The in vitro antitumor effects of CD20-Flex BiFP against lymphoma. Raji (A) and Daudi (B) cells were incubated with increasing concentrations of CD20 antibodies and Flex-Ig fusion protein in the presence of human complement at 37°C for 4 h. CDC activity was calculated by a standard lactate dehydrogenase assay as described in “Materials and methods.” These graphs are representative of at least 3 experiments, each showing similar results. (C) ADCC activity against Daudi cells using human PBMCs as effector cells at an effector-to-target cell (E:T) ratio of 25:1. The ADCC activity of CD20-Flex BiFP at varying concentrations was measured using a standard lactate dehydrogenase assay as described in “Materials and methods.” Data are expressed as means ± SD (n = 3). (D) Inhibition of CD20 antibody-induced cell death by ZVAD. Before Raji cells were incubated with 10 μg/mL rituximab or CD20-Flex BiFP in the presence of cross-linker (x-link, goat anti-human κ F(ab′)2 fragment, 20 μg/mL), caspase inhibitor ZVAD was added over a range of different concentrations for 2 hours. Shown are means and SD of at least 3 experiments.
