Fig. 6

Costimulated NKG2D CAR T cells inhibit tumor growth of TNBC in vivo. a NKG2D ligand expression on TNBC cell line MDA-MB-231 detected by NKG2D-Fc chimeric protein. b Growth curve of MDA-MB-231 tumor (n = 5) treated with the control PBS, UNT T cells (3 × 10⁷), or NKG2D CAR T cells (3 × 10⁷, ~ 30% CAR+) by intravenous injection 40 and 45 days post tumor inoculation. At the end of the experiment, the tumors treated with GFP-NKG2D-BBz or GFP-NKG2D-27z CAR T cells were significantly smaller than those in the control group and GFP-NKG2D-z group (P < 0.001). c Bioluminescence images was applied to monitor and quantify MDA-MB-231 fLuc(+) tumor growth in NSG mice immediately before and 3 weeks after first CAR-T cell injection. d CD4+ and CD8+ GFP-NKG2D-BBz and GFP-NKG2D-27z CAR T cells were initially present at low numbers in peripheral circulation, suggesting NKG2DL-specific CAR T cell migration to specific tumor locales. Mean cell concentration (cells/ul) ± SEM for all evaluable mice in each treatment group is shown (n = 5). e NKG2D-z CAR, but not costimulated NKG2D CAR, was poorly expressed on the surface of transduced (GFP+) T cells, suggesting CAR downregulation. f, g CD27 and 4-1BB signaling enhances the survival of circulating human CD4+ and CD8 +T cells in vivo 3 weeks after first dose of T cell infusion(P < 0.01) and costimulated NKG2D CAR expression on the T cell surface is stable and increased in vivo