Fig. 7
PF-06747143 induces CLL-B cell killing by ADCC and CDC. a CDC assay was performed by treating CLL-B cells (1 × 106/mL) with PF-06747143, in presence of complete or heat-inactivated 5% human serum. The cells were incubated for 4 h and the cytotoxicity was determined using flow cytometry with CD19/CD5/Annexin V staining. b The ADCC assay in patient B-CLL cells was performed using 1:1 ratio of the target/effector cell (T/E) and incubated for 6 h at 37 °C. The IgG1 control antibody, PF-06747143, and rituximab were tested in a 1:3 titration curve, ADCC activity was determined using Bio-Glo™ luciferase assay, and the luminescence results are expressed in relative light units (RLU). The samples were analyzed in duplicates with the mean and SD shown for each group. The data was analyzed using Prism 4 GraphPad software. c ADCC activity was evaluated in JVM-13 tumor cells by incubating the cells with PF-06747143, m15-IgG1, m15-IgG4, or respective negative control antibodies for 4 h, in the presence of NK92 158V effector killer cells (effector/target cell ratio 10:1). Cell lysis was measured by ToxiLight bioluminescent cytotoxicity assay. Experiments were performed in quadruplicates with the mean ± SEM shown for each group