KIR-CARs/Dap12 trigger robust antigen-specific cytotoxicity in vitro.

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KIR-CARs/Dap12 trigger robust antigen-specific cytotoxicity in vitro. KIR-CARs/Dap12 trigger robust antigen-specific cytotoxicity in vitro. A, schematic diagram of a mesothelin-specific KIR-CAR. B, schematic diagram of the lentiviral vector used for codelivery of a KIR-CAR and DAP12. C, primary human T cells were stimulated with anti-CD3/anti-CD28 T-cell activator beads (Dynabeads CD3/CD28 CTS; Life Technologies). After 24 hours of stimulation, the T cells were transduced with a lentiviral vector encoding the indicated CAR or mock-transduced (Mock). Cells were expanded for 9 days and analyzed by flow cytometry for the expression of the indicated CAR using biotinylated goat-anti-mouse F(ab)2 followed by streptavidin-PE. Results are representative of at least 3 independent experiments. D, antigen-specific cytotoxic activity of T cells generated as described in C was assessed by a 4-hour 51Cr-release assay as described in Supplementary Materials and Methods using K562 cells (Kwt) or K562 cells engineered to express human mesothelin (Kmeso). Enxiu Wang et al. Cancer Immunol Res 2015;3:815-826 ©2015 by American Association for Cancer Research