Figure 1.

Jurkat reporter cells (Promega) stably expressing Fc γ receptor (FcγR) I, FcγRIIA or FcγRIIIA, respectively, were incubated with increasing concentrations of anti-CD2 antibody. Data normalized to luminescence induced by the highest concentration of siplizumab (mean ± SD; n=3). Reporter cells bind the Fc-fragment of a target-bound IgG antibody with their FcγR which induces expression of a luciferase reporter gene resulting in a luminescence signal upon addition of assay substrate (Promega). Cells were incubated with increasing doses of siplizumab, deglycosylated (DG) siplizumab, Fc-silent (FcS) anti-CD2 IgG1, FcS anti-CD2 IgG2 and FcS anti-CD2 IgG4, respectively. (A) Cell-based FcγRI signaling assay. (B) Cell-based FcγRIIA signaling assay. (C) Cell-based FcγRIIIA signaling assay.