Expression data for CD8 TILs subpopulations sorted by Tim3 and PD1

Reversing the dysfunctional T cell state that arises in cancer and chronic viral infections is the focus of therapeutic interventions; however, current therapies are effective in only some patients and some tumor types. To gain a deeper molecular understanding of the dysfunctional T cell state, we analyzed population and single-cell RNA profiles of CD8+ tumor-infiltrating lymphocytes (TILs) and used genetic perturbations to identify a distinct gene module for T cell dysfunction that can be uncoupled from T cell activation. This distinct dysfunction module is downstream of intracellular metallothioneins that regulate zinc metabolism and can be identified at single-cell resolution. We further identify Gata-3, a zinc-finger transcription factor in the dysfunctional module, as a regulator of dysfunction, and use CRISPR/Cas9 genome editing to show that it drives a dysfunctional phenotype in CD8+ TILs. Our results open novel avenues for targeting dysfunctional T cell states, while leaving activation programs intact. Samples were generated of naive (CD62LhiCD44low) and effector/memory (CD62LlowCD44hi) CD8+ cells from non tumor-bearing Balb/c mice, CD8+Tim3-PD1- (DN) TILs, CD8+Tim3-PD1+(SP), and CD8+Tim3+PD1+ (DP) TILs. Batch indicated in sample name (2157, 1962, 1635, 1655 and 1716).