PD-1 regulates tumor-infiltrating CD8+ T cells in both a cell-intrinsic  and cell-extrinsic fashion [scRNA-seq]
Although PD-1 inhibitors are FDA-approved for over 25 different cancers, the mechanisms contributing to response remain incompletely understood. To elucidate the impact of PD-1 deleted CD8+ T cells on PD-1-expressing CD8+ T cells in the same tumor microenvironment, we developed an inducible PD-1 knockout (KO) model in which PD-1 is deleted on approximately 50% of cells. PD-1 deletion beginning at day 7 after implantation of MC38 tumor cells induced robust tumor control. Remarkably, PD-1-expressing CD8+ T cells in the tumor had increased functionality similar to PD-1 KO CD8+ T cells. Using single cell RNA seq and TCR seq, we found that the major transcriptional changes following PD-1 deletion were shared by PD-1 KO and PD-1-expressing CD8+ T cells, though PD-1 KO clones preferentially expanded. These data suggest PD-1 inhibitors not only exert cell-intrinsic effects but also may promote increased T cell function via non-cell autonomous mechanisms, which has important implications for design of PD-1-based cancer immunotherapies. Overall design: Cre+ and Cre- PD-1 fl/fl mice bearing subcutaneous MC38 tumors were treated with tamoxifen such that roughly 50% of cells experienced deletion of exons 2-4 of Pdcd1. CD8+ T cells were obtained and use to generate gene expression, V(D)J, and feature barcoding libraries.