In this study, canine IFN? was fused by a flexible linker with canine serum albumin to construct the fusion protein IFN?-CSA for the purpose to design a long-acting canine IFN?. The fusion protein was successfully expressed in baculovirus-infected Sf9 insect cells and was purified by salting-out and ion exchange chromatography. The IFN?-CSA fusion possessed potent anti-viral assay against vesicular stomatitis virus in cultured cells. IFN?-CSA was also stable at 37 °C up to 72 h compared with 8 h for IFN? alone. In vivo pharmacokinetics demonstrated a significantly longer half-life for IFN?-CSA (15.42 h) than for canine reIFN? (1.51 h) in KM mice. These results indicate that IFN?-CSA expression in the baculovirus system was successful and provide a promising long-acting cytokine for veterinary clinical applications.
Interferon (IFN)-? plays an important role in antiviral, anti-proliferative, immunomodulatory and pro-inflammatory activities. However, the short therapeutic half-life of IFN-? lessens its efficacy. Albumin fusion strategy is one of the most effective ways to improve the pharmacokinetic properties of cytokines. In this study, N- and C-terminal canine albumin fusions with canine IFN-? were expressed in the baculovirus expression system. The fusion proteins stimulated Stat1 phosphorylation at levels similar to that of the recombinant IFN. The antiviral, anti-proliferative and promote apoptosis activity of CSA-IFN-? was lower than IFN-?-CSA and both were less than that of recombinant IFN-?. In vivo pharmacokinetics demonstrated a significantly longer half-life for CSA-IFN-? (21.73 h) than for IFN-?-CSA (6.51 h) and canine reIFN-? (2.22 h) in Wistar rats. CSA-IFN-? was also more effective than IFN-?-CSA and canine reIFN-? at inhibiting growth of canine renal malignant histiocytosis in nude mice. Our results indicated that a canine serum albumin fusion at the N-terminus of IFN-? prolongs its half-life and improves its in vivo antitumor activity.
