Development and Characterization of Darbepoetin

Studies on human erythropoietin (EPO) demonstrated that there is a direct relationship between the sialic acid-containing carbohydrate content of the molecule and its serum half-life and in vivo biological activity, but an inverse relationship with its receptor-binding affinity. These observations led to the hypothesis that increasing the carbohydrate content, beyond that found naturally, would lead to a molecule with enhanced biological activity. Hyperglycosylated recombinant human EPO (rHuEPO) analogs were developed to test this hypothesis. Darbepoetin alfa (Aranesp), which was engineered to contain five N-linked carbohydrate chains (two more than rHuEPO), has been evaluated in preclinical animal studies. Due to its increased sialic acid-containing carbohydrate content, darbepoetin alfa is biochemically distinct from rHuEPO, having an increased molecular weight and greater negative charge. Compared with rHuEPO, it has an approximate threefold longer serum half-life, greater in vivo potency, and can be administered less frequently to obtain the same biological response. Darbepoetin alfa is currently being evaluated in human clinical trials for treatment of anemia and reduction in its incidence. [ONCOLOGY 16(Suppl 11):13-22, 2002] Erythropoietin (EPO) is a glycoprotein hormone that is the primary regulator of erythropoiesis, maintaining the body’s red blood cell mass at an optimum level. [1,2] In response to a decrease in tissue oxygenation, EPO synthesis increases in the kidney. The secreted hormone binds to specific receptors on the surface of red blood cell precursors in the bone marrow, leading to their survival, proliferation and differentiation, and ultimately to an increase in hematocrit. [3,4] Since its introduction more than a decade ago, recombinant human EPO (rHuEPO) has become the standard of care in treating the anemia associated with chronic renal failure (CRF). It is highly effective at correcting the anemia, restoring energy levels, and increasing patient well-being and quality of life. [5-8] It has also been approved for the treatment of anemia associated with cancer, HIV infection, and use in the surgical setting to decrease the need for allogeneic blood transfusions. For all indications, it has proven to be remarkably well tolerated and highly efficacious. [9-11] The recommended and usual therapy with rHuEPO is two to three times per week by subcutaneous or intravenous injection. For CRF patients, the duration of therapy is for the life of the patient, or until a successful kidney transplant restores kidney function, including the production of the natural hormone. For cancer patients, rHuEPO therapy is indicated for as long as the anemia persists, generally through the entire course of chemotherapy. It can be a hardship to administer rHuEPO two to three times per week, particularly for those patients who do not otherwise need to be seen in the clinic this frequently. In these cases, the patient needs to make a special trip to the clinic for his or her rHuEPO therapy. As is true for all growth factors, reduction in dose frequency results in a significant loss in efficiency. That is, the total weekly dose required when rHuEPO is administered one time per week is greater than when it is administered as two to three divided doses. It was anticipated that this clinical need could be addressed by creating a molecule with enhanced in vivo bioactivity to allow for less frequent dosing of patients. To create a molecule with enhanced activity, research was initially directed towards elucidating those factors and structural features that control the in vivo activity of EPO. [12] This research led to the discovery and development of darbepoetin alfa, a novel erythropoiesis stimulating protein (Aranesp), that can be administered less frequently than epoetin. [13]