[Expression of rhEPO-L-Fc fusion protein and analysis of its bioactivity and pharmacokinetics].
To prolong serum half-life of human Erythropoietin for better efficacy, a new form of recombinant human erythropoietin (rhEpo-L-Fc) was generated by fusion of a full length human erythropoietin gene and the Fc fragment of human IgG1 with flexible linker sequence. The fusion gene rhEPO-L-Fc was constructed by PCR, then inserted into expression vector pOptiVEC-TOPO, and expressed in Chinese Hamster Ovary cells deficient in the DHFR enzyme(CHO-dhfr-). The chimeric protein was purified by Protein A affinity chromatography, showed expected molecular weight and demonstrated a similar bioactivity compared to that of the native recombinant human erythropoietin (rhEPO) in an EPO-dependent cell-based assay. In vivo pharmacokinetic studies showed that the rhEPO-L-Fc had an elimination half-life of 27 h. In vivo efficacy studies showed that a single dose administration of rhEPO-L-Fc in rats increased the reticulocyte number in the peripheral blood significantly. These results demonstrated that the new engineered rhEPO-L-Fc may become alternative therapeutic approach to extend the half-time of rhEPO to treat anemia.
Zhu
Qiang
Q
Department of Biochemistry and Molecular Biology, College of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Huang
Zhihua
Z
Huang
Yuliang
Y
Qin
Yang
Y
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004921:Erythropoietin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000493:pharmacokinetics*; D005260:Female; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000096:biosynthesis* / Q000235:genetics; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics*; D011994:Recombinant Proteins
D007141:Immunoglobulin Fc Fragments; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D004921:Erythropoietin