Figure 5. Tg-Pmel T cells effectively infiltrate and persist in vivo in the tumor if adoptively transferred in conjunction with anti-VEGFR2 CAR transduced T cells.

C57BL/6 mice bearing B16 melanoma tumor were sublethally irradiated with 5 Gy TBI and treated with 10⁶ Thy1.1⁺ Wt T cells or 10⁶ Thy1.1⁺ Tg-Pmel T cells mixed with 5 × 10⁶ Ly5.1⁺ syngeneic open repertoire Wt T cells transduced with a retroviral vector expressing the control CAR (SP6 CAR) or an anti-VEGFR2 CAR (DC101 CAR). Animals concurrently received a single dose of 2 × 10⁷ pfu vaccinia virus expressing hgp100 and twice daily rhIL-2 administration for 3 days. Control group received no treatment. Tumors and spleens of 3 mice from each group were excised at different time points post therapy and single cell suspensions were made as described in materials and methods. Cell preparations were stained with FITC-labeled anti-rat Thy1.1 and PE-labeled mouse anti-mouse Ly5.1 antibodies and analyzed by flow cytometry. Representative flow cytometry data from single cell preparations of spleen and tumor tissues from one mouse in each group obtained on day 3, 6, and 9 post T cell treatment indicating the percentage Thy1.1⁺ and Ly5.1⁺ cells gated in the total viable cell population are shown.