Fig. 4

CD16-CAR T cell activation and cytoxicity in complex co-culture systems. a Schematic overview of constructs used in this figure; the anti-MCSP antibody (LC007 or glycoengineered LC007GE), CD16-CAR with VH, FL, VL variants and Vdel. b In total, 20 × 10⁴ CD16 VH-CAR-, CD16 VL-CAR-, CD16 FL-CAR-, CD16-del-transduced T cells were co-cultured for 48 h with 2 × 10⁴ MCSP⁺ A375 melanoma cells in presence of either 10 µg/ml of the anti-MCSP antibody LC007, 10 µg/ml of the anti-MCSP glycoengineered antibody LC007GE or without antibody. Each condition was additionally tested in the presence or absence of 10 mg/ml or 1 mg/ml polyclonal immunoglobulin or 2 × 10⁶ PBMC. IFN-γ production was measured by ELISA. c IN total, 10⁵ CD16 VH-CAR-, CD16 VL-CAR-, CD16 FL-CAR-, CD16-del-transduced T cells were added after overnight incubation to 10⁴ MCSP⁺ A375 melanoma cells, in presence of either 10 µg/ml of the anti-MCSP antibody LC007, 10 µg/ml anti-MCSP glycoengineered antibody LC007GE or without antibody. Each condition was additionally tested in the presence or absence of 10 mg/ml or 1 mg/ml polyclonal human immunoglobulins (IVIgs) or 10⁶ PBMC. The time point when T cells and antibodies were added to the system are indicated by arrows. Cytotoxicity was measured in real-time, as a mean of cell viability by xCELLigence technology. All graphs show mean values of at least three technical replicate and each experiment shown is a representative figure of at least three independent experiments. For figure B a two-sided unpaired Student's t test was used and for C a two-way ANOVA with Bonferroni-correction was used to determine the p-values. A p-value < 0.05 was considered statistically significant