Assessment of the pre-clinical immunogenicity of a new VEGF receptor Fc-fusion protein FP3 with ELISA and BIACORE.

Abstract

A new VEGF receptor fusion protein FP3 was shown to have promising antitumor potency better than Bevacizumab. Characterization of its immune response is essential to the safe and effective administration in clinical trials. In this study, both BIACORE and ELISA assays were employed to assess pre-clinical immunogenicity of FP3 in monkeys. Serum samples from 20 rhesus monkeys were analyzed for the generation of anti-FP3 antibody after intravenous administration of three doses of FP3 (n = 6 per group) or buffer control (n = 2). Sera samples were obtained at 2, 4, 6, 8, 10 weeks after the first administration. It showed BIACORE presented linear correlation with the dilution of anti-FP3 antibody and the results of ELISA. Two weeks after the initial FP3 injection, anti-FP3 antibody was detected in about 20% FP3-treated monkeys. The ratio of positive samples and the titer of antibody increased along with the FP3-treatment time. Six weeks following FP3 injection almost all the samples were anti-FP3 antibody positive. Moreover, the titer of anti-FP3 antibody but not the ratio of positive samples was also enhanced when the dose of FP3 was elevated. Furthermore, the immunoglobulin types and subclasses of anti-FP3 antibody serum components were mainly identified as IgG1 and IgG4, not IgM. Serum antibodies are characterized that they could not block FP3 binding to VEGF and were non-neutralizing. Our data implied that proteins with full human sequences may also have the potential to induce immune response in rhesus monkeys, and BIACORE could be an effective approach to detect the immunogenicity of protein therapeutics in clinic.