Ajay K. Singh, MB, MRCP

Disclosures

January 07, 2008

In This Article

Introduction

The use of erythropoietin (Epo) in treating anemia of chronic kidney disease (CKD) has transformed the lives of millions of individuals.[1] In the pre-epoetin era, patients had to live with the symptoms of severe anemia, multiple transfusions, and frequent hospitalization. Recombinant erythropoietin (rhEpo) was approved in 1989 for the treatment of CKD anemia. Widespread use of Epo in the management of CKD anemia ensued. Over the past 2 decades, the market for Epo has dramatically increased. It is now estimated that $22 billion worth of Epo has been sold.[2] This vast market opportunity has not gone without notice in the pharmaceutical industry, and many Epo analogues and derivatives have entered the marketplace. These newer Epos are now grouped under the term erythropoietin-stimulating agents or ESAs. In addition, follow-on epoetins (FOEs), similar to the innovator epoetin, are now commonplace, particularly in emerging countries. The purpose of this mini-review is to critically evaluate the impact of both ESAs and FOEs in the management of CKD anemia.

 

Erythropoietin Molecular Physiology

Epo is an acidic glycoprotein with a molecular weight of 30.4 kDa ( Table ).[3,4]

The carbohydrate part of the molecule (approximately 40%) is composed of 3 N-linked and 1 O-linked glycan moieties. N-glycans are key to the biological activity of Epo; the addition of N-glycans into rhEpo increases the carbohydrate content and results in prolonged survival of the molecule. From a structural perspective, Epo has a basic structure like other cytokines (eg, the interleukins). It is composed of 4 amphipathic {alpha}-helical bundles. There are 2 domains located on Helix C and Helix D that are necessary for binding to the Epo receptor (EpoR). Following binding to the EpoR, a JAK2-mediated signal transduction cascade ensues (Figure 1). Epo's binding affinity is also critical to its action. Epoetin-alfa has a high binding affinity, whereas newer derivatives such as continuous erythropoietin receptor activator (CERA) have much lower affinities that are thought to contribute to the longer half-life of these molecules.

Figure 1.

Epo signal transduction.

The human Epo gene is located on the long arm of chromosome 7 (q11-q22).[5] The Epo gene contains 5 exons and 4 introns. Tissue-dependent expression of the Epo gene is dependent on 5' promoter sequences.[6] Several transcription factors bind to the promoter and regulate erythropoietin gene expression, including factors that stimulate (GATA-4) or inhibit (GATA-2 and NF-kappaB) Epo gene expression. The induction of Epo gene expression under hypoxic conditions is dependent on a 3' enhancer element.[7,8] Specific hypoxia-inducible transcription factors (HIF alpha/beta) bind hypoxia response elements within this enhancer sequence. Both GATA and HIF have emerged as therapeutic targets for modulating Epo gene expression and activity.

 

ESA Derivatives That Prolong Epo Action

In the years following the launch of rhEpo, it became apparent that N-linked glycosylation increased the half-life of the Epo molecule. In 2001, darbepoetin, a longer-acting derivative of epoetin-alfa, was approved. In 2007, CERA, a derivative of epoetin-beta, received regulatory approval. Both darbepoetin-alfa and CERA have a prolonged half life compared with epoetin-alfa or epoetin-beta (Figure 2); these agents have been extensively tested in phase 2 and 3 programs and allow less frequent dosing of Epo in the management of CKD anemia.

Figure 2.

Comparison of mean half-lives.

Darbepoetin-alfa has a half-life approximately 3 times that of epoetin-alfa (terminal half-life: 25 hours vs 8 hours).[9] It has 5 N-linked glycosylation sites (2 more than normally present in endogenous Epo) that increase its molecular weight from 30.4 to 37.1 kDa. N-glycosylation increases the carbohydrate content of darbepoetin from 40% to 51%. In patients with chronic kidney disease, darbepoetin is generally administered once every 2 weeks, but even monthly administration has been demonstrated to be effective.[10,11] In hemodialysis patients, the drug is usually given intravenously; but in nondialysis CKD patients, subcutaneous administration, at equivalent dosing, is more convenient and equally effective.

CERA is a pegylated form of epoetin-beta. CERA is a 60-kDa molecule, approximately twice the molecular weight of epoetin-alfa.[12,13] A large methoxy-polyethylene glycol polymer is incorporated into the epoetin molecule, essentially doubling its molecular weight compared with epoetin. This modification markedly prolongs CERA's half-life in humans to about 135 hours. In addition, CERA binds to EpoR more slowly than Epo. These 2 attributes result in CERA triggering the JAK2 Epo signal transduction cascade without being internalized. Consequently, CERA has more sustained biologic activity. CERA has been tested extensively and can be used less frequently than epoetin-alfa or darbepoetin-alfa -- it can be administered either once every other week or once monthly.[14,15,16]

Other ESAs are in the drug development pipeline but have not been tested in any meaningful manner in clinical trials. These emerging agents take advantage of technology that structurally modifies the rhEpo molecule to prolong its half-life or change its binding affinity to the EpoR. One class of agents are the recombinant dimeric Epos.[17,18] Here, rhEpo is linked to form dimers and trimers. This results in both an increased half-life and bioactivity. Another agent is termed synthetic erythropoiesis protein (SEP).[19] This is a synthetic 50.8-kDa macromolecule (a 166-amino-acid polypeptide) with a sequence similar to but not identical to that of native Epo. Negatively charged noncarbohydrate branched polymers have been attached to this protein at 2 sites by chemical ligation. SEP has a half-life approximately 2.5-fold longer than that of epoetin-alfa. Various strategies to fuse Epo to other proteins has also been pursued. One approach has been to add peptides at the carboxy-terminal end of the molecule.[20] Other approaches include fusing epoetin-alfa with the Fc region of human IgG,[21] fusing erythropoietin with GM-CSF[22,23] (GM-CSF stimulates erythroid progenitor cells in the bone marrow or using Epo mimetic fusion proteins that are structurally distinct from epoetin but activate EpoR. Unfortunately, this compound was also found to be a potent stimulator of anti-Epo antibodies.[24]

Small-molecule Epo mimetic peptides (EMPs) provide a novel paradigm to stimulate Epo. EMPs are peptide sequences that bind and activate EpoR. For example, Wrighton and colleagues[25,26] have identified a 20-amino-acid peptide (GGTYSCHFGPLTWVCKPQGG) that binds EpoR and has both in vivo and in vitro biologic activity. An example of an EMP that has entered clinical trials with much promise is Hematide (Affymax Inc., Palo Alto, California).[27] This is a pegylated dimeric EMP that has no sequence homology to Epo but binds and activates EpoR. PEGylation enhances the stability and prolongs the half-life of the molecule (t1/2 of up to 60 hours). Hematide is now being evaluated in phase 2 studies both in renal patients and in patients with cancer.[28,29] Other EMPs are also in the pipeline.

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