Revisiting the treatment of anemia in the setting of chronic kidney disease, hematologic malignancies, and cancer: perspectives with opinion and commentary.

INTRODUCTION: Anemia has and will continue to be a central theme in medicine particularly as clinicians are treating a burgeoning population of complex multi-organ system processes. As a result of multiple randomized controlled trials (RCTs), meta-analyses, and societal recommendations overly restrictive paradigms and under-administration of erythropoiesis stimulating agents (ESAs) have likely been followed by clinicians among all specialties. AREAS COVERED: A review of anemia in the context of chronic kidney disease, hematologic malignancies, and cancer is presented with focus on the establishment of ESAs as integral in the treatment of anemia. Multiple RCTs and meta-analyses studying the use of ESAs are presented with focus upon their application to clinical practice. A 'compendium' is proffered describing the evolution, establishment, and implications of ESA administration initially among those with CKD with rapid subsequent application to the Hematology-Oncology population of patients. Literature search methodologies have included MEDLINE (1985-2020), PubMed (1996-2020), Cochrane Central Trials (1985-2020), EMBASE (2000-2020), and ClinicalTrials.gov (2000-2020). EXPERT OPINION: Upon evaluation of risks and benefits of ESAs focused opinion and commentary is made supporting more liberal use of these agents and strongly suggesting that the current underlying treatment 'pendulum' has perhaps shifted too far to the 'under-treatment' side in many cases.

Synergistic Reduction of Apoptosis With Diazoxide and Erythropoietin in Spinal Cord Ischemic Injury.

BACKGROUND: Paraplegia remains a devastating complication of thoracoabdominal aortic intervention. Metabolic stress induces expression of beta common receptor subunit of erythropoietin (EPO) receptor (ßcR) to exert a neuroprotective effect in spinal cord ischemia reperfusion injury (SCIR). Diazoxide (DZ) has been shown to induce ischemic tolerance. We previously reported that DZ upregulated ßcR expression and enhanced the neuroprotective effects of EPO through the upregulation of ßcR. We hypothesize that ßcR expression induced by DZ before ischemia amplifies the antiapoptotic effects of EPO in a murine model of SCIR. METHODS: Experimental groups included phosphate-buffered saline (PBS) pretreatment + PBS immediately before the operation, PBS+EPO, DZ+PBS, DZ+EPO, and sham. Spinal cord ischemia was induced by a 4-minute thoracic aortic cross-clamp. Functional scoring (Basso Mouse Score) was done at 12-hour intervals for 48 hours. Spinal cords were harvested for histologic analysis, and antiapoptotic factors (caspase 3, 8, and 9, B-cell lymphoma-2, and neuroglobin) were evaluated by Western blot analysis. RESULTS: The motor function of DZ+EPO group was significantly preserved compared with all other groups. The levels of cleaved caspase 8 and 3 in DZ+EPO were significantly lower than in the other groups. Mice treated with DZ+EPO had significantly fewer terminal deoxynucleotide transferase-mediated deoxy uridine triphosphate nick-end labeling-positive cells than other groups. CONCLUSIONS: Optimized upregulation of ßcR by DZ can increase the extrinsic antiapoptotic effects of EPO. Better understanding of this synergetic mechanism may serve to help prevent ischemic complications caused by aortic intervention.

Activation of the ß-common receptor by erythropoietin impairs acetylcholine-mediated vasodilation in mouse mesenteric arterioles.

Clinically, erythropoietin (EPO) is known to increase systemic vascular resistance and arterial blood pressure. However, EPO stimulates the production of the potent vasodilator, nitric oxide (NO), in culture endothelial cells. The mechanism by which EPO causes vasoconstriction despite stimulating NO production may be dependent on its ability to activate two receptor complexes, the homodimeric EPO (EPOR2 ) and the heterodimeric EPOR/ß-common receptor (ßCR). The purpose of this study was to investigate the contribution of each receptor to the vasoactive properties of EPO. First-order, mesenteric arteries were isolated from 16-week-old male C57BL/6 mice, and arterial function was studied in pressure arteriographs. To determine the contribution of each receptor complex, EPO-stimulating peptide (ESP), which binds and activates the heterodimeric EPOR/ßCR complex, and EPO, which activates both receptors, were added to the arteriograph chamber 20 min prior to evaluation of endothelium-dependent (acetylcholine, bradykinin, A23187) and endothelium-independent (sodium nitroprusside) vasodilator responses. Only ACh-induced vasodilation was impaired in arteries pretreated with EPO or ESP. EPO and ESP pretreatment abolished ACh-induced vasodilation by 100% and 60%, respectively. EPO and ESP did not affect endothelium-independent vasodilation by SNP. Additionally, a novel ßCR inhibitory peptide (ßIP), which was computationally developed, prevented the impairment of acetylcholine-induced vasodilation by EPO and ESP, further implicating the EPOR/ßCR complex. Last, pretreatment with either EPO or ESP did not affect vasoconstriction by phenylephrine and KCl. Taken together, these findings suggest that acute activation of the heterodimeric EPOR/ßCR in endothelial cells leads to a selective impairment of ACh-mediated vasodilator response in mouse mesenteric resistance arteries.

Erythropoietin mimetic peptides and erythropoietin fusion proteins for treating anemia of chronic kidney disease.

PURPOSE OF REVIEW: First generation erythropoiesis stimulating agents (ESAs) have short duration of action which requires administration once weekly or greater. Second generation ESAs were developed which have longer duration of action and can be administered one to two times monthly. Erythropoietin (EPO) mimetic peptides (EMPs) activate the EPO receptor but have no structural analogy to EPO, offering the potential for lower cost as they are not biologic drugs. The first approved EMP, peginesatide, was withdrawn from the market within a year of its approval because of fatal anaphylactic reactions. In this review, we summarize recent progress regarding the development of newer, possibly less toxic, EMPs. We also summarize the development of EPO fusion proteins which fuse EPO with a portion of an immunoglobulin molecule or another EPO molecule, achieving a longer duration of action and less frequent dosing. RECENT FINDINGS: AGEM400(hydroxyethyl starch) and pegolsihematide are EMPs in phase II clinical trials. Three EPO fusion proteins are under development, two in phase I and one in phase II. SUMMARY: The future success of EMPs is limited by the prior experience with peginesatide and EPO fusion proteins do not offer cost savings or longer duration of action than currently available ESAs.

Lenalidomide Stabilizes the Erythropoietin Receptor by Inhibiting the E3 Ubiquitin Ligase RNF41.

In a subset of patients with non-del(5q) myelodysplastic syndrome (MDS), lenalidomide promotes erythroid lineage competence and effective erythropoiesis. To determine the mechanism by which lenalidomide promotes erythropoiesis, we investigated its action on erythropoietin receptor (EpoR) cellular dynamics. Lenalidomide upregulated expression and stability of JAK2-associated EpoR in UT7 erythroid cells and primary CD71+ erythroid progenitors. The effects of lenalidomide on receptor turnover were Type I cytokine receptor specific, as evidenced by coregulation of the IL3-Rα receptor but not c-Kit. To elucidate this mechanism, we investigated the effects of lenalidomide on the E3 ubiquitin ligase RNF41. Lenalidomide promoted EpoR/RNF41 association and inhibited RNF41 auto-ubiquitination, accompanied by a reduction in EpoR ubiquitination. To confirm that RNF41 is the principal target responsible for EpoR stabilization, HEK293T cells were transfected with EpoR and/or RNF41 gene expression vectors. Steady-state EpoR expression was reduced in EpoR/RNF41 cells, whereas EpoR upregulation by lenalidomide was abrogated, indicating that cellular RNF41 is a critical determinant of drug-induced receptor modulation. Notably, shRNA suppression of CRBN gene expression failed to alter EpoR upregulation, indicating that drug-induced receptor modulation is independent of cereblon. Immunohistochemical staining showed that RNF41 expression decreased in primary erythroid cells of lenalidomide-responding patients, suggesting that cellular RNF41 expression merits investigation as a biomarker for lenalidomide response. Our findings indicate that lenalidomide has E3 ubiquitin ligase inhibitory effects that extend to RNF41 and that inhibition of RNF41 auto-ubiquitination promotes membrane accumulation of signaling competent JAK2/EpoR complexes that augment Epo responsiveness. Cancer Res; 76(12); 3531-40. ©2016 AACR.

New Treatment Approaches for the Anemia of CKD.

Normocytic normochromic anemia is a common complication in chronic kidney disease and is associated with many adverse clinical consequences. Erythropoiesis-stimulating agents (ESAs) and adjuvant iron therapy represent the primary treatment for anemia in chronic kidney disease. The introduction of ESAs into clinical practice was a success story, mediating an increase in hemoglobin concentrations without the risk for recurrent blood transfusions and improving quality of life substantially. However, recombinant ESAs are still expensive and require a parenteral route of administration. Moreover, concern has arisen following randomized clinical trials showing that higher hemoglobin targets and/or high ESA doses may cause significant harm. This, together with changes in ESA reimbursement policy in some countries, has resulted in a significant reduction in ESA prescribing and the hemoglobin level targeted during therapy. Several attempts are being made to develop new drugs with improved characteristics and/or easier manufacturing processes compared with currently available ESAs, including new treatment approaches that may indirectly improve erythropoiesis. We give an update on the new investigational strategies for increasing erythropoiesis, examining in depth their characteristics and possible advantages in the clinical setting and the caveats to be aware of at the present stage of development.

Targeting EPO and EPO receptor pathways in anemia and dysregulated erythropoiesis.

INTRODUCTION: Recombinant human erythropoietin (rhEPO) is a first-line therapeutic for the anemia of chronic kidney disease, cancer chemotherapy, AIDS (Zidovudine therapy), and lower-risk myelodysplastic syndrome. However, rhEPO frequently elevates hypertension, is costly, and may affect cancer progression. Potentially high merit therefore exists for defining new targets for anti-anemia agents within erythropoietin (EPO) and EPO receptor (EPOR) regulatory circuits. AREAS COVERED: EPO production by renal interstitial fibroblasts is subject to modulation by several regulators of hypoxia-inducible factor 2a (HIF2a) including Iron Response Protein-1, prolyl hydroxylases, and HIF2a acetylases, each of which holds potential as anti-anemia drug targets. The cell surface receptor for EPO (EPOR) preassembles as a homodimer, together with Janus Kinase 2 (JAK2), and therefore it remains attractive to develop novel agents that trigger EPOR complex activation (activating antibodies, mimetics, small-molecule agonists). Additionally, certain downstream transducers of EPOR/JAK2 signaling may be druggable, including Erythroferrone (a hepcidin regulator), a cytoprotective Spi2a serpin, and select EPOR-associated protein tyrosine phosphatases. EXPERT OPINION: While rhEPO (and biosimilars) are presently important mainstay erythropoiesis-stimulating agents (ESAs), impetus exists for studies of novel ESAs that fortify HIF2a's effects, act as EPOR agonists, and/or bolster select downstream EPOR pathways to erythroid cell formation. Such agents could lessen rhEPO dosing, side effects, and/or costs.

Erythropoietin Receptors and IgG Autoantibody Expression on Nucleated Erythrocytes in Some Cases of Immuno-Related Pancytopenia.

BACKGROUND: In recent years, we have detected autoantibodies in bone marrow (BM) hemopoietic cells in some patients with idiopathic cytopenia of undetermined significance (ICUS), termed immunorelated pancytopenia (IRP). However, we know little about the targets of these autoantibodies. METHODS: Twenty-six newly diagnosed IRP patients with IgG autoantibody on nucleated erythrocytes and 20 healthy donors as controls were enrolled in this study. The serum erythropoietin (EPO) level was examined by ELISA. Expression of EPO receptor (EPOR) and IgG autoantibody on the membrane of nucleated erythrocytes were detected by flow cytometry before and after autoantibody stripping. RESULTS: The serum EPO level of the untreated patients was 199.9 ± 106.4 mIU/mL, which was significantly higher than that of normal controls (13.2 ± 8.41 mIU/mL, p < 0.01). EPOR expression on nucleated erythrocytes in the patients was 1.38 ± 0.73%, lower than that of normal controls (2.33 ± 1.73%), but there was no significant difference; EPOR in these patients was inversely correlated with IgG autoantibody on erythrocytes (r = -0.479, p = 0.013). The regression equation was Y = 0.116-0.479X; EPOR expression on the membrane increased significiantly (5.63 ± 4.99%, p < 0.01) after stripping the autoantibodies. After immunosuppressive treatment, median hemoglobin increased from 72 g/L to 98 g/L, and median reticulocytes increased from 1.46% to 3.56%. CONCLUSIONS: IgG autoantibodies might block or competitively inhibit EPOR on nucleated erythrocytes in some cases of ICUS.

Impairment of the Cellular Distribution and Stability of the Erythropoietin Receptor Through the Direct Targeting of Aristolochic Acid.

Aristolochic acid (AA) nephropathy is complicated with early onset and severe anemia. The molecular pathological mechanism of AA-induced anemia remains unclear. The aim of this study was to evaluate the putative pathological roles of the erythropoietin receptor (EPOR) in AA-induced anemia in both AA nephropathy zebrafish and cultured human renal tubular cells (HK2). Immunofluorescence staining experiments revealed that AA colocalizes with the EPOR in zebrafish embryos as well as in the cytoplasm of HK2 cells. After exogenous EPO stimulation, the EPOR was detected in the plasma membrane of HK cells. However, cotreatment with AA and EPO inhibited EPOR signaling and its membrane localization upon EPO stimulation. The results of studies with a protein synthesis inhibitor and a lysosome inhibitor revealed that AA accelerates the lysosomal degradation of EPOR. The molecular docking results suggest that AA may interact with the N-terminus of EPOR. Together with the results of light absorption and in vitro competition assays, we concluded that AA treatment impairs EPOR membrane localization, accelerates its lysosomal degradation, and consequently downregulates EPOR signaling by direct targeting. The results of this study may further detail the pathological mechanism of severe anemia complicated with AA nephropathy.

[The treatment of chemotherapy-induced anemia in lung cancer patients]. / A tüdodaganatos betegek kemoterápia okozta anémiájának kezelése.

Anemia is common among patients with malignant tumors, due to the disease and chemotherapy. Active oncotherapy, combination chemotherapy of lung cancer is accompanied with many side effects which may impair the patient's quality of life and compromise the effectiveness of chemotherapy, the most frequent one of them being chemotherapy-induced anemia. Anemia decreases not only the patient's quality of life, but also worsens the dose-intensity of chemotherapy. One of the potential treatments of chemotherapy-induced anemia is erythropoietin-stimulating agents (ESAs) for the appropriate indications. Several national and international studies have shown that ESA therapy effectively increases hemoglobin level. However, more recently, contradictory results were published on ESA treatment in terms of survival and tumor progression. The reason for this may be that the tumor cells and endothelial cells may as well express erythropoietin receptor, the role of which has not yet been fully elucidated in tumor progression.

Indoxyl sulfate counteracts endothelial effects of erythropoietin through suppression of Akt phosphorylation.

BACKGROUND: Erythropoietin (EPO) is used to treat anemia in patients with chronic kidney disease (CKD). A wide variation in individual response to EPO, however, is often observed, causing EPO resistance. EPO exhibits not only hematopoietic but also extra-hematopoietic functions such as endothelial effects. Indoxyl sulfate, a uremic toxin, is involved in endothelial dysfunction, and consequently, the pathogenesis of CKD-associated cardiovascular disease. The aim of the present study was to determine the effect of indoxyl sulfate on the extra-hematopoietic functions of EPO in human umbilical vein endothelial cells (HUVECs). METHODS AND RESULTS: HUVECs were incubated with or without indoxyl sulfate or an Akt inhibitor, and then stimulated with or without EPO. Indoxyl sulfate suppressed EPO-induced survival/proliferation, anti-apoptosis function, phosphorylation of endothelial nitric oxide synthase, and the expression of thrombospondin-1, an erythroid-stimulating factor, in HUVECs. Although EPO induced phosphorylation of both Akt and extracellular signal-regulated kinases (ERK) in HUVECs, indoxyl sulfate suppressed phosphorylation of Akt but not ERK. An Akt kinase inhibitor or Akt small interfering RNA suppressed all the EPO-induced cellular effects in HUVECs. As a site of action of indoxyl sulfate on EPO signaling, indoxyl sulfate attenuated EPO-induced tyrosine phosphorylation of EPO receptor (EPOR) in HUVECs. CONCLUSIONS: Indoxyl sulfate negatively regulates the EPOR-Akt pathway in endothelial cells, and might contribute to EPO resistance and endothelial dysfunction in patients with CKD.

The protective effects of erythropoietin on rat glomerular podocytes in culture are modulated by extracellular matrix proteins.

BACKGROUND/AIMS: Podocytes are typically cultured on collagen I; however, collagen I is absent from healthy glomerular basement membranes. Erythropoietin (EPO) is thought to protect podocytes in vivo. Here, we studied how various types of extracellular matrix (ECM) proteins and EPO affect podocytes in culture. METHODS: Primary rat podocytes were replated on collagen I, collagen IV, whole ECM extract, laminin, or bare plastic. Cellular adhesion (8 hours after plating), proliferation (5 days, 10 % serum), and resistance to serum deprivation (3 days, 0.5 % serum) were assessed. BrdU incorporation and expression of podocyte-specific markers were employed as measures of cellular proliferation and differentiation, respectively. qPCR was used to verify expression of EPO receptor in cultured podocytes. RESULTS: Cellular adhesion was similar on all ECM proteins and unaffected by EPO. Proliferation was accelerated by laminin and the ECM extract, but the final cell density was similar on all ECM surfaces. Collagen IV supported the serum-deprived cells better than the other ECM proteins. EPO (2-20 ng/ml) improved viability of serum-deprived podocytes on collagen I, collagen IV, and ECM, but not on laminin or bare plastic. The cells expressed mRNA for EPO receptor. CONCLUSION: The physiological ECM proteins are more supportive of primary podocytic cultures compared with collagen I. The protective effects of EPO during serum deprivation are modulated by the cultivation surface.

Erythropoietin, but not asialoerythropoietin or carbamyl-erythropoietin, attenuates monocrotaline-induced pulmonary hypertension in rats.

Erythropoietin (EPO) has long been utilized for the treatment of renal anemia. The erythropoietin receptor (EPOR) is also expressed in the cardiovascular and central nervous systems in addition to an erythroid lineage, to provide an organoprotective role against several types of cellular stress. Pulmonary hypertension (PH) is a poor prognostic disease caused by primary and secondary pulmonary vascular injury. We observed the effects of EPO derivatives on monocrotaline-induced PH in rats on the supposition that EPO may protect small arteries from injury. Asialoerythropoietin (AEPO) lacks sialic acids in the termini of carbohydrate chains that results in rapid clearance from blood. Carbamyl-erythropoietin (CEPO) interacts with EPOR/ßc heterodimers, but not with EPOR homodimers expressed in erythroid cells. Monocrotaline-injected rats were treated with continuous intravenous injection of 2500 ng/kg/day of EPO, AEPO, or CEPO for 21 days, and lung histology, cardiac function, and mRNA expression in the lungs were examined. Wall thickening of small arteries in the lungs and PH were improved by administration of EPO, but not by its non-hematopoietic derivatives, AEPO, or CEPO. Erythropoietin administration increased mRNA expression of the anti-apoptotic molecule, Bcl-xL, and maintained expression of the CD31 antigen. We conclude that lungs may express EPOR homoreceptors, but not heteroreceptors. Adequate serum erythropoietin levels may be essential for pulmonary protective effects.

Tumor necrosis factor α primes cerebral endothelial cells for erythropoietin-induced angiogenesis.

Erythropoietin (EPO) enhances angiogenesis in the ischemic brain. Stroke induces secretion of tumor necrosis factor α (TNF-α). We investigated the effect of TNF-α on EPO-induced in vitro angiogenesis in cerebral endothelial cells. Using a capillary-like tubular formation assay, we found that transient incubation of primary rat cerebral microvascular endothelial cells (RECs) with TNF-α substantially upregulated EPO receptor (EPOR) expression and addition of EPO into TNF-α-treated RECs significantly augmented the capillary-like tube formation. Blockage of TNF receptor 1 (TNFR1) suppressed TNF-α-upregulated EPOR expression and abolished EPO-induced tube formation. Attenuation of endogenous EPOR with small interfering RNA (siRNA) also inhibited EPO-enhanced tube formation. Treatment of RECs with EPO activated nuclear factor-kappa B (NF-κB) and Akt. Incubation of the TNF-α-treated endothelial cells with EPO activated vascular endothelial growth factor (VEGF), VEGF receptor 2 (VEGFR2), angiopoietin 1 (Ang1), and Tie2. Blockage of VEGFR2 and Tie2 resulted in reduction of EPO-augmented tube formation. These data indicate that interaction of TNF-α with TNFR1 sensitizes cerebral endothelial cells for EPO-induced angiogenesis by upregulation of EPOR, which amplifies the effect of EPO on activation of the VEGF/VEGFR2 and Ang1/Tie2 pathways. Our results provide the evidence for crosslink between TNF and EPOR to coordinate the onset of angiogenesis in cerebral endothelial cells.

Erythropoietin protects podocytes from damage by advanced glycation end-products.

BACKGROUND: Podocyte damage and accumulation of advanced glycation end-products (AGEs) are implicated in the development and progression of diabetic nephropathy. We have previously shown that changes in podocyte pathophysiology, such as hypertrophy and reduced migration, are closely linked with the induction of the cell cycle inhibitor p27(Kip1) and a decrease in neuropilin-1 (NRP1) expression. We investigated whether the erythropoietin receptor activators CERA and epoetin-ß may prevent AGE-mediated changes in podocytes. METHODS: Differentiated mouse podocytes in culture were challenged by AGE-modified bovine serum albumin (BSA) or control BSA in the presence or absence of CERA as well as epoetin-ß. Cell cycle analysis and determination of apoptosis markers were performed. p27(Kip1) and NRP1 expression was measured by RT-PCR and Western blots. RESULTS: Differentiated mouse podocytes in culture expressed erythropoietin receptors which were phosphorylated after incubation with CERA or epoetin-ß. CERA or epoetin-ß prevented the p27(Kip1)-dependent cell cycle arrest and cellular hypertrophy induced by AGE-BSA incubation. Furthermore, the p27(Kip1)-dependent AGE-BSA-induced decrease in cell viability and decrease in cell proliferation was ameliorated in the presence of CERA or epoetin-ß. Following erythropoietin treatment, AGE-BSA failed to further reduce NRP1 expression, resulting in improved podocyte migration. CONCLUSION: Treatment with the erythropoietin receptor activators epoetin-ß or CERA protected podocytes from AGE-BSA-mediated damage via an effect on p27(Kip1) and NRP1 expression. Consequently, early treatment with erythropoietin may help to prevent diabetic nephropathy.

[Erythropoietin: pleiotropic actions]. / Aspetti pleiotropici dell'eritropoietina.

The erythropoietin is produced by the kidney and other organs. EPO does not only affect erythroid cells, but also other blood cell lines, such as myeloid cells, lymphocytes and megakaryocytes. This hormone can also enhance phagocytes function of the polymorph nuclear cells and reduces the activation of macrophages, thus modulating the inflammatory process. Hematopoietic and endothelial cells probably have the same cellular origin, and the discovery of erythropoietin receptors also on mesangial and myocardial cells and smooth muscle fibro-cells has prompted the study of the pleiotropic actions of this hormone. Through its receptors, spread out over the body, it carries out many actions which range from the erythrogenesis after hypoxic stimuli to the tissue protection of the heart and the brain after ischemia. Erythropoietin also acts in the endothelial proliferation of new vessels involving the tumor genesis, but it opens new frontiers to the employment of rHuEPO in the Regenerative Medicine.

Darbepoetin-mediated cardioprotection after myocardial infarction involves multiple mechanisms independent of erythropoietin receptor-common beta-chain heteroreceptor.

BACKGROUND AND PURPOSE: Darbepoetin, a long-acting erythropoietin derivative, attenuates cardiomyocyte apoptosis and improves short-term (3 days) cardiac function, but the mechanisms responsible are unknown. We investigated potential mechanisms by which darbepoetin exerts cardioprotection following myocardial infarction in mice and the significance of the erythropoietin receptor (EPOR)-common beta-chain (c-beta-chain) heteroreceptor. EXPERIMENTAL APPROACH: Mice underwent 60 min coronary occlusion followed by treatment with vehicle or a single dose of darbepoetin. Effects on gene expression, apoptosis and neutrophil accumulation in infarcted left ventricle were assessed 24 h later. Cardiac function, effects on vascularization and fibrosis were assessed 28 days later. The significance of EPOR-c-beta-chain heteroreceptor was examined 28 days after infarction using mice deficient in c-beta-chain. KEY RESULTS: Twenty-four hours after darbepoetin, mRNAs encoding haeme oxygenase-1 (HO-1), iNOS and brain natriuretic peptide (BNP) were markedly elevated only in infarcted regions, and the frequency of apoptotic cells attenuated. Inflammation was also attenuated with reductions in neutrophil numbers. Darbepoetin also elevated mRNAs encoding angiogenic factors: placental growth factor, monocyte chemoattractant protein-1 and interleukin-1beta. Twenty-eight days after treatment, CD31+ vessels in the infarct zone doubled and fibrosis reduced. Cardiac haemodynamics were improved. Darbepoetin also improved cardiac haemodynamics in c-beta-chain-deficient mice, increased HO-1 and iNOS expression and vessel numbers and attenuated fibrosis. CONCLUSIONS AND IMPLICATIONS: Darbepoetin stimulates expression of haeme oxygenase, iNOS, BNP and angiogenic factors specifically in infarcted left ventricle that attenuates inflammation, apoptosis and fibrosis; elevate vessel numbers; and improve cardiac function. The EPOR-c-beta-chain heteroreceptor is not essential for these effects.

Erythropoietin activates mitochondrial biogenesis and couples red cell mass to mitochondrial mass in the heart.

RATIONALE: Erythropoietin (EPO) is often administered to cardiac patients with anemia, particularly from chronic kidney disease, and stimulation of erythropoiesis may stabilize left ventricular and renal function by recruiting protective effects beyond the correction of anemia. OBJECTIVE: We examined the hypothesis that EPO receptor (EpoR) ligand-binding, which activates endothelial NO synthase (eNOS), regulates the prosurvival program of mitochondrial biogenesis in the heart. METHODS AND RESULTS: We investigated the effects of EPO on mitochondrial biogenesis over 14 days in healthy mice. Mice expressing a mitochondrial green fluorescent protein reporter construct demonstrated sharp increases in myocardial mitochondrial density after 3 days of EPO administration that peaked at 7 days and surpassed hepatic or renal effects and anteceded significant increases in blood hemoglobin content. Quantitatively, in wild-type mice, complex II activity, state 3 respiration, and mtDNA copy number increased significantly; also, resting energy expenditure and natural running speed improved, with no evidence of an increase in left ventricular mass index. Mechanistically, EPO activated cardiac mitochondrial biogenesis by enhancement of nuclear respiratory factor-1, PGC-1alpha (peroxisome proliferator-activated receptor gamma coactivator 1alpha), and mitochondrial transcription factor-A gene expression in wild-type but not in eNOS(-/-) or protein kinase B (Akt1)(-/-) mice. EpoR was required, because EpoR silencing in cardiomyocytes blocked EPO-mediated nuclear translocation of nuclear respiratory factor-1. CONCLUSIONS: These findings support a new physiological and protective role for EPO, acting through its cell surface receptor and eNOS-Akt1 signal transduction, in matching cardiac mitochondrial mass to the convective O(2) transport capacity as erythrocyte mass expands.

Lenalidomide for treatment of myelodysplastic syndromes: current status and future directions.

Lenalidomide was approved by the US Food and Drug Administration (FDA) for treatment of transfusion-dependent lower-risk myelodysplastic syndrome patients with deletion (del) (5q) alone or with additional karyotype abnormalities. The approval was based on high rates of prolonged transfusion independence and complete cytogenetic response in this subset. In lower-risk non-del(5q) patients, meaningful erythroid responses also were reported with a low frequency of cytogenetic improvement, although inferior to that observed in the del(5q) patients. There is now a better understanding of the mechanism of the karyotype-dependent drug action, explaining the disparate response rates and frequency of myelosuppression. In del(5q) patients, lenalidomide suppresses the clone by inhibiting the nuclear sequestration of the haplodeficient cell cycle regulatory protein cdc25c, thereby promoting selective G2 arrest and apoptosis. In non-del(5q) patients, lenalidomide enhances erythropoietin receptor signaling. Future directions include use of biologic and molecular markers as predictive tools to select patients and use of combination strategies to overcome resistance to lenalidomide in del(5q) patients or enhance erythropoiesis in non-del 5 patients.