Hyper N-glycosylated human interferon alpha2b produced in HEK293 cells presents higher antitumor activity than the CHO-K1 derived protein

GUGLIOTTA, A.; CEAGLIO, N.; RAUD, B.; FORNO, A. G.; MAURO, L; KRATJE, R.; OGGERO, M.

Lausanne

Congreso; 25th Meeting of the European Society for Animal Cell Technology (ESACT); 2017

European Society for Animal Cell Technology (ESACT)

Background and noveltyGlycosylation constitutes an important attribute of glycoproteins since it defines many of their properties. In this context, CHO and HEK cells are interesting hosts for their production as biotherapeutics. The potentiality of using the human cell line over the rodent model to produce a hyper N-glycosylated protein is herein analyzed.Experimental approachA long-lasting N-glycoengineered human interferon‑α2b (IFN4N) was produced in CHO and HEK cell lines and purified by immunoaffinity chromatography. Both molecules were compared in terms of glycosidic profile, pharmacokinetics and in vitro and in vivo biological activity.Results and discussionIFN4NCHO presented higher average molecular mass and more acidic isoforms compared to IFN4NHEK. Accordingly, a significantly 2-fold higher sialic acid content was found in IFN4NCHO compared to IFN4NHEK. These results were in agreement with monosaccharides quantification and WAX chromatography, which supports the existence of highly sialylated and ramified structures for IFN4NCHO glycans, in contrast with smaller and truncated structures from IFN4NHEK. Mass spectrometry analysis revealed that FA2G2S2, FA3G3S3, FA4G4S4 and FA4G4Lac1S4 were predominant in IFN4NCHO whereas FA2B/FA3, FA2G1 and FA2BG1 were the main IFN4NHEK-bearing glycans. Unexpectedly, those remarkable differences had not a considerable impact on their pharmacokinetic properties, showing quite similar elimination profiles. Also, despite the in vitro antiviral specific biological activity of both proteins was the same, IFN4NHEK was more efficient as an antiproliferative agent. Further experiments with nude mice implanted with PC-3 cells confirmed the highest antitumor efficiency of IFN4NHEK. Our results show the importance of an appropriate host selection to set up a bioprocess and potentiate the use of HEK cells for the production of a new hyper N-glycosylated protein-based pharmaceutical."