Figure 6.

Impact of multiple coinhibitory receptor blockade during engineered T cell therapy. (A) T cell therapy protocol ±αPD-L1, αTIM-3, αLag3 (mAbs) or isotype controls in KPC mice. (B) Number of TCR_Msln cells normalized to tumor gram at 8 and 28 days postinfusion. Data are mean±SEM. (C) Representative flow cytometry plots of intracellular cytokine staining gated on TCR_Msln cells. (D) Proportion of TCR_Msln cells producing cytokines following ±4 hour ex vivo restimulation with Msln_406-414 peptide. Data are mean±SEM and n=3–4 mice per group. (E) Tumor weights at endpoint. Data are mean±SEM and n=3–4 mice per group. (F) Representative flow cytometry plots of Granzyme B and CD103 gated on donor T cells. Graphs are the proportion of TCR_Msln cells expressing Granzyme B ex vivo and the mean fluorescence intensity (MFI) of Granzyme B per donor T cell. Data are mean±SEM and n=3–4 mice per group. (G) Representative flow cytometry plots of 41BB expression by engineered T cells. Graphs are the proportion of TCR_Msln cells (left) or endogenous CD8⁺ T cells (right) that express 41BB at 8 days post infusion. Data are mean±SEM and n=3–4 mice per group. (H) Masson’s trichrome of tumor and lung tissue from representative treated mice. Scale bar, 50 µm. (I) Representative cleaved caspase 3 (CC3) staining of tumor and lung at 8 days post infusion. n=3–4 mice per group. (J) Number of CC3⁺ cells in tumor or lung at day 8 or day 28 post T cell infusion. Data are mean±SEM. **P<0.01 (unpaired, two-tailed Student’s t-test). n=3–4 mice per group and 3–4 10×field of views per tissue section.