Figure 2.

In vitro mode of action of B7-H6/CD3 ITE. A, Schematic graphic of B7-H6/CD3 ITE. B and C, Binding to HCT-15 (B) and T cells (C). Each datapoint represents a single measurement. D-F, Correlation of B7-H6 cell surface density and B7-H6/CD3 ITE-induced lysis of 10 colorectal (D), 10 gastric (E), and 14 pancreatic (F) cancer cell lines incubated with T cells in a ratio of 1:10 in the presence of 1.5 μg/mL B7-H6/CD3 ITE for 72 hours. G–M, Human T cells and B7-H6–positive HCT-15 cells or B7-H6–negative CHO-3E7 cells were cocultivated in the presence of increasing concentrations of B7-H6/CD3 ITE for 72 hours. G, B7-H6 dependency of B7-H6/CD3 ITE-induced cell lysis [effector to target cell ratio (E:T) 10:1]. H, B7-H6/CD3-induced lysis of HCT-15 cells by CD4⁺ and CD8⁺ T cells (E:T 10:1). I, E:T ratio dependency of B7-H6/CD3 ITE-induced lysis of HCT-15 cells. J, B7-H6 dependency of B7-H6/CD3 ITE-induced upregulation of CD25 and CD69 on CD4⁺ and CD8⁺ T cells (E:T 10:1). K, B7-H6 dependency of B7-H6/CD3 ITE-induced upregulation of extracellular CD107a and intracellular granzyme B (GrzB) in CD4⁺ and CD8⁺ T cells (E:T 10:1). L, B7-H6 dependency of B7-H6/CD3 ITE-induced secretion of IL2 and IFNγ by T cells (E:T 10:1). M, B7-H6 dependency of B7-H6/CD3 ITE-induced proliferation of T cells (E:T 10:1) after 6 days of coculture. Each datapoint represents a single measurement. For G, H, I, and L, each datapoint represents the mean of duplicates, error bars represent the SD. For J and K, each datapoint represents data from duplicates which were pooled for the flow cytometry analysis.