Formulation Optimization Of Recombinant Human Erythropoietin Sustained-release Microspheres

The present study is aimed to examine effects of microspherespreparation conditions on improving Erythropoietin (EPO) stability and invivo pharmacodynamic using SEC-HPLC and BALB/C mice. The reasonwhy sustained-release of protein drugs hasn’t been successfully applied aftermore than30years of research is mainly due to the fragile high structure ofthese macromolecules and adverse immune response induced by theirstructural changes. Water-oil and water-air interfaces in the process ofpreparation are recognized as the reasons causing protein aggregation. Toavoid such interfaces a unique approach involving pre-loading the proteininto solvent-resistant polysaccharide glassy particles was proposed by ourlaboratory. We applied the technology for EPO microsphere preparation andoptimization in this project.EPO is a protein hormone consisting of165amino acids and30400Dain molecular weight which is well used for treatment of patients with anemiadue to kidney diseases, cancer, chemotherapy, or HIV infection. EPO tops theglobal sales of recombinant protein drugs and easily denatured in theformulation process of sustained-release microspheres. Sustained release ofEPO not only has substantial clinical needs, but also can fully prove thefeasibility of the above sustained-release technology.In the present study, in vitro release factors of EPO microspheres were studied using UT-7cell proliferation method, and the formulationoptimization was studied in accordance with BALB/C mice in vivo efficacy.The optimized EPO microspheres showed considerable efficacy whilesignificantly reduced administration frequency compared to EPO solutionswhich could enhanced the patients’ compliance completely. EPO recoveredfrom microspheres by S/O/W and S/O/hO showed few or no aggregates bySEC-HPLC assay. In vitro release profiles showed less burst release andincomplete release.The SEC-HPLC, in vivo and in vitro release assays provided consistentevidences that microencapsulation by m-S/O/W and m-S/O/hO method maygreatly reduce EPO aggregates and preserve its activity. The completion ofthis study will not only solve sustained-release problems of EPO, the world’slargest prescription drug market, but also establish a new sustained-releasegeneric approach for a large number of similar recombinant protein drugs.