Intratumoral CD4+ T Cells Mediate Anti-tumor Cytotoxicity in Human Bladder Cancer

Responses to anti-PD-1 immunotherapy occur but are infrequent in bladder cancer. The specific T cells that mediate tumor rejection are unknown. T cells from human bladder tumors and non-malignant tissue were assessed with single-cell RNA and paired T cell receptor (TCR) sequencing of 30,604 T cells from 7 patients. We find that the states and repertoires of CD8(+) T cells are not distinct in tumors compared with non-malignant tissues. In contrast, single-cell analysis of CD4(+) T cells demonstrates several tumor-specific states, including multiple distinct states of regulatory T cells. Surprisingly, we also find multiple cytotoxic CD4(+) T cell states that are clonally expanded. These CD4(+) T cells can kill autologous tumors in an MHC class II-dependent fashion and are suppressed by regulatory T cells. Further, a gene signature of cytotoxic CD4(+) T cells in tumors predicts a clinical response in 244 metastatic bladder cancer patients treated with anti-PD-L1.