Figure 6. EGFR806-CAR T cells are selectively activated by tumor-specific EGFR expression in a human iPS cell teratoma model.

(A) Teratoma-only model: mice were injected subcutaneously with human iPS (hiPS) cells in the left leg muscle, and EGFR806-CAR T cells were injected after teratoma has established. Immunolabeling of the teratoma (right 2 panels) shows minimal infiltration of CD3⁺ cells (green) into EGFR⁺ (white) regions of teratoma. (B) Teratoma-glioma model: mice were injected with hiPS cells in the left leg. After establishment of a palpable teratoma, U87 glioma cells expressing CD19t (U87-CD19t) were injected in the right flank. CAR T cells were infused 10 days after glioma cell injection. Representative images show immunolabeling demonstrating similar degrees of infiltration of CD3⁺ CAR T cells (green) into EGFR⁺ teratoma (left 2 columns) and EGFR⁺ U87-CD19t glioma (right column) after treatment with EGFR806 CAR (top), Erbitux CAR (middle) and CD19 CAR (bottom) T cells, with differing degrees of activation as indicated by Granzyme B (GrzB) labeling in red. (C) Left: infiltration of CAR T cells into teratoma is much lower in animals that did not receive concurrent U87-CD19t glioma grafts. Right: GrzB⁺ CAR T cells in EGFR⁺ teratoma as a percentage of total CD3⁺ teratoma resident T cells. Bars represent average cell number in 20 40× images from different EGFR⁺ tumor regions. Error bars, SEM. (D) Left: infiltration of CAR T cells into U87-CD19t glioma grafts on the other side of teratoma-bearing mice. Right: bars represent percentage GrzB⁺ CAR T cells of total CD3⁺ T cells in EGFR⁺ U87-CD19t glioma. Bars represent average cell number in 20 40× images from different glial tumor regions. Error bars, SEM. ^* P < 0.05; ^** P < 0.01; ^*** P < 0.001; ^**** P < 0.0001. Scale bar, 20 μm. All images were acquired at 40× magnification.