. Justify each recommendation by describing the linkage between the recommendation and its supporting evidence. Indicate the quality of evidence and the recommendation strength, based on the criteria described in Topic 9. Each guideline chapter contains recommendations for management of CKD patients at risk for or with anemia. Each recommendation builds on a supporting rationale with evidence tables if available. The strength of the recommendation and the quality of evidence are provided in parenthesis within each recommendation. 15. Potential benefits and harm Describe anticipated benefits and potential risks associated with implementation of guideline recommendations. The benefits and harm for each comparison of interventions are provided in summary tables and summarized in evidence profiles. The estimated balance between potential benefits and harm was considered when formulating the recommendations. 16. Patient preferences Describe the role of patient preferences when a recommendation involves a substantial element of personal choice or values. Many recommendations are Level 2 or “discretionary” which indicates a greater need to help each patient arrive at a management decision consistent with her or his values and preferences. 17. Algorithm Provide (when appropriate) a graphical description of the stages and decisions in clinical care described by the guideline. See Chapter 4. 18. Implementation considerations Describe anticipated barriers to application of the recommendations.

ment decision consistent with her or his values and preferences. 17. Algorithm Provide (when appropriate) a graphical description of the stages and decisions in clinical care described by the guideline. See Chapter 4. 18. Implementation considerations Describe anticipated barriers to application of the recommendations. Provide reference to any auxiliary documents for providers or patients that are intended to facilitate implementation. Suggest review criteria for measuring changes in care when the guideline is implemented. These recommendations are global. Review criteria were not suggested because implementation with prioritization and development of review criteria have to proceed locally. Furthermore, most recommendations are discretionary, requiring substantial discussion among stakeholders before they can be adopted as review criteria. Suggestions were provided for future research. CKD, chronic kidney disease; GRADE, Grading of Recommendations Assessment, Development, and Evaluation; KDIGO, Kidney Disease: Improving Global Outcomes; RCT, randomized controlled trial.

John J V McMurray, MD, FRCP, FESC (Work Group Co-Chair), is Professor of Medical Cardiology at BHF Glasgow Cardiovascular Research Centre and Head of Section of Academic Cardiology at University of Glasgow. Dr McMurray received his medical degree from University of Manchester and completed additional clinical training in Edinburgh, Dundee and Glasgow. He conducts clinical research in a wide span of areas including heart failure, left ventricular dysfunction, coronary heart disease, diabetes, and kidney failure. As such, Dr McMurray is a member of the Executive Committee or Steering Committee for a number of large ongoing multinational trials: ARISTOTLE, ASCEND-HF, ASPIRE, ATMOSPHERE, Dal-OUTCOMES, EMPHASIS-HF, NAVIGATOR, PARADIGM-HF, RED-HF, TREAT and VIVIDD. He is also Past President of the Heart Failure Association of the European Society of Cardiology and has authored close to 500 original publications, reviews, and book chapters. Dr McMurray is currently on a number of journal editorial boards including: Cardiovascular Drugs and Therapy, Circulation: Heart Failure, European Heart Journal, European Journal of Heart Failure, Heart, Heart Failure Reviews, International Journal of Cardiology and Journal of the Renin-Angiotensin-Alderosterone System. Dr McMurray′s employer, Glasgow University, received support from Amgen for his role as Executive Committee member of clinical trials (RED-HF; TREAT; ATOMIC-HF). Dr McMurray′s salary from his employer is independent from the monies received by Glasgow University from commercial or non-commercial organizations

John J V McMurray, MD, FRCP, FESC (Work Group Co-Chair), is Professor of Medical Cardiology at BHF Glasgow Cardiovascular Research Centre and Head of Section of Academic Cardiology at University of Glasgow. Dr McMurray received his medical degree from University of Manchester and completed additional clinical training in Edinburgh, Dundee and Glasgow. He conducts clinical research in a wide span of areas including heart failure, left ventricular dysfunction, coronary heart disease, diabetes, and kidney failure. As such, Dr McMurray is a member of the Executive Committee or Steering Committee for a number of large ongoing multinational trials: ARISTOTLE, ASCEND-HF, ASPIRE, ATMOSPHERE, Dal-OUTCOMES, EMPHASIS-HF, NAVIGATOR, PARADIGM-HF, RED-HF, TREAT and VIVIDD. He is also Past President of the Heart Failure Association of the European Society of Cardiology and has authored close to 500 original publications, reviews, and book chapters. Dr McMurray is currently on a number of journal editorial boards including: Cardiovascular Drugs and Therapy, Circulation: Heart Failure, European Heart Journal, European Journal of Heart Failure, Heart, Heart Failure Reviews, International Journal of Cardiology and Journal of the Renin-Angiotensin-Alderosterone System. Dr McMurray′s employer, Glasgow University, received support from Amgen for his role as Executive Committee member of clinical trials (RED-HF; TREAT; ATOMIC-HF). Dr McMurray′s salary from his employer is independent from the monies received by Glasgow University from commercial or non-commercial organizations Patrick S Parfrey, MD, FRCPC, FRSC (Work Group Co-Chair), is a University Research Professor at Memorial University and staff nephrologist at Eastern Health, Newfoundland. Dr Parfrey received his medical degree from University College Cork, Ireland and is active in clinical epidemiology research in kidney disease, particularly as it relates to cardiovascular disease, anemia and genetic diseases. He also supervised post-graduate work of more than 50 students and has authored over 300 publications. Dr Parfrey is past Associate Editor of CJASN, past president of the Canadian Society of Nephrology, an Officer of the Order of Canada and Fellow of the Royal Society of Canada and has received financial support from both Amgen and Ortho Biotech, as Chair of the Data Monitoring Committee of the Normal Hematocrit Study; Co-Primary Investigator of the Canada–Europe Trial; Executive Committee Member of TREAT; and Co-Chair of the Executive Committee of EVOLVE.

ellow of the Royal Society of Canada and has received financial support from both Amgen and Ortho Biotech, as Chair of the Data Monitoring Committee of the Normal Hematocrit Study; Co-Primary Investigator of the Canada–Europe Trial; Executive Committee Member of TREAT; and Co-Chair of the Executive Committee of EVOLVE. Grant/Research Support: Amgen Speaker: Amgen

ellow of the Royal Society of Canada and has received financial support from both Amgen and Ortho Biotech, as Chair of the Data Monitoring Committee of the Normal Hematocrit Study; Co-Primary Investigator of the Canada–Europe Trial; Executive Committee Member of TREAT; and Co-Chair of the Executive Committee of EVOLVE. Grant/Research Support: Amgen Speaker: Amgen John W Adamson, MD, completed his undergraduate work at the University of California, Berkeley, and received a MD degree from UCLA. Following training in Internal Medicine and Hematology at the University of Washington, he spent two years at the NIH, returning to the faculty in Seattle in 1969. He rose through the ranks to become professor and head of the Division of Hematology in 1980 and was named a Clinical Research Professor of the American Cancer Society in 1988. In 1989, he moved to New York City as President of the New York Blood Center and director of its research institute. In 1998 he moved to Milwaukee as Executive Vice President for Research at the Blood Center of Wisconsin and Director of its Blood Research Institute. Four years ago he joined the faculty at the University of California, San Diego, as Clinical Professor of Medicine in Hematology/Oncology where he serves as head of the Hematology/Oncology section at the VA Medical Center and Associate Director of the Hematology/Oncology Fellowship program at UCSD. Dr Adamson has published numerous scientific articles and reviews and has previously served as Editor-in-Chief of Blood; founding editor of Current Opinion in Hematology; President of the American Society of Hematology; and President of the International Society for Experimental Hematology. His interests lie in the areas of anemia diagnosis and management, pathophysiology of the myeloproliferative neoplasms, and the molecular biology of iron metabolism.

of Current Opinion in Hematology; President of the American Society of Hematology; and President of the International Society for Experimental Hematology. His interests lie in the areas of anemia diagnosis and management, pathophysiology of the myeloproliferative neoplasms, and the molecular biology of iron metabolism. Advisor/Consultant: Affymax; Akebia; AMAG; Amgen; Hospira; Watson Speaker: AMAG; Watson Pedro Aljama, MD, PhD, received his MD from the University of Cadiz in 1971 and PhD from the University of Seville in 1975. Professor Aljama then continued his training at the Royal Victoria lnfirmary, Newcastle, United Kingdom, where he was a Medical Officier, Registrar and then Senior Registrar in Nephrology, and a Lecturer in Renal Medicine at the University of Newcastle upon Tyne (1977–1979). He returned to Spain in 1980 as a Senior Registrar at Reina Sofia Hospital, University of Cordoba, and was appointed Professor of Medicine and Nephrology in 1987. He is past President of the Spanish Society of Nephrology and presently a member of the International Society of Nephrology, the European Society for Clinical Investigation, the British Society of Nephrology and the European Renal Association-European Dialysis and Transplant Association. Professor Aljama has authored over 250 scientific papers and 50 book chapters. Advisor/Consultant: Amgen; Roche; Vifor Grant/Research Support: Janssen-Cilag; Roche Speaker: Amgen; Vifor

Pedro Aljama, MD, PhD, received his MD from the University of Cadiz in 1971 and PhD from the University of Seville in 1975. Professor Aljama then continued his training at the Royal Victoria lnfirmary, Newcastle, United Kingdom, where he was a Medical Officier, Registrar and then Senior Registrar in Nephrology, and a Lecturer in Renal Medicine at the University of Newcastle upon Tyne (1977–1979). He returned to Spain in 1980 as a Senior Registrar at Reina Sofia Hospital, University of Cordoba, and was appointed Professor of Medicine and Nephrology in 1987. He is past President of the Spanish Society of Nephrology and presently a member of the International Society of Nephrology, the European Society for Clinical Investigation, the British Society of Nephrology and the European Renal Association-European Dialysis and Transplant Association. Professor Aljama has authored over 250 scientific papers and 50 book chapters. Advisor/Consultant: Amgen; Roche; Vifor Grant/Research Support: Janssen-Cilag; Roche Speaker: Amgen; Vifor Jeffrey S Berns, MD, is Professor of Medicine at the Perelman School of Medicine at the University of Pennsylvania and the Penn Presbyterian Medical Center of Philadelphia, University of Pennsylvania Health System. Dr Berns is also the Associate Dean for Graduate Medical Education, Nephrology Fellowship Program Director and Associate Chief of Renal, Electrolyte and Hypertension Division at the University of Pennsylvania Health System. He obtained his medical degree from Case Western Reserve University and completed his nephrology fellowship at Yale University School of Medicine. His professional activities include his service as a long-standing Work Group member of the KDOQI Anemia guideline from 1995–2007 and currently he is the KDOQI Vice Chair for Guideline Commentaries and Updates and also a member of the National Quality Forum ESRD Steering Committee. Dr Berns has authored over 130 publications and is on the editorial board of Clinical Nephrology, CJASN, and Seminars in Dialysis. In recognition for his contributions, he received the Leonard Berwick Memorial Teaching Award in 2008 and the Penn Medicine Patient Advocacy Award in 2010.

ty Forum ESRD Steering Committee. Dr Berns has authored over 130 publications and is on the editorial board of Clinical Nephrology, CJASN, and Seminars in Dialysis. In recognition for his contributions, he received the Leonard Berwick Memorial Teaching Award in 2008 and the Penn Medicine Patient Advocacy Award in 2010. Advisor/Consultant: Affymax; Amgen; Takeda Julia Bohlius, MD, MScPH, is a physician who is trained in both hematology/oncology and public health. Dr Bohlius is Editor of the Cochrane Haematological Malignancies Group and has experience in the conduct of both literature-based and individual patient data meta-analyses. Since 2001 she is a leading systematic reviewer on ESAs in cancer patients and has worked on international health technology assessments and clinical guideline projects for ESAs and other growth factors in cancer patients. While she started her clinical and scientific career at the University of Cologne, Germany, she now works as a Senior Research Fellow at the Institute of Social and Preventive Medicine, University of Bern, Switzerland. Dr Bohlius reported no relevant financial relationships

Julia Bohlius, MD, MScPH, is a physician who is trained in both hematology/oncology and public health. Dr Bohlius is Editor of the Cochrane Haematological Malignancies Group and has experience in the conduct of both literature-based and individual patient data meta-analyses. Since 2001 she is a leading systematic reviewer on ESAs in cancer patients and has worked on international health technology assessments and clinical guideline projects for ESAs and other growth factors in cancer patients. While she started her clinical and scientific career at the University of Cologne, Germany, she now works as a Senior Research Fellow at the Institute of Social and Preventive Medicine, University of Bern, Switzerland. Dr Bohlius reported no relevant financial relationships Tilman B Drüeke, MD, FRCP, is Emeritus Director of Research at the INSERM laboratory ERI-12, UFR de Médecine et Pharmacie, Université de Picardie Jules Verne, Amiens, France. He received his MD degree at the University of Tübingen Medical School, Germany in 1968. From 1969 through 2009, he practiced his medical and scientific activities at Necker Hospital/Necker Medical School, Université Paris V, Paris, France. Professor Drüeke's research interests focus on chronic renal failure, hemodialysis, metabolic and endocrine abnormalities, anemia, cardiovascular complications and arterial hypertension. He is a member of several scientific societies, committees and advisory boards and a former Co-Chair of the KDIGO CKD-MBD Guideline Work Group. Professor Drüeke is Editor Emeritus of Nephrology Dialysis Transplantation, former Associate Editor of the CJASN, an editorial board member of JASN and presently Associate Editor of Kidney International. He has published more than 500 original articles and reviews in peer-reviewed journals.

KD-MBD Guideline Work Group. Professor Drüeke is Editor Emeritus of Nephrology Dialysis Transplantation, former Associate Editor of the CJASN, an editorial board member of JASN and presently Associate Editor of Kidney International. He has published more than 500 original articles and reviews in peer-reviewed journals. Advisor/Consultant: Amgen; Roche; Vifor Speaker: Amgen; Chugai; Vifor Fredric O Finkelstein, MD, obtained his medical degree from Columbia University and completed his nephrology fellowship at Yale University Medical School where he is also presently a Clinical Professor of Medicine. Over the span of his career, he has lectured extensively throughout the world and has held more than 30 visiting teaching positions. In addition, he is currently Chair of the International Liaison Committee of the International Society of Peritoneal Dialysis. He is also Co-Chair of the Dialysis Committee of the International Society of Nephrology and an author of over 200 publications. Dr Finkelstein has dedicated substantial research towards the understanding of quality of life and psychosocial issues for dialysis and non-dialysis patients alike. He has served on the editorial board of Peritoneal Dialysis International since 2004 and Kidney International since 2010. Advisor/Consultant: Akebia, Amgen; Baxter Grant/Research Support: Amgen

Fredric O Finkelstein, MD, obtained his medical degree from Columbia University and completed his nephrology fellowship at Yale University Medical School where he is also presently a Clinical Professor of Medicine. Over the span of his career, he has lectured extensively throughout the world and has held more than 30 visiting teaching positions. In addition, he is currently Chair of the International Liaison Committee of the International Society of Peritoneal Dialysis. He is also Co-Chair of the Dialysis Committee of the International Society of Nephrology and an author of over 200 publications. Dr Finkelstein has dedicated substantial research towards the understanding of quality of life and psychosocial issues for dialysis and non-dialysis patients alike. He has served on the editorial board of Peritoneal Dialysis International since 2004 and Kidney International since 2010. Advisor/Consultant: Akebia, Amgen; Baxter Grant/Research Support: Amgen Steven Fishbane, MD, received his medical degree from Albert Einstein College of Medicine where he also completed his nephrology fellowship. He is currently Vice President of the North Shore-LIJ Health System in Manhasset, NY, as well as Professor of Medicine at SUNY Stony Brook School of Medicine. Dr Fishbane is the Director of Clinical Trials for the Department of Medicine of North Shore-LIJ University Hospitals. Having participated as a KDOQI Anemia Guideline Work Group member, he maintains an active research interest in this area and has written over 150 publications. In addition to serving as a reviewer for numerous journals, he currently sits on the editorial board of CJASN and Kidney International. In recognition for his commitment on enhancing healthcare delivery and assessment, Dr Fishbane was the recipient of the Physician Leadership in Quality Improvement Award from IPRO in 2002 and the Volunteerism Award of the National Kidney Foundation Serving Greater New York in 2010.

rd of CJASN and Kidney International. In recognition for his commitment on enhancing healthcare delivery and assessment, Dr Fishbane was the recipient of the Physician Leadership in Quality Improvement Award from IPRO in 2002 and the Volunteerism Award of the National Kidney Foundation Serving Greater New York in 2010. Advisor/Consultant: Affymax; Akebia; Fibrogen; Rockwell Medical Technologies Tomas Ganz, PhD, MD, is Professor of Medicine and Pathology at the David Geffen School of Medicine at UCLA. Dr Ganz received his PhD from the California Institute of Technology in Applied Physics and MD from UCLA. He was then trained in Internal Medicine and Pulmonary/Critical Care Medicine at the UCLA Medical Center. His major focus was on research on the biological role of peptide mediators in innate immunity and iron metabolism. More recently, he has investigated the pathogenesis of anemia of inflammation and iron overload states, and worked on the development of hepcidin agonists and antagonists. Dr Ganz has served as an Associate Editor of Blood and a member of the Erythrocyte and Leukocyte Biology (ELB) Study Section of the National Institutes of Health. In 2005, he received the Marcel Simon Award of the International Bioiron Society for the discovery of hepcidin. Advisor/Consultant: Alnylam; Intrinsic LifeSciences; Merganser Biotech; Ortho Biotech/Centocor; Pieris; Xenon; Employee: Intrinsic LifeSciences; Merganser Biotech Equity Interest: Intrinsic LifeSciences; Merganser Biotech Grant/Research Support: Amgen; Xenon

Tomas Ganz, PhD, MD, is Professor of Medicine and Pathology at the David Geffen School of Medicine at UCLA. Dr Ganz received his PhD from the California Institute of Technology in Applied Physics and MD from UCLA. He was then trained in Internal Medicine and Pulmonary/Critical Care Medicine at the UCLA Medical Center. His major focus was on research on the biological role of peptide mediators in innate immunity and iron metabolism. More recently, he has investigated the pathogenesis of anemia of inflammation and iron overload states, and worked on the development of hepcidin agonists and antagonists. Dr Ganz has served as an Associate Editor of Blood and a member of the Erythrocyte and Leukocyte Biology (ELB) Study Section of the National Institutes of Health. In 2005, he received the Marcel Simon Award of the International Bioiron Society for the discovery of hepcidin. Advisor/Consultant: Alnylam; Intrinsic LifeSciences; Merganser Biotech; Ortho Biotech/Centocor; Pieris; Xenon; Employee: Intrinsic LifeSciences; Merganser Biotech Equity Interest: Intrinsic LifeSciences; Merganser Biotech Grant/Research Support: Amgen; Xenon Iain C Macdougall, BSc, MD, FRCP, is a Consultant Nephrologist and Professor of Clinical Nephrology at King's College Hospital, London, UK. He is a combined medical and science graduate of Glasgow University, Scotland, from which he was awarded a First Class Honours BSc in Pharmacology in 1980, and his medical degree in 1983. Professor Macdougall then completed his general medical and nephrology training at hospitals in Glasgow, Cardiff, and London. He developed a research interest in renal anemia while a Clinical Research Fellow in Cardiff (1988–1991) and extended this interest during his appointment at St Bartholomew's Hospital (1991–1996), where he studied the potential role of proinflammatory cytokines in mediating resistance to epoetin. He has been involved in numerous advisory boards in renal anemia management worldwide, including the Working Parties responsible for both the 1999 and the 2004 versions of the European Best Practice Guidelines, along with the Work Group that produced the latest US KDOQI Anemia Guidelines (2006; update 2007). He was a previous Board member of the KDIGO initiative, and a Council member of the European Renal Association from 2004 until 2007. He has been the UK lead on several pivotal clinical trials of anemia management in patients with chronic kidney disease, including CREATE and TREAT, and he chairs the Anaemia Clinical Study Group of the UK Kidney Research Consortium.

itiative, and a Council member of the European Renal Association from 2004 until 2007. He has been the UK lead on several pivotal clinical trials of anemia management in patients with chronic kidney disease, including CREATE and TREAT, and he chairs the Anaemia Clinical Study Group of the UK Kidney Research Consortium. Advisor/Consultant: Affymax; Amgen; Ortho Biotech; Roche; Takeda; Vifor Grant/Research Support: Affymax; Amgen; Vifor Speaker: Amgen; Ortho Biotech; Takeda; Vifor

itiative, and a Council member of the European Renal Association from 2004 until 2007. He has been the UK lead on several pivotal clinical trials of anemia management in patients with chronic kidney disease, including CREATE and TREAT, and he chairs the Anaemia Clinical Study Group of the UK Kidney Research Consortium. Advisor/Consultant: Affymax; Amgen; Ortho Biotech; Roche; Takeda; Vifor Grant/Research Support: Affymax; Amgen; Vifor Speaker: Amgen; Ortho Biotech; Takeda; Vifor Ruth A McDonald, MD, is Professor of Pediatrics at University of Washington and Clinical Director of Nephrology at Children's Hospital and Regional Medical Center in Seattle, Washington. She completed her medical degree at University of Minneosta School of Medicine where she was a recipient of the Top Medical Graduate: Hewlett-Packard Award. Dr McDonald is currently involved in numerous multicenter clinical studies including a controlled trial of Anti-CD20 monoclonal antibody therapy in historically unsensitized renal transplant recipients with donor-specific antibodies; a Phase II study to determine safety and immunomodulatory functions of induction therapy with Campath 1 H, combined with mycophenolate mofetil and sirolimus; a surveillance study of viral infections in renal transplant recipients and many others. She is also a member of eight professional organizations including American Society of Pediatric Nephrology, American Society of Transplantation, International Pediatric Transplant Association and past Work Group member of the KDIGO Clinical Practice Guideline for the Care of Kidney Transplant Recipients. Among her teaching responsibilities, she has trained over 25 fellows and has also served as Medical Student Research Mentor. Dr McDonald has authored over 60 publications and has given close to 40 invited and extra-institutional lectures in the past 10 years.

tice Guideline for the Care of Kidney Transplant Recipients. Among her teaching responsibilities, she has trained over 25 fellows and has also served as Medical Student Research Mentor. Dr McDonald has authored over 60 publications and has given close to 40 invited and extra-institutional lectures in the past 10 years. Dr McDonald reported no relevant financial relationships Lawrence P McMahon, MBBS, MD, is Director, Department of Renal Medicine at Eastern Health Integrated Renal Services and Professor Nephrology at Monash University. Prior to his present appointments, he was Associate Professor at University of Melbourne School of Medicine; Director of Nephrology Services and Obstetric Medical Services at Western Health; and Consortium Director of Physician Training at Greater Western Consortium. Dr McMahon has participated in guideline development activities for the Australian and New Zealand Society of Nephrology and is presently the President, National Council of Society of Obstetric Medicine of Australian and New Zealand. He has written more than 50 publications and serves as a regular reviewer for more than a dozen journals, including his role as Associate Editor of Nephrology Dialysis Transplantation. Grant/Research Support: Amgen; Roche

Lawrence P McMahon, MBBS, MD, is Director, Department of Renal Medicine at Eastern Health Integrated Renal Services and Professor Nephrology at Monash University. Prior to his present appointments, he was Associate Professor at University of Melbourne School of Medicine; Director of Nephrology Services and Obstetric Medical Services at Western Health; and Consortium Director of Physician Training at Greater Western Consortium. Dr McMahon has participated in guideline development activities for the Australian and New Zealand Society of Nephrology and is presently the President, National Council of Society of Obstetric Medicine of Australian and New Zealand. He has written more than 50 publications and serves as a regular reviewer for more than a dozen journals, including his role as Associate Editor of Nephrology Dialysis Transplantation. Grant/Research Support: Amgen; Roche Gregorio T Obrador, MD, MPH, is Professor of Medicine and Dean at the Universidad Panamericana School of Medicine in Mexico City. He also serves as Adjunct Physician at the Tufts Medical Center's Division of Nephrology and as staff nephrologist at Dalinde Medical Center in Mexico City. He earned his medical degree from the University of Navarra, Pamplona, Spain, completed his Internal Medicine residency at the Western Pennsylvania Hospital, Pittsburgh, USA, and obtained his Nephrology training at Boston University, USA. While undertaking a clinical research fellowship at the Tufts-New England Medical Center and a Master of Public Health at Harvard University, he demonstrated that the management of patients with CKD prior to stage 5 is suboptimal, and that this is an important factor for the high morbidity and mortality observed in these patients. He has been a member of the KDOQI's Advisory Board, the NKF/KDOQI Anemia Work Goup, and the KDIGO Transplant Guideline Work Group. Currently he is a member of the WHO's Non-Communicable diseases Network (NCDnet), Co-Chair of the Global Kidney Disease Prevention Network (KDPN), Co-Chair of the Latin American Clinical Practice Guidelines for the Prevention, Diagnosis and Treatment of CKD (Stages 1–5), and President of the Board of Directors of the Mexican Kidney Foundation. In 2009 he received the National Kidney Foundation's International Distinguished Medal. Dr Obrador is a member of the editorial board of CJASN and has served as reviewer for other nephrology journals. He has given more than 100 lectures in national and international forums and has several publications in the area of CKD.

2009 he received the National Kidney Foundation's International Distinguished Medal. Dr Obrador is a member of the editorial board of CJASN and has served as reviewer for other nephrology journals. He has given more than 100 lectures in national and international forums and has several publications in the area of CKD. Grant/Research Support: Amgen; Roche Giovanni FM Strippoli, MD, PhD, MPH, is a nephrologist and an epidemiologist trained both in Italy and at the University of Sydney School of Public Health, Sydney, Australia where he completed a Master of Public Health and a PhD in medicine-clinical epidemiology. Dr Strippoli is an editor of the Cochrane Renal Group, and Adjunct Associate Professor of Epidemiology at the School of Public Health, and the Renal Research Coordinator at Mario Negri Sud Consortium in Italy. He also serves as scientific director of Diaverum AB. His research interests include evidence-based nephrology, with a focus on systematic reviews in the area of prognosis and treatment of renal conditions, design and conduct of randomized controlled trials in the field of prevention of chronic kidney disease and cardiovascular risk. Dr Strippoli has a substantial scientific output with independent funding in these areas. He is also the principal investigator of LIRICO, a trial on the Long Term Impact of Renin Angiotensin System Inhibitors on Cardiorenal Outcomes in people with albuminuria, and C.E. DOSE, a trial on the clinical evaluation of the Dose of Erythropoietins in people on hemodialysis. Employee: Diaverum AB

Giovanni FM Strippoli, MD, PhD, MPH, is a nephrologist and an epidemiologist trained both in Italy and at the University of Sydney School of Public Health, Sydney, Australia where he completed a Master of Public Health and a PhD in medicine-clinical epidemiology. Dr Strippoli is an editor of the Cochrane Renal Group, and Adjunct Associate Professor of Epidemiology at the School of Public Health, and the Renal Research Coordinator at Mario Negri Sud Consortium in Italy. He also serves as scientific director of Diaverum AB. His research interests include evidence-based nephrology, with a focus on systematic reviews in the area of prognosis and treatment of renal conditions, design and conduct of randomized controlled trials in the field of prevention of chronic kidney disease and cardiovascular risk. Dr Strippoli has a substantial scientific output with independent funding in these areas. He is also the principal investigator of LIRICO, a trial on the Long Term Impact of Renin Angiotensin System Inhibitors on Cardiorenal Outcomes in people with albuminuria, and C.E. DOSE, a trial on the clinical evaluation of the Dose of Erythropoietins in people on hemodialysis. Employee: Diaverum AB Günter Weiss, MD, is Professor of Clinical Immunology and Infectious Diseases, Department of Internal Medicine, and Head of research laboratory for Molecular Immunology and Infectious Diseases at Medical University of Innsbruck. Dr Weiss had enrolled in Leopold Franzens University and University of Innsbruck for his medical studies and his ongoing research encompasses a wide array of topics including: anemia of chronic disease; primary and secondary iron overload; host pathogen interaction with a particular focus on the role of macrophages and natural resistance genes; and regulatory interactions between iron, immunity and infection. Dr Weiss has authored 190 original publications in peer reviewed journals including reviews on anemia of chronic disease and iron metabolism in inflammation and infection.

n with a particular focus on the role of macrophages and natural resistance genes; and regulatory interactions between iron, immunity and infection. Dr Weiss has authored 190 original publications in peer reviewed journals including reviews on anemia of chronic disease and iron metabolism in inflammation and infection. Grant/Research Support: Amgen Speaker: Vifor Andrzej Wie?cek, MD, PhD, FRCP, is Professor of Internal Medicine and Chief, Department of Nephrology, Endocrinology and Metabolic Diseases, at Silesian University School of Medicine, Katowice, Poland. Dr Wie?cek's research interests include anemia management in CKD, treatments for primary and secondary hypertension, elucidation of hormonal abnormalities in uremia, and endocrine function of adipose tissue. In addition to being a participating member of the European Renal Best Practice Anaemia Working Group, he is Past President of Polish Society of Nephrology and has served on the KDIGO Board. Dr Wie?cek is now a member of KDIGO Implementation Task Force Leader for Eastern Europe region and Secretary-Treasurer for the ERA-EDTA. As a prolific author with over 530 publications, he is currently Subject Editor for Nephrology Dialysis Transplantation. Advisor/Consultant: Abbott; Affymax; Sandoz Speaker: Amgen KDIGO CHAIRS Bertram L Kasiske, MD, is Professor of Medicine at the University of Minnesota, USA. He received his medical degree from the University of Iowa and completed his Internal Medicine residency and fellowship training in Nephrology at Hennepin County Medical Center where he is currently Director of Nephrology.

KDIGO CHAIRS Bertram L Kasiske, MD, is Professor of Medicine at the University of Minnesota, USA. He received his medical degree from the University of Iowa and completed his Internal Medicine residency and fellowship training in Nephrology at Hennepin County Medical Center where he is currently Director of Nephrology. Dr Kasiske is former Deputy Director of the United States Renal Data System and former Editor-in-Chief of The American Journal of Kidney Diseases. He has served as Secretary/Treasurer and on the Board of Directors of the American Society of Transplantation, and on the Organ Procurement and Transplantation Network/United Network of Organ Sharing Board of Directors, and the Scientific Advisory Board of the National Kidney Foundation. He is currently serving on the Board of Councilors of the International Society of Nephrology. He is the Principal Investigator for a National Institutes of Health-sponsored, multi-center study of long term outcomes after kidney donation. He is the Director of the Scientific Registry of Transplant Recipients. He has over 160 scientific publications in major peer reviewed journals, and 230 review articles, editorials and textbook chapters. Dr Kasiske is also a recipient of the NKF's Garabed Eknoyan Award in 2003. Advisor/Consultant: Litholink Grant/Research Support: Bristol-Myers Squibb; Merck-Schering Plough

Dr Kasiske is former Deputy Director of the United States Renal Data System and former Editor-in-Chief of The American Journal of Kidney Diseases. He has served as Secretary/Treasurer and on the Board of Directors of the American Society of Transplantation, and on the Organ Procurement and Transplantation Network/United Network of Organ Sharing Board of Directors, and the Scientific Advisory Board of the National Kidney Foundation. He is currently serving on the Board of Councilors of the International Society of Nephrology. He is the Principal Investigator for a National Institutes of Health-sponsored, multi-center study of long term outcomes after kidney donation. He is the Director of the Scientific Registry of Transplant Recipients. He has over 160 scientific publications in major peer reviewed journals, and 230 review articles, editorials and textbook chapters. Dr Kasiske is also a recipient of the NKF's Garabed Eknoyan Award in 2003. Advisor/Consultant: Litholink Grant/Research Support: Bristol-Myers Squibb; Merck-Schering Plough David C Wheeler, MD, FRCP, holds an academic position in Nephrology (Reader) at University College London, UK and is an Honorary Consultant Nephrologist at the Royal Free Hospital. His research is focused on the cardiovascular complications of chronic kidney disease and the role of vascular risk factors in progression of kidney damage. Dr Wheeler is a member of the International Steering Committee of the Study of Heart and Renal Protection (SHARP) and was UK National Coordinator for the trial. He is involved in several other randomized trials and observational studies involving patients with chronic kidney disease.

rs in progression of kidney damage. Dr Wheeler is a member of the International Steering Committee of the Study of Heart and Renal Protection (SHARP) and was UK National Coordinator for the trial. He is involved in several other randomized trials and observational studies involving patients with chronic kidney disease. He currently serves on the executive committee of KDIGO and previously contributed as a Work Group member to the KDIGO Guideline on Chronic Kidney Disease-Mineral and Bone Disorder. He has recently received an International Distinguished Medal from the US National Kidney Foundation in recognition of his contribution to guideline development. In the UK, he has previously served on the executive committee of the Renal Association and has been elected President for the term 2012–2014. Dr Wheeler has served on the editorial boards of the American Journal of Kidney Diseases and Journal of the American Society of Nephrology and currently acts as co-Deputy Editor for Nephrology Dialysis Transplantation. Advisor/Consultant: Amgen Honoraria: Abbott, Amgen, Fresenius, Shire Grant/Research Support: Abbott, Genzyme

He currently serves on the executive committee of KDIGO and previously contributed as a Work Group member to the KDIGO Guideline on Chronic Kidney Disease-Mineral and Bone Disorder. He has recently received an International Distinguished Medal from the US National Kidney Foundation in recognition of his contribution to guideline development. In the UK, he has previously served on the executive committee of the Renal Association and has been elected President for the term 2012–2014. Dr Wheeler has served on the editorial boards of the American Journal of Kidney Diseases and Journal of the American Society of Nephrology and currently acts as co-Deputy Editor for Nephrology Dialysis Transplantation. Advisor/Consultant: Amgen Honoraria: Abbott, Amgen, Fresenius, Shire Grant/Research Support: Abbott, Genzyme EVIDENCE REVIEW TEAM Ethan M Balk, MD, MPH, is Director, Evidence-based Medicine at the Tufts Center for Kidney Disease Guideline Development and Implementation, in Boston, MA, USA, Associate Director of the Tufts Evidence-based Practice Center, and Assistant Professor of Medicine at Tufts University School of Medicine. Dr Balk graduated from Tufts University School of Medicine and completed a fellowship in Clinical Care Research. As Project Director, he plays a substantial role in providing methodological expertise in the guideline development process and assists in the collection, evaluation, grading, and synthesis of evidence and the revisions of the final evidence report. Dr Balk also provides methodological guidance and training of Work Group members during meetings regarding topic refinement, key question formulation, data extraction, study assessment, evidence grading, and recommendation formulation. His primary research interests are evidence-based medicine, systematic review, clinical practice guideline development, and critical literature appraisal.

up members during meetings regarding topic refinement, key question formulation, data extraction, study assessment, evidence grading, and recommendation formulation. His primary research interests are evidence-based medicine, systematic review, clinical practice guideline development, and critical literature appraisal. Dr Balk reported no relevant financial relationships Ashish Upadhyay, MD, is Assistant Professor, Renal Section and Associate Director, Internal Medicine Residency Program at Boston University School of Medicine, Boston, MA, USA. Dr Upadhyay was previously Assistant Professor at Tufts University School of Medicine and staff physician in the William B. Schwartz, MD, Division of Nephrology at Tufts Medical Center. He joined the ERT in July 2009 and served as the Assistant Project Director for the KDIGO Management of Blood Pressure in CKD and Anemia in CKD Guidelines. Dr Upadhyay coordinated and assisted in the collection, evaluation, grading, and synthesis of evidence, and played a critical role in the revisions of the final evidence report. He also provided methodological guidance and training of Work Group members on topic refinement, key question formulation, data extraction, study assessment, evidence grading, and recommendation formulation. Dr Upadhyay's past research involved studying kidney disease epidemiology in the Framingham Heart Study. He has published in areas ranging from arterial stiffness in CKD and inflammation in kidney disease to dialysis complications and epidemiology of hyponatremia. Dr Upadhyay reported no relevant financial relationships

Ashish Upadhyay, MD, is Assistant Professor, Renal Section and Associate Director, Internal Medicine Residency Program at Boston University School of Medicine, Boston, MA, USA. Dr Upadhyay was previously Assistant Professor at Tufts University School of Medicine and staff physician in the William B. Schwartz, MD, Division of Nephrology at Tufts Medical Center. He joined the ERT in July 2009 and served as the Assistant Project Director for the KDIGO Management of Blood Pressure in CKD and Anemia in CKD Guidelines. Dr Upadhyay coordinated and assisted in the collection, evaluation, grading, and synthesis of evidence, and played a critical role in the revisions of the final evidence report. He also provided methodological guidance and training of Work Group members on topic refinement, key question formulation, data extraction, study assessment, evidence grading, and recommendation formulation. Dr Upadhyay's past research involved studying kidney disease epidemiology in the Framingham Heart Study. He has published in areas ranging from arterial stiffness in CKD and inflammation in kidney disease to dialysis complications and epidemiology of hyponatremia. Dr Upadhyay reported no relevant financial relationships Dana C Miskulin, MD, MS, is Assistant Professor of Medicine at Tufts University School of Medicine, Boston, MA, USA. She completed a fellowship in Clinical Care Research and participated in the conduct of systematic reviews and critical literature appraisals for this guideline. Her primary research interests are in comparative effectiveness research in dialysis patients, blood pressure treatment in dialysis patients, and autosomal dominant polycystic kidney disease.

nical Care Research and participated in the conduct of systematic reviews and critical literature appraisals for this guideline. Her primary research interests are in comparative effectiveness research in dialysis patients, blood pressure treatment in dialysis patients, and autosomal dominant polycystic kidney disease. Dr Miskulin reported no relevant financial relationships Amy Earley, BS, is a project coordinator at the Tufts Center for Kidney Disease Guideline Development and Implementation in Boston, MA, USA. She is key in coordinating the guideline development activities within the ERT, especially in the development of the evidence reports for all guidelines. Ms Earley also heads the actual evidence review, which includes running searches, screening, data extraction, drafting of tables and methods sections, proofing of guideline drafts and critical literature appraisals. She participates in the conduct of research projects at the Center and actively collaborates with other members of the Center on independent research topics and manuscript submissions. Ms Earley reported no relevant financial relationships Shana Haynes, MS, DHSc, is a research assistant at the Tufts Center for Kidney Disease Guideline Development and Implementation in Boston, MA, USA. She participates in all aspects of evidence review and guideline development. She screens abstracts and articles, extracts data, and assists in the drafting and editing of evidence tables. Dr Haynes also assists in the development of clinical practice guidelines and conducts systematic reviews and critical literature appraisals.

ipates in all aspects of evidence review and guideline development. She screens abstracts and articles, extracts data, and assists in the drafting and editing of evidence tables. Dr Haynes also assists in the development of clinical practice guidelines and conducts systematic reviews and critical literature appraisals. Dr Haynes reported no relevant financial relationships Jenny Lamont, MS, is a project manager and medical writer at the Tufts Center for Kidney Disease Guideline Development and Implementation in Boston, MA, USA. She participates in all aspects of evidence review and guideline development, assists in the preparation of talks and manuscripts, and edits KDIGO draft guidelines currently in progress. Ms Lamont reported no relevant financial relationships

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SECTION I: USE OF THE CLINICAL PRACTICE GUIDELINE This Clinical Practice Guideline document is based upon systematic literature searches last conducted in January 2011, supplemented with additional evidence through February 2012. It is designed to provide information and assist decision making. It is not intended to define a standard of care, and should not be construed as one, nor should it be interpreted as prescribing an exclusive course of management. Variations in practice will inevitably and appropriately occur when clinicians take into account the needs of individual patients, available resources, and limitations unique to an institution or type of practice. Every health-care professional making use of these recommendations is responsible for evaluating the appropriateness of applying them in any particular clinical situation. The recommendations for research contained within this document are general and do not imply a specific protocol.

n institution or type of practice. Every health-care professional making use of these recommendations is responsible for evaluating the appropriateness of applying them in any particular clinical situation. The recommendations for research contained within this document are general and do not imply a specific protocol. SECTION II: DISCLOSURE Kidney Disease: Improving Global Outcomes (KDIGO) makes every effort to avoid any actual or reasonably perceived conflicts of interest that may arise as a result of an outside relationship or a personal, professional, or business interest of a member of the Work Group. All members of the Work Group are required to complete, sign, and submit a disclosure and attestation form showing all such relationships that might be perceived or actual conflicts of interest. This document is updated annually and information is adjusted accordingly. All reported information is published in its entirety at the end of this document in the Work Group members' Biographic and Disclosure Information section, and is kept on file at the National Kidney Foundation (NKF), Managing Agent for KDIGO.

It is our hope that this document will serve several useful purposes. Our primary goal is to improve patient care. We hope to accomplish this, in the short term, by helping clinicians know and better understand the evidence (or lack of evidence) that determines current practice. By providing comprehensive evidence-based recommendations, this guideline will also help define areas where evidence is lacking and research is needed. Helping to define a research agenda is an often neglected, but very important, function of clinical practice guideline development. We used the Grading of Recommendations Assessment, Development and Evaluation (GRADE) system to rate the quality of evidence and the strength of recommendations. In all, there were no recommendations in this guideline for which the overall quality of evidence was graded ‘A,' whereas 4 (23.5%) were graded ‘B,' 3 (17.7%) were graded ‘C,' and 10 (58.8%) were graded ‘D.' Although there are reasons other than quality of evidence that underpin a grade 1 or 2 recommendation, in general, there is a correlation between the quality of overall evidence and the strength of the recommendation. Thus, there were 8 (47.1%) recommendations graded ‘1' and 9 (52.9%) graded ‘2.' There were no recommendations graded ‘1A,' 4 (23.5%) were ‘1B,' 2 (11.8%) were ‘1C,' and 2 (11.8%) were ‘1D.' There were no recommendations graded ‘2A' or ‘2B,' 1 (5.9%) was ‘2C,' and 8 (47.1%) were ‘2D.' There were 4 (19.1%) statements that were not graded.

(47.1%) recommendations graded ‘1' and 9 (52.9%) graded ‘2.' There were no recommendations graded ‘1A,' 4 (23.5%) were ‘1B,' 2 (11.8%) were ‘1C,' and 2 (11.8%) were ‘1D.' There were no recommendations graded ‘2A' or ‘2B,' 1 (5.9%) was ‘2C,' and 8 (47.1%) were ‘2D.' There were 4 (19.1%) statements that were not graded. Some argue that recommendations should not be made when evidence is weak. However, clinicians still need to make decisions in their daily practice, and they often ask, ‘What do the experts do in this setting?' We opted to give guidance, rather than remain silent. These recommendations are often rated with a low strength of recommendation and a low quality of evidence, or were not graded. It is important for the users of this guideline to be cognizant of this (see Notice). In every case these recommendations are meant to be a place for clinicians to start, not stop, their inquiries into specific management questions pertinent to the patients they see in daily practice. We wish to thank Dr Gavin Becker who co-chaired the Work Group with David Wheeler, along with all of the Work Group members who volunteered countless hours of their time developing this guideline. We also thank the Evidence Review Team members and staff of the National Kidney Foundation who made this project possible. Finally, we owe a special debt of gratitude to the many KDIGO Board members and individuals who volunteered time reviewing the guideline, and making very helpful suggestions. Bertram L Kasiske, MD KDIGO Co-Chair David C Wheeler, MD, FRCP KDIGO Co-Chair

Work Group Co-Chairs Gavin J Becker, MD, FRACP Royal Melbourne Hospital Melbourne, Australia David C Wheeler, MD, FRCP University College London London, United Kingdom Work Group Dick de Zeeuw, MD, PhD University Medical Center Groningen Groningen, Netherlands Toshiro Fujita, MD University of Tokyo School of Medicine Tokyo, Japan Susan L Furth, MD, PhD The Children's Hospital of Philadelphia Philadelphia, PA, USA Hallvard Holdaas, MD, PhD Hospital Rikshospitalet Oslo, Norway Shanthi Mendis, MBBS, MD, FRCP, FACC World Health Organization Geneva, Switzerland Suzanne Oparil, MD University of Alabama Birmingham, AL, USA Vlado Perkovic, MBBS, FRACP, FASN, PhD George Institute for International Health Sydney, Australia Cibele Isaac Saad Rodrigues, MD, PhD Catholic University of São Paulo São Paulo, Brazil Mark J Sarnak, MD, MS Tufts Medical Center Boston, MA, USA Guntram Schernthaner, MD Rudolfstiftung Hospital Vienna, Austria Charles R V Tomson, DM, FRCP Southmead Hospital Bristol, United Kingdom Carmine Zoccali, MD CNR-IBIM Clinical Research Unit, Ospedali Riuniti Reggio Calabria, Italy Evidence Review Team Tufts Center for Kidney Disease Guideline Development and Implementation, Tufts Medical Center, Boston, MA, USA: Katrin Uhlig, MD, MS, Project Director; Director, Guideline Development Ashish Upadhyay, MD, Assistant Project Director Amy Earley, BS, Project Coordinator Shana Haynes, MS, DHSc, Research Assistant Jenny Lamont, MS, Project Manager In addition, support and supervision were provided by: Ethan M Balk, MD, MPH; Program Director, Evidence Based Medicine

The 2012 Kidney Disease: Improving Global Outcomes (KDIGO) Clinical Practice Guideline for the Management of Blood Pressure in Chronic Kidney Disease aims to provide guidance on blood pressure management and treatment for all non-dialysis-dependent CKD patients and kidney transplant recipients. Guideline development followed an explicit process of evidence review and appraisal. Treatment approaches are addressed in each chapter and guideline recommendations are based on systematic reviews of relevant trials. Appraisal of the quality of the evidence and the strength of recommendations followed the GRADE approach. Ongoing areas of controversies and limitations of the evidence are discussed and additional suggestions are also provided for future research. Keywords: blood pressure; chronic kidney disease; clinical practice guideline; evidence-based recommendation; KDIGO; systematic review

Chapter 2: Lifestyle and pharmacological treatments for lowering blood pressure in CKD ND patients GENERAL STRATEGIES 2.1: Individualize BP targets and agents according to age, co-existent cardiovascular disease and other co-morbidities, risk of progression of CKD, presence or absence of retinopathy (in CKD patients with diabetes) and tolerance of treatment. (Not Graded) 2.2: Inquire about postural dizziness and check for postural hypotension regularly when treating CKD patients with BP-lowering drugs. (Not Graded) LIFESTYLE MODIFICATION 2.3: Encourage lifestyle modification in patients with CKD to lower BP and improve long-term cardiovascular and other outcomes: 2.3.1: We recommend achieving or maintaining a healthy weight (BMI 20 to 25). (1D) 2.3.2: We recommend lowering salt intake to <90 mmol (<2 g) per day of sodium (corresponding to 5 g of sodium chloride), unless contraindicated. (1C) 2.3.3: We recommend undertaking an exercise program compatible with cardiovascular health and tolerance, aiming for at least 30 minutes 5 times per week. (1D) 2.3.4: We suggest limiting alcohol intake to no more than two standard drinks per day for men and no more than one standard drink per day for women. (2D) Chapter 3: Blood pressure management in CKD ND patients without diabetes mellitus 3.1: We recommend that non-diabetic adults with CKD ND and urine albumin excretion <30 mg per 24 hours (or equivalent*) whose office BP is consistently >140 mm Hg systolic or >90 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤140 mm Hg systolic and ≤90 mm Hg diastolic. (1B)

tus 3.1: We recommend that non-diabetic adults with CKD ND and urine albumin excretion <30 mg per 24 hours (or equivalent*) whose office BP is consistently >140 mm Hg systolic or >90 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤140 mm Hg systolic and ≤90 mm Hg diastolic. (1B) 3.2: We suggest that non-diabetic adults with CKD ND and urine albumin excretion of 30 to 300 mg per 24 hours (or equivalent*) whose office BP is consistently >130 mm Hg systolic or >80 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤130 mm Hg systolic and ≤80 mm Hg diastolic. (2D) 3.3: We suggest that non-diabetic adults with CKD ND and urine albumin excretion >300 mg per 24 hours (or equivalent*) whose office BP is consistently >130 mm Hg systolic or >80 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤130 mm Hg systolic and ≤80 mm Hg diastolic. (2C) 3.4: We suggest that an ARB or ACE-I be used in non-diabetic adults with CKD ND and urine albumin excretion of 30 to 300 mg per 24 hours (or equivalent*) in whom treatment with BP-lowering drugs is indicated. (2D) 3.5: We recommend that an ARB or ACE-I be used in non-diabetic adults with CKD ND and urine albumin excretion >300 mg per 24 hours (or equivalent*) in whom treatment with BP-lowering drugs is indicated. (1B) *Approximate equivalents for albumin excretion rate per 24 hours—expressed as protein excretion rate per 24 hours, albumin/creatinine ratio, protein/creatinine ratio, and protein reagent strip results—are given in Table 1, Chapter 1.

3.5: We recommend that an ARB or ACE-I be used in non-diabetic adults with CKD ND and urine albumin excretion >300 mg per 24 hours (or equivalent*) in whom treatment with BP-lowering drugs is indicated. (1B) *Approximate equivalents for albumin excretion rate per 24 hours—expressed as protein excretion rate per 24 hours, albumin/creatinine ratio, protein/creatinine ratio, and protein reagent strip results—are given in Table 1, Chapter 1. Chapter 4: Blood pressure management in CKD ND patients with diabetes mellitus 4.1: We recommend that adults with diabetes and CKD ND with urine albumin excretion <30 mg per 24 hours (or equivalent*) whose office BP is consistently >140 mm Hg systolic or >90 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤140 mm Hg systolic and ≤90 mm Hg diastolic. (1B) 4.2: We suggest that adults with diabetes and CKD ND with urine albumin excretion >30 mg per 24 hours (or equivalent*) whose office BP is consistently >130 mm Hg systolic or >80 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤130 mm Hg systolic and ≤80 mm Hg diastolic. (2D) 4.3: We suggest that an ARB or ACE-I be used in adults with diabetes and CKD ND with urine albumin excretion of 30 to 300 mg per 24 hours (or equivalent*). (2D) 4.4: We recommend that an ARB or ACE-I be used in adults with diabetes and CKD ND with urine albumin excretion >300 mg per 24 hours (or equivalent*). (1B)

4.2: We suggest that adults with diabetes and CKD ND with urine albumin excretion >30 mg per 24 hours (or equivalent*) whose office BP is consistently >130 mm Hg systolic or >80 mm Hg diastolic be treated with BP-lowering drugs to maintain a BP that is consistently ≤130 mm Hg systolic and ≤80 mm Hg diastolic. (2D) 4.3: We suggest that an ARB or ACE-I be used in adults with diabetes and CKD ND with urine albumin excretion of 30 to 300 mg per 24 hours (or equivalent*). (2D) 4.4: We recommend that an ARB or ACE-I be used in adults with diabetes and CKD ND with urine albumin excretion >300 mg per 24 hours (or equivalent*). (1B) *Approximate equivalents for albumin excretion rate per 24 hours—expressed as protein excretion rate per 24 hours, albumin/creatinine ratio, protein/creatinine ratio, and protein reagent strip results—are given in Table 1, Chapter 1. Chapter 5: Blood pressure management in kidney transplant recipients (CKD T) 5.1: We suggest that adult kidney transplant recipients whose office BP is consistently >130 mm Hg systolic or >80 mm Hg diastolic be treated to maintain a BP that is consistently ≤130 mm Hg systolic and ≤80 mm Hg diastolic, irrespective of the level of urine albumin excretion. (2D) 5.2: In adult kidney transplant recipients, choose a BP-lowering agent after taking into account the time after transplantation, use of calcineurin inhibitors, presence or absence of persistent albuminuria, and other co-morbid conditions. (Not Graded)

Chapter 5: Blood pressure management in kidney transplant recipients (CKD T) 5.1: We suggest that adult kidney transplant recipients whose office BP is consistently >130 mm Hg systolic or >80 mm Hg diastolic be treated to maintain a BP that is consistently ≤130 mm Hg systolic and ≤80 mm Hg diastolic, irrespective of the level of urine albumin excretion. (2D) 5.2: In adult kidney transplant recipients, choose a BP-lowering agent after taking into account the time after transplantation, use of calcineurin inhibitors, presence or absence of persistent albuminuria, and other co-morbid conditions. (Not Graded) Chapter 6: Blood pressure management in children with CKD ND 6.1: We recommend that in children with CKD ND, BP-lowering treatment is started when BP is consistently above the 90th percentile for age, sex, and height. (1C) 6.2: We suggest that in children with CKD ND (particularly those with proteinuria), BP is lowered to consistently achieve systolic and diastolic readings less than or equal to the 50th percentile for age, sex, and height, unless achieving these targets is limited by signs or symptoms of hypotension. (2D) 6.3: We suggest that an ARB or ACE-I be used in children with CKD ND in whom treatment with BP-lowering drugs is indicated, irrespective of the level of proteinuria. (2D)

6.2: We suggest that in children with CKD ND (particularly those with proteinuria), BP is lowered to consistently achieve systolic and diastolic readings less than or equal to the 50th percentile for age, sex, and height, unless achieving these targets is limited by signs or symptoms of hypotension. (2D) 6.3: We suggest that an ARB or ACE-I be used in children with CKD ND in whom treatment with BP-lowering drugs is indicated, irrespective of the level of proteinuria. (2D) Chapter 7: Blood pressure management in elderly persons with CKD ND 7.1: Tailor BP treatment regimens in elderly patients with CKD ND by carefully considering age, co-morbidities and other therapies, with gradual escalation of treatment and close attention to adverse events related to BP treatment, including electrolyte disorders, acute deterioration in kidney function, orthostatic hypotension and drug side effects. (Not Graded)

There is a strong association between chronic kidney disease (CKD) and an elevated blood pressure (BP) whereby each can cause or aggravate the other. BP control is fundamental to the care of patients with CKD and is relevant at all stages of CKD regardless of the underlying cause. Clinical practice guidelines (CPGs) have been published on this topic by many authoritative bodies over the past decade, the most comprehensive being the National Kidney Foundation's (NKF) Kidney Disease Outcomes Quality Initiative (KDOQI) Clinical Practice Guidelines on Hypertension and Antihypertensive Agents in Chronic Kidney Disease, which was based on evidence collected up to 2001 (http://www.kidney.org/professionals/KDOQI/guidelines_bp/index.htm).1 The Kidney Disease: Improving Global Outcomes (KDIGO) Board believed that it would be clinically useful to update this CPG to incorporate the evidence gathered since then. KDIGO therefore commissioned an evidence review to include the recent literature and assembled a Work Group with the mandate of writing an updated guideline relevant to an international audience. This KDIGO Guideline, entitled “Management of Blood Pressure in Chronic Kidney Disease,” is the result of these efforts. Scope of this guideline This Guideline has been developed to provide advice on the management of BP in patients with non–dialysis-dependent CKD (CKD ND) (see Reference Keys).

There is a strong association between chronic kidney disease (CKD) and an elevated blood pressure (BP) whereby each can cause or aggravate the other. BP control is fundamental to the care of patients with CKD and is relevant at all stages of CKD regardless of the underlying cause. Clinical practice guidelines (CPGs) have been published on this topic by many authoritative bodies over the past decade, the most comprehensive being the National Kidney Foundation's (NKF) Kidney Disease Outcomes Quality Initiative (KDOQI) Clinical Practice Guidelines on Hypertension and Antihypertensive Agents in Chronic Kidney Disease, which was based on evidence collected up to 2001 (http://www.kidney.org/professionals/KDOQI/guidelines_bp/index.htm).1 The Kidney Disease: Improving Global Outcomes (KDIGO) Board believed that it would be clinically useful to update this CPG to incorporate the evidence gathered since then. KDIGO therefore commissioned an evidence review to include the recent literature and assembled a Work Group with the mandate of writing an updated guideline relevant to an international audience. This KDIGO Guideline, entitled “Management of Blood Pressure in Chronic Kidney Disease,” is the result of these efforts. Scope of this guideline This Guideline has been developed to provide advice on the management of BP in patients with non–dialysis-dependent CKD (CKD ND) (see Reference Keys). BP We have avoided using the term ‘hypertension' in our title because this implies that there is a BP value above or below which morbidity or mortality changes in a stepwise fashion, hence suggesting that it is possible to set a universal BP target. In reality, it proved difficult to define precise targets appropriate for all CKD subpopulations, consistent with the notion that the ‘ideal' BP may differ between patients, once other factors are considered. These factors include specific features of CKD such as the severity of albuminuria or proteinuria, the presence of other risk factors for cardiovascular disease (CVD) and co-morbidities. Another reason for our choice of terminology is that agents introduced primarily to treat high BP may have actions that may not be directly linked to BP-lowering (e.g., the anti-albuminuric effects of angiotensin-converting enzyme inhibitors [ACE-Is] and angiotensin-receptor blockers [ARBs]).

(CVD) and co-morbidities. Another reason for our choice of terminology is that agents introduced primarily to treat high BP may have actions that may not be directly linked to BP-lowering (e.g., the anti-albuminuric effects of angiotensin-converting enzyme inhibitors [ACE-Is] and angiotensin-receptor blockers [ARBs]). Definition of CKD The Work Group defined CKD according to the standard KDOQI classification system2 as endorsed by KDIGO.3 Populations of interest The populations covered in this guideline are: Adults with CKD ND without diabetes mellitus Adults with CKD ND with diabetes mellitus Adults with CKD ND who have received a kidney transplant (CKD T) Children with CKD ND Elderly with CKD ND The scope of this guideline did not include BP management in patients with dialysis-dependent CKD 5 (CKD 5D) since this has been the topic of a recent KDIGO consensus conference4 and has been covered by two recent systematic reviews.5, 6 There are other groups of patients with CKD for whom specific recommendations might be welcome, but who are not represented in sufficient numbers in randomized controlled trials (RCTs) to constitute a sufficiently robust evidence base. The evidence review team (ERT) was asked to present the evidence separately for adults with CKD and diabetes, since these individuals constitute the single largest subgroup of CKD patients in the world.

ented in sufficient numbers in randomized controlled trials (RCTs) to constitute a sufficiently robust evidence base. The evidence review team (ERT) was asked to present the evidence separately for adults with CKD and diabetes, since these individuals constitute the single largest subgroup of CKD patients in the world. The separation of the evidence base according to diabetes status meant that there were two separate datasets for the Work Group to review. Although the two sets of recommendations had much in common, the Work Group decided that they differed sufficiently in detail to warrant two separate chapters. Adults who received a kidney transplant, children, and the elderly were also thought to deserve dedicated chapters, although the evidence base for each of these subpopulations is rather small. The Work Group was unable to identify sufficient evidence to make recommendations according to severity (stage) of CKD, although common sense dictates that pharmacological management should differ at least between mild CKD (patients with normal glomerular filtration rate [GFR]) and advanced CKD (patients with low GFR). However, the Work Group did consider the modification of drug dosages and risks related to the various classes of BP-lowering agents in the context of CKD in Chapter 2.

al management should differ at least between mild CKD (patients with normal glomerular filtration rate [GFR]) and advanced CKD (patients with low GFR). However, the Work Group did consider the modification of drug dosages and risks related to the various classes of BP-lowering agents in the context of CKD in Chapter 2. Clearly there are many other populations that could have been considered. CKD patients with glomerulonephritis are the subject of a recent KDIGO Guideline,7 so they were not considered separately here. Although management of BP in the pregnant CKD patient is an important issue, there is insufficient evidence in this subgroup to allow recommendations to be made.8 Furthermore, the Work Group did not consider the management of BP in patients with acute kidney injury. Interventions Interventions primarily aiming at modifying BP include advice on lifestyle and administration of pharmacological agents that reduce BP. The efficacies of both strategies have been widely studied in the general population with high BP. The pharmacology of anti-hypertensive agents was detailed in the 2004 KDOQI guideline.1 Of the available RCTs that met our inclusion criteria, most involved agents interfering with the renin-angiotensin-aldosterone system (RAAS). Accordingly, these agents may be over-represented in this Guideline, and if so, it is because of the availability of the evidence rather than a deliberate focus by the Work Group.

Of the available RCTs that met our inclusion criteria, most involved agents interfering with the renin-angiotensin-aldosterone system (RAAS). Accordingly, these agents may be over-represented in this Guideline, and if so, it is because of the availability of the evidence rather than a deliberate focus by the Work Group. Evidence for interventions Because CKD is common and BP levels are often elevated in CKD populations, the management of BP in CKD patients could have an enormous global impact. Given that the focus of the Guideline is on management and the comparative effectiveness of various interventions, the preferred and most robust evidence is derived from large-scale RCTs which assessed hard clinical outcomes. The ERT was asked to include RCTs with a minimum of 50 patients in each arm and interventions included pharmacological agents (alone or in combination), lifestyle modifications, and trials assessing various levels of BP control. Outcomes of interest were mortality, cardiovascular events and changes in kidney function including urine albumin or protein excretion.

imum of 50 patients in each arm and interventions included pharmacological agents (alone or in combination), lifestyle modifications, and trials assessing various levels of BP control. Outcomes of interest were mortality, cardiovascular events and changes in kidney function including urine albumin or protein excretion. Reduction in BP, particularly when achieved using agents that interfere with the RAAS, can lead to acute reductions in kidney function and albuminuria; thus the minimal duration of follow-up in RCTs required for their inclusion in the evidence review was set at 1 year for kidney function, cardiovascular outcomes, and mortality and 3 months for urine albumin or protein levels. Because there were so few trials assessing lifestyle modifications, BP reduction was included as an outcome, with the minimum follow-up period set at 6 weeks. The approach to the evidence review is described in detail in Methods for Guideline Development. The ERT conducted a systematic review of RCTs involving individuals with CKD. This was supplemented with published systematic reviews and meta-analyses (which often included smaller RCTs). Work Group members further supplemented this yield with selected RCTs that included individuals at increased risk of CVD but who were not specifically chosen on the basis of having CKD. The Work Group also helped identify RCTs that included CKD subgroups. To a lesser extent, the Work Group made reference to observational evidence from large population studies where evidence from RCTs was perceived to be insufficient.

ncreased risk of CVD but who were not specifically chosen on the basis of having CKD. The Work Group also helped identify RCTs that included CKD subgroups. To a lesser extent, the Work Group made reference to observational evidence from large population studies where evidence from RCTs was perceived to be insufficient. Not all questions of interest have been the subject of RCTs; some issues do not lend themselves to be studied in this manner. To facilitate further discussion on major issues relevant to management of BP in CKD patients (for which there is some evidence but ongoing controversy remains), the Work Group included a chapter on Future Directions and Controversies (Chapter 8). For other issues widely accepted in practice, but not supported by evidence from RCTs, the Work Group wrote ungraded recommendations reflecting the consensus of its members. These ungraded statements are explained in detail in the accompanying narrative. The Work Group did not wish to provide advice on specific treatment questions for which there was no supporting evidence. By highlighting these gaps in knowledge, we aim to promote further research. During the preparation of this Guideline, the Work Group was aware that other international organizations were writing new or updating old guidelines that were potentially relevant to the management of BP in CKD patients. The Work Group kept in contact with these other organizations and sought to achieve consistency with their recommendations as much as possible.

Work Group was aware that other international organizations were writing new or updating old guidelines that were potentially relevant to the management of BP in CKD patients. The Work Group kept in contact with these other organizations and sought to achieve consistency with their recommendations as much as possible. Measurement of BP The Work Group recognized that many reviews on the methodology of BP measurement have been published9, 10 and that this topic was covered in detail in the 2004 KDOQI Guideline.1 Previous publications have highlighted inconsistencies between conventional office (or clinic) BP measurements and other methods, such as self-measurement of BP at home or ambulatory blood pressure monitoring (ABPM).11, 12, 13 Many recommendations regarding when and how to use ABPM in hypertensive patients not known to have CKD have also been published. Although few studies have assessed the value of ABPM CKD patients, the small, short-term studies that do exist reflect the inconsistency between office BP measurements and other BP measurements and also suggest that ABPM gives a better indication of overall BP and kidney prognosis than office BP measurements.11, 12, 13 Despite this, to date there has only been one large RCT of BP control in CKD patients (all of whom were children) in which ABPM was used as the method for BP assessment.14 We therefore cannot provide evidence-based recommendations regarding the use of ABPM to evaluate BP in CKD patients but existing evidence is reviewed in Chapter 8.

o date there has only been one large RCT of BP control in CKD patients (all of whom were children) in which ABPM was used as the method for BP assessment.14 We therefore cannot provide evidence-based recommendations regarding the use of ABPM to evaluate BP in CKD patients but existing evidence is reviewed in Chapter 8. Since office BP measurements are used in almost all RCTs of interventions that modify BP in CKD, this Guideline can only make recommendations about BP assessed by this method. Because office readings are known to vary from day to day, management decisions should be based on repeated measurements,15 as emphasized in this guideline by the use of the term ‘consistently' (e.g., Recommendation 4.1 … maintain a BP that is consistently ≤140 mm Hg systolic …). The term is used simply to imply that the BP has been measured more than once and that there was meaningful agreement between the measurements. The Work Group also discussed whether to consider pulse pressure and/or pulse wave velocity, measures of arterial compliance that may provide important prognostic information in CKD patients. However, there is a paucity of data from RCTs showing that any particular intervention reliably alters these measures and subsequently influences mortality or morbidity. Thus the Work Group was not able to make any evidence-based recommendations relating to these measurements. However, these issues are of interest for the future of BP assessment in CKD patients and are discussed in further detail in Chapter 8.

ly alters these measures and subsequently influences mortality or morbidity. Thus the Work Group was not able to make any evidence-based recommendations relating to these measurements. However, these issues are of interest for the future of BP assessment in CKD patients and are discussed in further detail in Chapter 8. Albuminuria and proteinuria Some BP-lowering agents are particular effective at reducing albuminuria or proteinuria, suggesting that BP management should differ depending on the amount of albumin or protein in the urine.16, 17, 18, 19 Accordingly, as in the KDOQI 2004 Guidelines and the majority of other CPGs addressing BP control in patients with CKD or diabetes, the Work Group has attempted to stratify treatment effects according to urinary albumin excretion. Based on a recent KDIGO Controversies Conference and data from the CKD Prognosis Consortium, the Work Group used three categories (levels) of albuminuria.20 Wherever possible, the Work Group modified its recommendations to fit these categories, although since not all RCTs use this classification system, consistency was not achievable. The three categories of urinary albumin excretion are as follows: >300 mg per 24 h (or ‘macroalbuminuria'), 30 to 300 mg per 24 h (or ‘microalbuminuria'), and <30 mg per 24 h (Table 1). When other measures (such as assessment of proteinuria, ratios of urinary albumin or urinary protein to urine creatinine, or protein reagent strip readings) were used in RCTs, these measures were translated to albumin excretion rates (AERs) per 24 h, recognizing that these converted values are approximations at best. Recommendations and suggestions for interventions based on albumin levels expressed in milligrams per 24 h can also be converted (Table 1).

stification to recommend different BP levels for these two parameters. Doing so might also lead to confusion, since we would be recommending two different BP levels possibly with two evidence ratings and would not be able to provide coherent advice for managing patients between the recommended threshold and target BPs. Studies that have not specifically targeted CKD patients demonstrate that BP is a continuous risk factor for CVD outcomes.21 BP targets could differ depending on the presence of other CVD risk factors in each patient. This approach contrasts with the ‘one size fits all' philosophy that has previously been endorsed. There are far less data in CKD patients to inform the best approach. In RCTs involving CKD patients who are randomized to different BP targets, the achieved differences between groups are usually less than the targeted differences. Intention-to-treat analyses allow conclusions to be drawn based on target BP levels rather than achieved BP levels. The Work Group generally followed this convention and based recommendations on target levels BP levels rather than those achieved in the RCTs. It also considered the evidence derived from RCTs in which patients were not randomized to BP targets but achieved BPs were reported. The logic for using target BP levels in RCTs rather than the achieved BP levels observed as the basis for setting guideline targets has been questioned;22 this concern is one reason for our conservative approach to BP target setting in this Guideline.

ts were not randomized to BP targets but achieved BPs were reported. The logic for using target BP levels in RCTs rather than the achieved BP levels observed as the basis for setting guideline targets has been questioned;22 this concern is one reason for our conservative approach to BP target setting in this Guideline. Outcomes The major outcomes relevant to BP control in CKD patients are kidney disease progression and cardiovascular events (including stroke). Kidney outcomes Although it is possible for a diagnosis of CKD to be made in an individual with a normal GFR and AER and even a normal BP (for example on the basis of an imaging study, as in early adult polycystic kidney disease), most patients recruited into RCTs addressing BP and its management in CKD have a reduced GFR or persistently elevated albumin excretion. Entry criteria for RCTs involving CKD patients are usually based on these parameters, changes in which may form the basis for kidney end points.

arly adult polycystic kidney disease), most patients recruited into RCTs addressing BP and its management in CKD have a reduced GFR or persistently elevated albumin excretion. Entry criteria for RCTs involving CKD patients are usually based on these parameters, changes in which may form the basis for kidney end points. Kidney function Changes in kidney function are important outcomes in clinical trials assessing the effects of various BP-management regimens in CKD patients. Although the most important events are the requirement for renal replacement therapy or death due to kidney failure, many studies have used surrogates such as changes in GFR or the percentage of patients in whom the serum creatinine (SCr) level doubles. Such numerical end points may be particularly relevant in trials that include patients with early-stage CKD, among whom kidney failure and death are uncommon events. One problem with the assessment of such surrogates is that the therapeutic agent used to modify BP may also directly alter kidney function. For example, ACE-Is are known to reduce GFR through a vasodilator effect on the efferent arteriole. This effect may be beneficial in the early stages of CKD when a reduced intra-glomerular pressure is protective, but might be detrimental at a later stage when kidney function is severely compromised and dialysis may be imminent, at which time GFR may increase if ACE-Is are withdrawn.23 Thus, a drug may modify GFR via a mechanism that does not directly involve changes in systemic BP and the impact of this effect on the patient may vary according to CKD stage. The Work Group bore such considerations in mind when assessing the evidence and viewed consistency in the change of GFR outcomes across various CKD stages as a strong indicator of the benefits of a particular agent on kidney function.

emic BP and the impact of this effect on the patient may vary according to CKD stage. The Work Group bore such considerations in mind when assessing the evidence and viewed consistency in the change of GFR outcomes across various CKD stages as a strong indicator of the benefits of a particular agent on kidney function. Albuminuria The level of albuminuria in CKD predicts not only the prognosis with respect to kidney function but also morbidity and mortality from CVD events including stroke.16, 17, 18, 19 Urinary albumin excretion is influenced by BP and by many of the agents used to reduce BP, particularly ACE-Is and ARBs. The concept of using albuminuria as a surrogate marker for CKD progression and CVD outcomes is widely accepted, with the reduction of urine albumin levels often being regarded as a target for therapy. This would mean that treatment would be escalated to reduce albuminuria to a preferred level, regardless of BP. Treating to an albumin target usually involves an escalation of RAAS blockade, which can be achieved by restricting dietary salt intake, increasing doses of an ACE-I or an ARB, combining the two classes of medication, or by adding a thiazide diuretic, an aldosterone-receptor blocker or a direct renin inhibitor (DRI).

. Treating to an albumin target usually involves an escalation of RAAS blockade, which can be achieved by restricting dietary salt intake, increasing doses of an ACE-I or an ARB, combining the two classes of medication, or by adding a thiazide diuretic, an aldosterone-receptor blocker or a direct renin inhibitor (DRI). While a strong case has been made for targeting a reduction of albuminuria, particularly with agents that interfere with the RAAS, there have been no large studies in CKD patients reporting long term differences in GFR or CVD outcomes where reduction in urinary albumin levels (regardless of BP) was the primary objective. There is also uncertainty as to whether the dose of a particular agent that is required to achieve BP control is necessarily the same as the dose required for albuminuria reduction.24 The Work Group thus decided that it was premature to recommend an albuminuria reduction target strategy for all cases of CKD but felt this deserved further discussion in Chapter 8.

dose of a particular agent that is required to achieve BP control is necessarily the same as the dose required for albuminuria reduction.24 The Work Group thus decided that it was premature to recommend an albuminuria reduction target strategy for all cases of CKD but felt this deserved further discussion in Chapter 8. Cardiovascular outcomes Recognition that premature CVD is a major cause of death in CKD has led to CVD risk management becoming a recognized component of the care of the CKD patient. In planning appropriate interventions, one strategy is simply to extrapolate data from CVD outcomes trials in the general population. This approach has been challenged because the benefits of interventions predicted in observational studies25 are not always observed in RCTs involving CKD patients.26, 27 In CKD-ND patients,28 unlike CKD patients on dialysis (CKD 5D),29 a higher BP is generally associated with a higher CVD risk, making BP-lowering an attractive goal in an effort to reduce cardiovascular morbidity and mortality. Although no RCTs assessing BP lowering agents have been specifically designed or powered to assess cardiovascular event rates as the primary outcome in any group of CKD patients, several studies assessing cardiovascular outcomes have included CKD patients and this information was considered in making the recommendations.

Although no RCTs assessing BP lowering agents have been specifically designed or powered to assess cardiovascular event rates as the primary outcome in any group of CKD patients, several studies assessing cardiovascular outcomes have included CKD patients and this information was considered in making the recommendations. Intended Users of this Guideline This Guideline is primarily aimed at health care professionals caring for individuals with CKD, including nephrologists, nurses, and pharmacists, as well as at physicians involved in the care of patients with diabetes and primary care providers. The Guideline is not aimed at health care administrators, policy makers, or regulators, although the explanatory text might be of value to these groups and assist in enhancing implementation and adherence to BP-lowering strategies. The Guideline is also not designed to be used in the development of clinical performance measures. Some of the difficulties in implementation and in auditing BP target achievement are discussed in Chapter 8.

ight be of value to these groups and assist in enhancing implementation and adherence to BP-lowering strategies. The Guideline is also not designed to be used in the development of clinical performance measures. Some of the difficulties in implementation and in auditing BP target achievement are discussed in Chapter 8. DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. Table 1 Relationship among categories for albuminuria and proteinuriaa Categories Measure Normal to mildly increased Moderately increased Severely increased AER (mg/24 h) <30 30–300 >300 PER (mg/24 h) <150 150–500 >500 ACR (mg/mmol) <3 3–30 >30 (mg/g) <30 30–300 >300

DISCLAIMER While every effort is made by the publishers, editorial board, and ISN to see that no inaccurate or misleading data, opinion or statement appears in this Journal, they wish to make it clear that the data and opinions appearing in the articles and advertisements herein are the responsibility of the contributor, copyright holder, or advertiser concerned. Accordingly, the publishers and the ISN, the editorial board and their respective employers, office and agents accept no liability whatsoever for the consequences of any such inaccurate or misleading data, opinion or statement. While every effort is made to ensure that drug doses and other quantities are presented accurately, readers are advised that new methods and techniques involving drug usage, and described within this Journal, should only be followed in conjunction with the drug manufacturer's own published literature. Table 1 Relationship among categories for albuminuria and proteinuriaa Categories Measure Normal to mildly increased Moderately increased Severely increased AER (mg/24 h) <30 30–300 >300 PER (mg/24 h) <150 150–500 >500 ACR (mg/mmol) <3 3–30 >30 (mg/g) <30 30–300 >300 PCR (mg/mmol) <15 15–50 >50 (mg/g) <150 150–500 >500 Protein reagent strip Negative to trace Trace to + + or greater ACR, albumin/creatinine ratio; AER, albumin excretion rate; PCR, protein/creatinine ratio, PER, protein excretion rate. Albuminuria and proteinuria can be measured using excretion rates in timed urine collections, ratio of concentrations to creatinine concentration in spot urine samples, and using reagent strips in spot urine samples. Relationships among measurement methods within a category are not exact. The relationships between AER and ACR and between PER and PCR are based on the assumption that average creatinine excretion rate is approximately 1.0 g/24 h or 10 mmol/24 h. The conversions are rounded for pragmatic reasons. (For an exact conversion from mg/g of creatinine to mg/mmol of creatinine, multiply by 0.113.) Creatinine excretion varies with age, sex, race and diet; therefore the relationship among these categories is approximate only. ACR <10 mg/g (<1mg/mmol) is considered normal; ACR 10–29 mg/g (1.0–2.9mg/mmol) is considered ‘high normal.' The relationship between urine reagent strip results and other measures depends on urine concentration.

varies with age, sex, race and diet; therefore the relationship among these categories is approximate only. ACR <10 mg/g (<1mg/mmol) is considered normal; ACR 10–29 mg/g (1.0–2.9mg/mmol) is considered ‘high normal.' The relationship between urine reagent strip results and other measures depends on urine concentration. a Tentatively adopted by KDIGO CKD Work Group.

INTRODUCTION This section outlines lifestyle and pharmacological methods to reduce BP in patients with non-dialysis-dependent CKD (CKD ND). Because these strategies were covered in detail in the 2004 KDOQI Clinical Practice Guidelines on Hypertension and Antihypertensive Agents in Chronic Kidney Disease (http://www.kidney.org/professionals/KDOQI/guidelines_bp/index.htm),1 we concentrate on issues relating to BP control in CKD patients that have arisen since 2004. Additional information that may be of help to the clinician (although not specifically relevant to CKD patients) can be found in the Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (JNC 7) (http://www.nhlbi.nih.gov/guidelines/hypertension/jnc7full.pdf).9 GENERAL STRATEGIES It is generally accepted that a stepwise combination of lifestyle modifications and drug therapy should be used to lower BP in CKD patients, with escalation of efforts depending on factors such as the severity of the BP elevation, the co-morbidities present and the age of the patient.2.1: Individualize BP targets and agents according to age, co-existent cardiovascular disease and other co-morbidities, risk of progression of CKD, presence or absence of retinopathy (in CKD patients with diabetes) and tolerance of treatment. (Not Graded)

elevation, the co-morbidities present and the age of the patient.2.1: Individualize BP targets and agents according to age, co-existent cardiovascular disease and other co-morbidities, risk of progression of CKD, presence or absence of retinopathy (in CKD patients with diabetes) and tolerance of treatment. (Not Graded) RATIONALE We recognize that individual decision making is required regarding BP targets and agents with the risks and benefit being taken into consideration; however, since there is little evidence from RCTs to guide these decisions, this recommendation has not been graded. The potential benefits of lower BP include a decreased risk of both CVD and progression of CKD. To assess the likely benefit in a given patient, the clinician needs to consider such issues as the prior rate of CKD progression, the expected course of the specific disease, the level of urinary albumin excretion and the presence or absence of other risks of CVD. Potential adverse effects generic to treatment used to lower BP include decreases in cerebral perfusion (contributing to dizziness, confusion and falls) and acute deterioration in kidney function.

d course of the specific disease, the level of urinary albumin excretion and the presence or absence of other risks of CVD. Potential adverse effects generic to treatment used to lower BP include decreases in cerebral perfusion (contributing to dizziness, confusion and falls) and acute deterioration in kidney function. It is widely acknowledged that achievement of a reduction in BP can be difficult in CKD patients, particularly in the elderly, those with co-morbidities, and those with diabetes mellitus.1, 9, 30 Increased conduit-artery stiffness, resulting in high pulse pressure (with high systolic and low diastolic pressures) is common in CKD patients, the elderly and patients with diabetes.31, 32, 33, 34, 35, 36 Arterial stiffening is associated with an increased risk of CVD independent of other recognized risk factors.37, 38, 39 With a high pulse pressure, efforts to reduce systolic BP in older patients and those with coronary artery disease (CAD) can result in lowering diastolic BP to levels well below diastolic targets, which may be associated with greater morbidity or mortality.40, 41 A J-shaped relationship between achieved BP and outcome has been observed in the elderly and in patients with vascular disease, possibly suggesting that BP can be reduced too far in these patients.40, 42, 43 Discussion of this issue is further elaborated in Chapters 7 and 8. Unfortunately, in CKD patients, the available evidence proved to be insufficient to allow the Work Group to define the lowest BP targets (see Chapter 8).

vascular disease, possibly suggesting that BP can be reduced too far in these patients.40, 42, 43 Discussion of this issue is further elaborated in Chapters 7 and 8. Unfortunately, in CKD patients, the available evidence proved to be insufficient to allow the Work Group to define the lowest BP targets (see Chapter 8). Similarly, when considering the choice of BP-lowering agents, decision making should be tailored to the individual patient. For instance, ACE-Is and ARBs are potentially harmful in the presence of significant renovascular disease or volume depletion, or when used in combination with nonsteroidal anti-inflammatory drugs (NSAIDs) or cyclooxygenase-2 (COX-2) inhibitors (as outlined later in this chapter). The presence of diabetic retinopathy in a CKD patient may also influence BP target and choice of agent as outlined in Chapter 4. Based on these considerations, the Work Group concluded that it is good clinical practice to assess the risks and benefits of BP-lowering treatment in an individual patient and to tailor therapy accordingly.2.2: Inquire about postural dizziness and check for postural hypotension regularly when treating CKD patients with BP-lowering drugs. (Not Graded)

e Work Group concluded that it is good clinical practice to assess the risks and benefits of BP-lowering treatment in an individual patient and to tailor therapy accordingly.2.2: Inquire about postural dizziness and check for postural hypotension regularly when treating CKD patients with BP-lowering drugs. (Not Graded) RATIONALE Patients with CKD, particularly the elderly31 and diabetic patients with autonomic neuropathy, are prone to orthostatic hypotension,44, 45 which may be exacerbated by volume depletion. Many CKD patients will require combinations of drugs to control BP including vasodilators, which can cause or exacerbate postural hypotension. This can lead to postural dizziness, reduced adherence and in extreme cases, syncope or falls with consequent injury. Accordingly, it is sensible to regularly check for symptoms of postural dizziness and to compare lying, sitting and standing BP in CKD patients, particularly before and after altering the treatment regimen. LIFESTYLE MODIFICATION The impact of lifestyle-related factors on BP and the risk of cardiovascular and other diseases have been well documented. A number of observational studies in the general population have linked factors such as salt intake,46 weight and body mass index (BMI),47 exercise frequency,48 and alcohol intake49 with BP level. RCTs addressing many of these factors have been undertaken, the results of which have led the authors of BP guidelines for the general population9 (e.g., JNC 7) to make specific recommendations about the management of lifestyle as a key component of BP management.

ercise frequency,48 and alcohol intake49 with BP level. RCTs addressing many of these factors have been undertaken, the results of which have led the authors of BP guidelines for the general population9 (e.g., JNC 7) to make specific recommendations about the management of lifestyle as a key component of BP management. Individuals with CKD generally have higher9 BP levels than people with normal kidney function and their BP may be particularly sensitive to some factors related to lifestyle. For example, high salt intake may potentially have a greater impact on BP in patients with CKD than in those without CKD since CKD may reduce the ability to excrete the salt load in the urine. CKD patients may also be more sensitive to harms related to lifestyle interventions; for instance, an individual with tubular disease with salt wasting from the kidney could be at increased risk of hypovolemia if salt intake is restricted. Furthermore, some potential lifestyle interventions, such as increased physical exercise, may be difficult for patients with CKD owing to reduced energy levels.

ons; for instance, an individual with tubular disease with salt wasting from the kidney could be at increased risk of hypovolemia if salt intake is restricted. Furthermore, some potential lifestyle interventions, such as increased physical exercise, may be difficult for patients with CKD owing to reduced energy levels. Lifestyle modification offers the potential to lower BP in a simple, inexpensive, effective fashion while also improving a range of other outcomes (e.g., changes in lipid levels resulting from diet and exercise and liver function through moderation of alcohol intake). Because lifestyle changes are applicable to the general population and are potentially implementable at low expense worldwide, the Work Group felt many were sufficiently important to warrant an evidence grade of level 1, with the strength of the evidence varying in accordance to their potential to do harm in CKD patients. 2.3: Encourage lifestyle modification in patients with CKD to lower BP and improve long-term cardiovascular and other outcomes: 2.3.1: We recommend achieving or maintaining a healthy weight (BMI 20 to 25). (1D) RATIONALE Weight reduction lowers BP in the general population. Observational studies show that weight-loss strategies reduce BP in CKD patients. Weight-reduction strategies may result in other health benefits to CKD patients including reduction in urine albumin or protein levels, improved lipid profile and increased insulin sensitivity.

Weight reduction lowers BP in the general population. Observational studies show that weight-loss strategies reduce BP in CKD patients. Weight-reduction strategies may result in other health benefits to CKD patients including reduction in urine albumin or protein levels, improved lipid profile and increased insulin sensitivity. The prevalence of obesity is very high in Western countries and is increasing rapidly in developed and developing countries around the world. A strong relationship exists between body weight (usually defined as BMI) and BP levels in the general population.50, 51, 52 Compared with a person of normal weight, individuals who are overweight or obese tend to have higher BP levels, abnormalities in a range of other cardiovascular parameters (e.g., dyslipidemia52), and an increased risk of cardiovascular events.

(usually defined as BMI) and BP levels in the general population.50, 51, 52 Compared with a person of normal weight, individuals who are overweight or obese tend to have higher BP levels, abnormalities in a range of other cardiovascular parameters (e.g., dyslipidemia52), and an increased risk of cardiovascular events. Weight and BP. A weight-reducing diet has been clearly demonstrated to lower BP in overweight individuals in the general population. A systematic review53 published in 2006 identified 14 trials assessing the effects of dietary modification on BP in the general population, all but two of which assessed the effects of weight reduction in overweight persons. Many of the 14 trials also included other modifications to diet (e.g., increased fruit and vegetable intake and salt reduction) and lifestyle (e.g., increased exercise). Trials were 8 to 52 weeks in duration and mostly included participants with elevated BP levels. The quality of the trials was generally suboptimal. Overall, dietary modification reduced systolic BP by 6.0 mm Hg (95% confidence interval [CI] 3.4–8.6) and diastolic BP by 4.8 mm Hg (95% CI 2.7–6.9). High levels of heterogeneity in the trial results were observed.

mostly included participants with elevated BP levels. The quality of the trials was generally suboptimal. Overall, dietary modification reduced systolic BP by 6.0 mm Hg (95% confidence interval [CI] 3.4–8.6) and diastolic BP by 4.8 mm Hg (95% CI 2.7–6.9). High levels of heterogeneity in the trial results were observed. The available data regarding the effects of weight loss in CKD patients has been systematically reviewed by Navaneethan et al.54 Only two randomized trials were identified but 11 observational studies were also included. A range of surgical and non-surgical weight-loss interventions were assessed. All interventions, when taken together, resulted in significant reduction in weight among CKD patients. This was associated with a reduction in urinary protein excretion (described in two studies) but no overall effect on the GFR, possibly due to the short term nature of the studies. Effects on BP were not reported in the RCTs, whereas the observational studies reported consistently large, significant reductions in BP compared to baseline with both non-surgical weight loss (weighted mean difference in BP 9.0 mm Hg; 95% CI 3.7–14.2 mm Hg; P<0.0001) as well as surgical weight loss (weighted mean difference, 22.6 mm Hg; 95% CI 19.1–26.2; P<0.0001). Thus, weight loss likely improves BP in patients with CKD, although high-quality RCTs are needed to confirm this finding.

n-surgical weight loss (weighted mean difference in BP 9.0 mm Hg; 95% CI 3.7–14.2 mm Hg; P<0.0001) as well as surgical weight loss (weighted mean difference, 22.6 mm Hg; 95% CI 19.1–26.2; P<0.0001). Thus, weight loss likely improves BP in patients with CKD, although high-quality RCTs are needed to confirm this finding. Body weight and outcomes. In the general population, overweight and obesity have been clearly shown to be associated with an increased risk of cardiovascular events and death.52 A J-curve relationship has been described in many reports, revealing an increased risk in underweight individuals (e.g., those with a BMI <18.5) as well. RCTs have demonstrated that weight loss reduces the incidence of diabetes,55 but any beneficial effects on cardiovascular outcomes or survival remain to be proven. Indeed, a number of RCTs involving use of pharmacological agents to induce weight loss have been stopped early owing to unintended and unanticipated adverse effects of the agent being assessed (e.g., rimonabant and sibutramine).56, 57

ut any beneficial effects on cardiovascular outcomes or survival remain to be proven. Indeed, a number of RCTs involving use of pharmacological agents to induce weight loss have been stopped early owing to unintended and unanticipated adverse effects of the agent being assessed (e.g., rimonabant and sibutramine).56, 57 The data are less clear for patients with CKD. Obesity has been proposed as a possible potentiator of CKD progression; however, reliable data remain sparse. Many observational studies have suggested that among patients with advanced CKD who are dialysis-dependent, and particularly hemodialysis-dependent, clinical outcomes might actually be better for overweight individuals than for non-overweight individuals.58, 59 Other studies have reported conflicting results.60 It is possible that these observations are due to reverse causality, with the results driven by underlying malnutrition or inflammation in the lower-weight patients and they may also reflect differences in the proportions of muscle and fat in patients with CKD compared with people without CKD. These data should therefore be interpreted with caution. For overweight individuals, the method used to reduce body weight may be important within the context of CKD. Popular and widely recommended weight-loss diets are commonly high in potassium and protein and may therefore increase risks of hyperkalemia and CKD progression in patients with CKD. As the potential benefits and harms have not been specifically addressed in the CKD population, the use of these diets is not recommended.

opular and widely recommended weight-loss diets are commonly high in potassium and protein and may therefore increase risks of hyperkalemia and CKD progression in patients with CKD. As the potential benefits and harms have not been specifically addressed in the CKD population, the use of these diets is not recommended. Overall, the available data suggest that achieving or maintaining a body weight in the healthy range will lead to improved BP levels and better long-term CKD outcomes. This is particularly clear for individuals with CKD stages 1–2. Caution should be exercised in patients with more advanced CKD, because malnutrition may be associated with adverse outcomes. Since a high weight may be protective in CKD 5D patients, there could be risks associated with encouraging weight loss in those with advanced CKD. Hence, Recommendation 2.3.1 was graded 1D.2.3.2: We recommend lowering salt intake to <90 mmol (<2 g) per day of sodium (corresponding to 5 g of sodium chloride), unless contraindicated. (1C) RATIONALE Lowering salt intake reduces BP in the general population. In CKD patients with reduced GFR, salt retention is associated with an increase in BP.

Overall, the available data suggest that achieving or maintaining a body weight in the healthy range will lead to improved BP levels and better long-term CKD outcomes. This is particularly clear for individuals with CKD stages 1–2. Caution should be exercised in patients with more advanced CKD, because malnutrition may be associated with adverse outcomes. Since a high weight may be protective in CKD 5D patients, there could be risks associated with encouraging weight loss in those with advanced CKD. Hence, Recommendation 2.3.1 was graded 1D.2.3.2: We recommend lowering salt intake to <90 mmol (<2 g) per day of sodium (corresponding to 5 g of sodium chloride), unless contraindicated. (1C) RATIONALE Lowering salt intake reduces BP in the general population. In CKD patients with reduced GFR, salt retention is associated with an increase in BP. A relationship between average daily salt intake and BP levels has long been recognized, leading to calls from the World Health Organization (WHO) for salt intake to be restricted to improve BP levels (http://www.who.int/cardiovascular_diseases/guidelines/Full%20text.pdf).61 Restricting salt intake clearly lowers BP by a moderate amount, as demonstrated in a systematic review of seven trials,53 most of which assessed the impact of restricting salt intake to 4 to 6 g (70–100 mmol). Overall, BP levels were reduced as compared to baseline levels: systolic BP by 4.7 mm Hg (95% CI 2.2–7.2) and diastolic BP by 2.5 mm Hg (95% CI 1.8–3.3). Moderate heterogeneity was observed in the effects on systolic BP, but this was corrected when one outlier trial was excluded. Other systematic reviews including a different group of trials have suggested similar but somewhat smaller benefits.62

4.7 mm Hg (95% CI 2.2–7.2) and diastolic BP by 2.5 mm Hg (95% CI 1.8–3.3). Moderate heterogeneity was observed in the effects on systolic BP, but this was corrected when one outlier trial was excluded. Other systematic reviews including a different group of trials have suggested similar but somewhat smaller benefits.62 Alterations in salt handling are likely to be a significant contributor to elevated BP levels in patients with CKD. Although no large scale long term RCTs of salt restriction in CKD patients were found, there is no reason to believe that BP reductions should not also be observed. Reducing salt intake could have a greater capacity to lower BP in patients with CKD who have salt and water retention and this intervention should be routinely discussed with such individuals. A low-sodium diet has been shown to further reduce BP and urine albumin or protein levels in the short term in patients on ARBs63, 64, 65, 66 and may be a consideration for those with high BP who have a poor response to ACE-Is or ARBs. Some forms of CKD may be associated with salt wasting from the kidney. Affected individuals may be at higher than usual risk of volume depletion and electrolyte disturbances potentiated by salt restriction. Volume and electrolyte status should thus be carefully monitored in patients with CKD undergoing salt restriction. Recent studies suggesting that low urinary sodium excretion (hence perhaps low dietary sodium intake) associates with higher mortality in diabetes have yet to be confirmed by others or explained.67, 68

on. Volume and electrolyte status should thus be carefully monitored in patients with CKD undergoing salt restriction. Recent studies suggesting that low urinary sodium excretion (hence perhaps low dietary sodium intake) associates with higher mortality in diabetes have yet to be confirmed by others or explained.67, 68 Since salt restriction is an inexpensive and important contributor to lowering BP in the generally population worldwide, this intervention was deemed a level 1 recommendation. But since the evidence base for CKD patients included only small, short-term RCTs involving special circumstances, Recommendation 2.3.2 was graded 1C.2.3.3: We recommend undertaking an exercise program compatible with cardiovascular health and tolerance, aiming for at least 30 minutes 5 times per week. (1D) RATIONALE Increased physical exercise has been linked to a broad range of positive health outcomes through a wide variety of mechanisms. A clear inverse relationship between exercise and average daily BP has been demonstrated by a large volume of previous epidemiological data in the general population, although exercise may lead to modest and acute physiological increases in BP during the time of the activity.

comes through a wide variety of mechanisms. A clear inverse relationship between exercise and average daily BP has been demonstrated by a large volume of previous epidemiological data in the general population, although exercise may lead to modest and acute physiological increases in BP during the time of the activity. The effects of exercise on BP in the context of RCTs have been systematically reviewed in the general population.53 Most of the 21 RCTs included in the review examined the efficacy of 3 to 5 weekly sessions of aerobic exercise lasting 30 to 60 minutes. Overall, the exercise group had an average reduction in systolic BP of 6.1 mm Hg from baseline (95% CI 2.1–10.1) and in diastolic BP of 3.0 mm Hg (95% CI 1.1–4.9). The effects were slightly reduced when one outlier trial was excluded from the analysis (to average reductions of 4.6 and 2.6 mm Hg, respectively), but moderate heterogeneity among the results remained. No RCTs in the CKD population were found. A post hoc observational analysis69 of the Modification of Diet in Renal Disease (MDRD) study population did not identify a clear relationship between level of physical activity at baseline and the subsequent risk of death, although trends toward better outcomes for active individuals were observed. Two larger studies from the US Renal Data System found that CKD 5D patients who are sedentary have a higher risk of death than those who are active.70, 71 All of these studies are observational and more data are required.

equent risk of death, although trends toward better outcomes for active individuals were observed. Two larger studies from the US Renal Data System found that CKD 5D patients who are sedentary have a higher risk of death than those who are active.70, 71 All of these studies are observational and more data are required. The benefits of exercise on BP and on general health appear likely to be similar in the CKD and the general population, with no strong rationale for different recommendations. On this basis, Recommendation 2.3.3 was graded 1D.2.3.4: We suggest limiting alcohol intake to no more than two standard drinks per day for men and no more than one standard drink per day for women. (2D) RATIONALE Alcohol has been shown to produce both acute and chronic increases in BP, suggesting that restricting alcohol intake would lower BP. In a systematic review of four trials,53 restricting alcohol intake in the general population resulted in a 3.8 mm Hg reduction (95% CI 1.4–6.1) in systolic BP and a 3.2 mm Hg reduction (95% CI 1.4–5.0) in diastolic BP, with no evidence of heterogeneity among the results. No data specific to CKD patients were found, but the effects are expected to be similar.

restricting alcohol intake in the general population resulted in a 3.8 mm Hg reduction (95% CI 1.4–6.1) in systolic BP and a 3.2 mm Hg reduction (95% CI 1.4–5.0) in diastolic BP, with no evidence of heterogeneity among the results. No data specific to CKD patients were found, but the effects are expected to be similar. Most data suggest that up to two standard drinks per day for a man and 1 standard drink per day for a woman are likely to be safe. The definition of a standard drink varies from 8 to 19.7 g of alcohol in different countries (see http://whqlibdoc.who.int/hq/2000/who_msd_msb_00.4.pdf).72 10 g of alcohol is equivalent to 30 ml of spirits, 100 ml of wine, 285 ml of full-strength beer, and 425 ml of light beer. The benefits of alcohol moderation on BP and on general health appear likely to be similar in the CKD and the general population, with no strong rationale for different recommendations. On this basis, Recommendation 2.3.4 was graded 2D. OTHER INTERVENTIONS Cigarette smoking. Cigarette smoking and exposure to environmental tobacco smoke are clearly among the most potent modifiable risk factors for CVD in the general population and in patients with CKD. Although it does not have a clear, direct impact on long-term BP, the avoidance of exposure to cigarette smoke is a critical aspect of cardiovascular risk reduction but as yet there are no RCTs in the CKD population.

ong the most potent modifiable risk factors for CVD in the general population and in patients with CKD. Although it does not have a clear, direct impact on long-term BP, the avoidance of exposure to cigarette smoke is a critical aspect of cardiovascular risk reduction but as yet there are no RCTs in the CKD population. Dietary supplementation. The effects of potassium supplementation on BP have been assessed in a number of studies.53 These have produced conflicting results, with some but not all indicating a benefit. CKD patients often have reduced capacity for potassium excretion, particularly as the GFR falls, such that the risk of hyperkalemia may be increased. In the absence of specific studies demonstrating a benefit in CKD patients, we cannot recommend potassium supplementation to reduce BP in patients with CKD. The evidence base for magnesium supplementation is similar, with some but not all studies suggesting a benefit with respect to BP.53, 73 Although hypermagnesemia is not a common problem in CKD patients, magnesium supplementation cannot be recommended without specific data demonstrating its safety and efficacy. Fish-oil supplementation has been shown to produce small but significant reductions in BP in a number of RCTs and systematic reviews.53, 74 The mechanisms of these effects remain uncertain, however and the safety of fish oil has not been clearly demonstrated in CKD patients. Although some data supporting the use of fish oil exists for patients with IgA nephropathy,75 it is premature to recommend this treatment for BP lowering in the CKD population.

.53, 74 The mechanisms of these effects remain uncertain, however and the safety of fish oil has not been clearly demonstrated in CKD patients. Although some data supporting the use of fish oil exists for patients with IgA nephropathy,75 it is premature to recommend this treatment for BP lowering in the CKD population. BP-LOWERING AGENTS RCTs involving both CKD and non-CKD populations in which a target BP has been set at the levels recommended in this Guideline clearly show that most patients will require two or more antihypertensive agents to achieve these targets. Surveys of BP control in CKD patients indicate that three or more agents are frequently needed. With the exception of ARBs or ACE-Is in CKD patients with high levels of urinary albumin or protein excretion, there is no strong evidence to support the preferential use of any particular agent(s) in controlling BP in CKD; nor are there data to guide the clinician in the choice of second- and third-line medications. Since the 2004 KDOQI Guideline1 was published, there has been an increasing trend towards tailoring antihypertensive therapy to the individual patient, taking into account issues such as the presence or absence of high urine albumin excretion, co-morbidities, concomitant medications, adverse effects, and availability of the agents. Ultimately, the choice of agents is less important than the actual reduction in BP achieved, since BP reduction is the major measurable outcome in the individual patient.

as the presence or absence of high urine albumin excretion, co-morbidities, concomitant medications, adverse effects, and availability of the agents. Ultimately, the choice of agents is less important than the actual reduction in BP achieved, since BP reduction is the major measurable outcome in the individual patient. Other information of value in deciding on the optimal BP lowering regimen include data on drug half-life and dose adjustments in CKD stage 5D, which may be of help in guiding the use of BP lowering drugs in advanced CKD ND.4, 76 The optimal timing of administration of medication has not been studied in CKD patients. CKD patients who do not have the normal decrease in BP during sleep (non-dippers and reverse dippers) have worse cardiovascular and kidney outcomes when compared to dippers.11, 12, 77, 78, 79 Whether the recently reported strategy of evening dosing to produce nocturnal dipping will improve outcomes in CKD patients, as has been described in individuals with essential hypertension, remains to be established.80, 81, 82

s) have worse cardiovascular and kidney outcomes when compared to dippers.11, 12, 77, 78, 79 Whether the recently reported strategy of evening dosing to produce nocturnal dipping will improve outcomes in CKD patients, as has been described in individuals with essential hypertension, remains to be established.80, 81, 82 The ERT was not asked to search for evidence of the effectiveness of established anti-hypertensive agents in lowering BP in patients with CKD, since it is generally believed that all such drugs are effective, although the sensitivity in individual patients may vary, as may be the side effects. Instead, the ERT focused on two issues. Firstly, studies that compared different BP targets were identified. In these studies, only the BP targets were randomized; the protocols varied with respect to the sequence of drugs and escalation of dose. Secondly the ERT searched for studies that included a comparison of different combinations of anti-hypertensive agents. In these studies, only the choice of first-line drug was randomized, with study protocols varying with respect to drug dose, use of concomitant agents and BP thresholds for drug titration (Table 5, see Methods for Guideline Development).

that included a comparison of different combinations of anti-hypertensive agents. In these studies, only the choice of first-line drug was randomized, with study protocols varying with respect to drug dose, use of concomitant agents and BP thresholds for drug titration (Table 5, see Methods for Guideline Development). The KDOQI Clinical Practice Guidelines on Hypertension and Antihypertensive Agents in Chronic Kidney Disease (http://www.kidney.org/professionals/KDOQI/guidelines_bp/index.htm)1 contains details of clinical pharmacology and practical guidance on the use of the various agents to lower BP in CKD patients. Information on CKD- and CVD-related indications, side effects, dosages and contraindications relevant to all commonly used anti-hypertensive agents as well as strategies to improve adherence and warnings regarding the hazards of certain combinations are also noted therein. The Work Group believed that there was insufficient new evidence to warrant rewriting the clear guidance provided in the KDOQI Guideline. However, at the request of the KDIGO Board, the Work Group summarize specific aspects of the use of antihypertensive agents in CKD patients. We outline the information that can be drawn from the known pharmacology of agents or observations in non-CKD patients, emphasizing the difficulty in extrapolating to CKD patients, especially those with advanced CKD.

Board, the Work Group summarize specific aspects of the use of antihypertensive agents in CKD patients. We outline the information that can be drawn from the known pharmacology of agents or observations in non-CKD patients, emphasizing the difficulty in extrapolating to CKD patients, especially those with advanced CKD. Given that the prescribed drug regimen commonly involves many medications, it is reasonable to use strategies that might maximize the likelihood of adherence, including the use of cheaper drugs, convenient frequency of dosing and reduction in pill numbers. This can be achieved by prescribing once-daily medication and combination pills (which are simpler to take and in some circumstances may be less expensive than the individual agents) when possible.83 Renin–angiotensin–aldosterone system blockers Because of its pivotal role in regulation of BP, the RAAS system is an obvious target for BP-lowering medications. Although other agents, particularly beta-blockers, interfere with the RAAS pathway, the main RAAS inhibitors are ACE-Is, ARBs, aldosterone antagonists, and DRIs.

Renin–angiotensin–aldosterone system blockers Because of its pivotal role in regulation of BP, the RAAS system is an obvious target for BP-lowering medications. Although other agents, particularly beta-blockers, interfere with the RAAS pathway, the main RAAS inhibitors are ACE-Is, ARBs, aldosterone antagonists, and DRIs. ACE-Is and ARBs. ACE-Is block the conversion of angiotensin I to angiotensin II and the degradation of bradykinin. It seems likely that the accumulation of bradykinin leads to persistent dry cough, a recognized side effect which occurs in 5 to 20% of patients on ACE-Is. Angioneurotic edema can occur with both ACE-Is and ARBs, although the relative frequencies and the mechanism are not clear. ARBs act by competitively antagonizing the interaction between angiotensin II and angiotensin receptors and were first introduced as an alternative to ACE-Is in patients who had an ACE-I induced cough. ACE-Is and ARBs are valuable BP-reducing agents in CKD patients, are indicated if urinary albumin excretion is elevated and are safe to combine with most other BP-reducing agents. Clinically significant hyperkalemia and reductions in GFR can occur in patients receiving ACE-Is or ARBs, particularly in those who have renal-artery stenosis or reduced intravascular volume, or when these agents are used together with NSAIDs, COX-2 inhibitors, or potassium-sparing diuretics. The use of these drugs in women of child-bearing age should be balanced with the risk of pregnancy since they are potentially teratogenic (see Chapter 6).84, 85

ve renal-artery stenosis or reduced intravascular volume, or when these agents are used together with NSAIDs, COX-2 inhibitors, or potassium-sparing diuretics. The use of these drugs in women of child-bearing age should be balanced with the risk of pregnancy since they are potentially teratogenic (see Chapter 6).84, 85 The sequential marketing of ACE-Is first (captopril in 1977) and ARBs later (losartan in 1995) has influenced the design of RCTs involving these drug classes. The first large-scale RCT of RAAS blockade in diabetes involved patients with type 1 disease given captopril. By the time ARBs were introduced, the benefits of ACE-Is (in CKD patients with type 1 diabetes) were well established. Thus RCTs involving ARBs generally targeted individuals with type 2 diabetes. This has led to some bias in the evidence base underpinning recommendations for using ACE-Is or ARBs in the treatment of BP. There is no substantive evidence to suggest that ACE-Is and ARBs differ in their ability to reduce BP in patients with essential hypertension.86 In most health care settings, ACE-Is are less expensive than ARBs, which may influence the choice between an ACE-I or ARB. The most prominent BP-related effects of the blockade of angiotensin II by ACE-Is or ARBs are as follows: Generalized arterial vasodilatation, resulting in lower BP.

The sequential marketing of ACE-Is first (captopril in 1977) and ARBs later (losartan in 1995) has influenced the design of RCTs involving these drug classes. The first large-scale RCT of RAAS blockade in diabetes involved patients with type 1 disease given captopril. By the time ARBs were introduced, the benefits of ACE-Is (in CKD patients with type 1 diabetes) were well established. Thus RCTs involving ARBs generally targeted individuals with type 2 diabetes. This has led to some bias in the evidence base underpinning recommendations for using ACE-Is or ARBs in the treatment of BP. There is no substantive evidence to suggest that ACE-Is and ARBs differ in their ability to reduce BP in patients with essential hypertension.86 In most health care settings, ACE-Is are less expensive than ARBs, which may influence the choice between an ACE-I or ARB. The most prominent BP-related effects of the blockade of angiotensin II by ACE-Is or ARBs are as follows: Generalized arterial vasodilatation, resulting in lower BP. Vasodilatation of the efferent and afferent glomerular arterioles, particularly the efferent, resulting in decreased intra-glomerular pressure and hence reduction in both GFR and urine albumin excretion. This is believed to result in some degree of long-term renoprotection, at least in patients with albuminuria.87 On initiation of therapy a reversible reduction in GFR of up to 30% (accordingly a 30% increase in SCr concentration) has been regarded as reasonably attributable to this physiological mechanism. Greater reductions may indicate underlying renal artery stenosis.1, 88 It has been suggested that in advanced CKD, cessation of RAAS blockade may allow an increase in GFR of sufficient magnitude to delay end-stage kidney failure.23 This concept is further discussed in Chapter 8.

butable to this physiological mechanism. Greater reductions may indicate underlying renal artery stenosis.1, 88 It has been suggested that in advanced CKD, cessation of RAAS blockade may allow an increase in GFR of sufficient magnitude to delay end-stage kidney failure.23 This concept is further discussed in Chapter 8. Reduction in adrenal secretion of aldosterone. In about 50% of subjects prescribed ACE-Is or ARBs, aldosterone production is restored to at least pre-treatment levels over a period of months (a phenomenon termed aldosterone breakthrough).89 This may explain the efficacy of aldosterone antagonists in patients already taking an ACE-I or ARB. ACE-Is and ARBs may have other effects, including inhibition of fibrosis and enhancement of vascular and cardiac remodelling. Discussion of these effects, which may be of relevance to renoprotection, is beyond the scope of this Guideline.

Reduction in adrenal secretion of aldosterone. In about 50% of subjects prescribed ACE-Is or ARBs, aldosterone production is restored to at least pre-treatment levels over a period of months (a phenomenon termed aldosterone breakthrough).89 This may explain the efficacy of aldosterone antagonists in patients already taking an ACE-I or ARB. ACE-Is and ARBs may have other effects, including inhibition of fibrosis and enhancement of vascular and cardiac remodelling. Discussion of these effects, which may be of relevance to renoprotection, is beyond the scope of this Guideline. Dose considerations in CKD patients. Most available ACE-Is have active moieties that are largely excreted in the urine. Fosinopril and trandolapril are partially (in general, approximately 50%) excreted by the liver, such that the blood levels are less influenced by kidney failure than levels of other ACE-Is which are predominantly excreted by the kidneys. Since ACE-Is are generally titrated to achieve optimal clinical effect, the mode of excretion is not regarded as a major factor in dosing.76 If hyperkalemia occurs in CKD patients taking a renal excreted ACE-I, possible interventions include dietary advice, reducing the dose, switching to fosinopril or trandolapril, or adding a potassium-losing diuretic. All ARBs are substantially excreted by the liver, with the proportion of drug elimination ranging from 40% (in the case of candesartan) to >95% (in the case of irbesartan and telmisartan). As with ACE-Is, the dose in ARBs is usually adjusted according to clinical effect rather than kidney function.76

Dose considerations in CKD patients. Most available ACE-Is have active moieties that are largely excreted in the urine. Fosinopril and trandolapril are partially (in general, approximately 50%) excreted by the liver, such that the blood levels are less influenced by kidney failure than levels of other ACE-Is which are predominantly excreted by the kidneys. Since ACE-Is are generally titrated to achieve optimal clinical effect, the mode of excretion is not regarded as a major factor in dosing.76 If hyperkalemia occurs in CKD patients taking a renal excreted ACE-I, possible interventions include dietary advice, reducing the dose, switching to fosinopril or trandolapril, or adding a potassium-losing diuretic. All ARBs are substantially excreted by the liver, with the proportion of drug elimination ranging from 40% (in the case of candesartan) to >95% (in the case of irbesartan and telmisartan). As with ACE-Is, the dose in ARBs is usually adjusted according to clinical effect rather than kidney function.76 ACE-Is and ARBs should be used with caution or even avoided in certain CKD subgroups, particularly in patients with bilateral renal-artery stenosis or with intravascular fluid depletion, because of the risk of a large reduction in GFR. The normal capacity of the kidney to auto-regulate GFR in the face of fluctuations in BP is impaired in CKD and further compromised by the use of ACE-Is or ARBs. Hypotension (e.g., as a result of hypovolemia or sepsis) may cause an acute decline in GFR in patients with CKD taking ACE-Is or ARBs.90 Several case series have reported a high risk of acute kidney injury in diabetic patients on an ACE-I or ARB during sepsis91, 92, 93 and when they are used in combination with NSAIDs94 or diuretics.95 Reducing the dose or holding off on using ACE-Is or ARBs until recovery is sensible in patients who develop inter-current illnesses that lead to dehydration as a result of diarrhea, vomiting, or high fever.

ts on an ACE-I or ARB during sepsis91, 92, 93 and when they are used in combination with NSAIDs94 or diuretics.95 Reducing the dose or holding off on using ACE-Is or ARBs until recovery is sensible in patients who develop inter-current illnesses that lead to dehydration as a result of diarrhea, vomiting, or high fever. Indications for ACE-Is and ARBs. In this guideline, ACE-Is and ARBs are recommended for specific groups of CKD patients with increased urinary albumin excretion in which context use of these agents may be associated with better kidney96 and cardiovascular outcomes.97 In non-CKD patients, these drugs are indicated for the treatment of heart failure and for use soon after myocardial infarction, stroke, and in patients with high cardiovascular risk.98, 99, 100

y albumin excretion in which context use of these agents may be associated with better kidney96 and cardiovascular outcomes.97 In non-CKD patients, these drugs are indicated for the treatment of heart failure and for use soon after myocardial infarction, stroke, and in patients with high cardiovascular risk.98, 99, 100 The Oregon Health Resources Commission reported in 2005 on the use of ACE-Is in essential hypertension. No differences were found among various ACE-Is in terms of the BP-lowering effect and serious complications which were independent of gender, age, or African-American heritage.99 In 2006, the Commission reviewed the evidence for the use of ARBs.100 It reported that there were no data to suggest that any particular ARB was superior to another in the context of a variety of clinical scenarios, including essential hypertension and high cardiovascular risk; nor was there evidence of any ARB being associated with a higher risk of serious complications or differences in efficacy or side effects regardless of age, race, or gender. In reviewing studies specifically involving patients with CKD, no important differences in the effect of ARBs on BP or side effects were found.

k; nor was there evidence of any ARB being associated with a higher risk of serious complications or differences in efficacy or side effects regardless of age, race, or gender. In reviewing studies specifically involving patients with CKD, no important differences in the effect of ARBs on BP or side effects were found. Accordingly, ACE-Is or ARBs might be considered for use in patients with CKD who have heart failure, recent myocardial infarction, a history of stroke, or a high cardiovascular risk. However, it is not possible to make any recommendations for CKD patients in particular, since the data are largely from studies of non-CKD patients. In addition, because CKD patients are at higher risk of side effects, particularly hyperkalemia and reduction in GFR, the use of ACE-Is or ARBs may not have the same risk-to-benefit ratio in CKD patients as in non-CKD populations. Drug combinations. The antihypertensive and anti-albuminuric effects of ACE-Is and ARBs are complemented by dietary sodium restriction or administration of diuretics.63, 65, 66 ACE-Is and ARBs are therefore valuable adjuncts to diuretics for the treatment of high BP and vice versa. Co-administration of beta-blockers and calcium-channel blockers with ACE-Is or ARBs is also acceptable. One recent post hoc analysis of a large trial involving hypertensive individuals demonstrated that a combination of an ACE-I (benazepril) and calcium antagonist (amlodipine) was superior to the same ACE-I used with a diuretic (hydrochlorothiazide) in slowing CKD progression.101

s with ACE-Is or ARBs is also acceptable. One recent post hoc analysis of a large trial involving hypertensive individuals demonstrated that a combination of an ACE-I (benazepril) and calcium antagonist (amlodipine) was superior to the same ACE-I used with a diuretic (hydrochlorothiazide) in slowing CKD progression.101 Patients given NSAIDs, COX-2 antagonists or potassium-sparing diuretics can develop hyperkalemia if these drugs are used in combination with ACE-Is or ARBs. The combination of ACE-Is and/or ARBs with aldosterone-blocking antagonists is an area of current controversy that is covered in more detail below and in Chapter 8.

Patients given NSAIDs, COX-2 antagonists or potassium-sparing diuretics can develop hyperkalemia if these drugs are used in combination with ACE-Is or ARBs. The combination of ACE-Is and/or ARBs with aldosterone-blocking antagonists is an area of current controversy that is covered in more detail below and in Chapter 8. Aldosterone antagonists. The aldosterone antagonist spirolactone has been in use as a BP-lowering agent since the late 1950s. Prescribed as a diuretic in the treatment of edema and resistant hypertension, it fell into disuse with the advent of more powerful diuretics and antihypertensives. With the high doses initially used (up to 300 mg/day), spironolactone use was associated with side effects, particularly those due to its estrogen-like activity (gynecomastia and menstrual disturbances). Recognition that BP-lowering could be achieved with much lower doses of spironolactone (12.5–50 mg/day) has led to renewed interest in aldosterone antagonists over the past decade.102, 103, 104, 105 As a result, eplerenone, a mineralocorticoid-receptor blocker without estrogen-like effects, has been developed. In CKD, the major emphasis has been on using aldosterone antagonists to reduce urine albumin levels and as an adjunct to other antihypertensive agents in treating resistant hypertension. Aldosterone antagonists are of proven benefit in non-CKD patients with heart failure, including heart failure after myocardial infarction. Because of the risk of hyperkalemia and reduction in GFR, they should be used with caution in CKD patients.

unct to other antihypertensive agents in treating resistant hypertension. Aldosterone antagonists are of proven benefit in non-CKD patients with heart failure, including heart failure after myocardial infarction. Because of the risk of hyperkalemia and reduction in GFR, they should be used with caution in CKD patients. Dose considerations in CKD patients. Impaired renal excretion of native drug or active metabolites of spironolactone and eplerenone and an increased risk of hyperkalemia may limit their use in patients with CKD. Plasma potassium levels and kidney function should be monitored closely during the introduction of aldosterone antagonists and during intercurrent illnesses, particularly those associated with a risk of GFR reduction, as occurs with dehydration. Indications for aldosterone antagonists. In patients without CKD, aldosterone antagonists are recommended for the treatment of severe cardiac failure that is resistant to other therapies and for use after acute myocardial infarction complicated by cardiac failure. These agents also have a place in the management of essential hypertension that is resistant to other therapies. It is unclear whether this information can be extrapolated to CKD patients, particularly those with advanced CKD in whom the risks associated with the use of aldosterone antagonists, particularly of hyperkalemia, may be increased.