Development and biological activity of long-acting recombinant human interferon-alpha 2b

Background The type I human interferon (IFN) family consists of a group of cytokines with a multiplicity of biological activities, including antiviral, antitumor, and immunomodulatory effects. However, because the half-life of IFN is short, its clinical application is limited. Increasing the yield and biological activity of IFN while extending its half-life is currently the focus of IFN research. Results Two novel long-acting recombinant human IFN-alpha 2b (rhIFN-alpha 2b) proteins were designed in which the carboxyl-terminal peptide (CTP) of the human chorionic gonadotropin beta su bunit and N-linked glycosylation sequences were linked to rhIFN-alpha 2b. They were designated IFN-1CTPON (fused at the C-terminus of rhIFN-alpha 2b) and IFN-2CTPON (fused at both the C-terminus and N-terminus of rhIFN-alpha 2b). Monoclonal CHO cell strains stably and efficiently expressing the IFNs were successfully selected with methotrexate (MTX), and the highest expression levels were 1468 mg/l and 1196 mg/l for IFN-1CTPON and IFN-2CTPON, respectively. The proteins were purified with affinity chromatography and molecular sieve chromatography. IFN-1CTPON and IFN-2CTPON showed antiviral and antiproliferative activities in vitro. Notably, the half-life of IFN-1CTPON and IFN-2CTPON in vivo were three-fold and two-fold longer than that of commercially available rhIFN-alpha 2b. Conclusions CHO cell strains stably expressing long-acting rhIFN-alpha 2b were screened. The purified IFN-CTPON protein has biological activity and an extended half-life, and therefore potential applications.