TOX-expressing Terminally Exhausted Tumor-infiltrating CD8 T Cells Are Reinvigorated by Co-blockade of PD-1 and TIGIT in Bladder Cancer

Exhausted T cells in the tumor microenvironment are major targets of immunotherapies. However, the exhaustion status of CD8 tumor-infiltrating lymphocytes (TILs) in bladder cancer has not been comprehensively evaluated. Herein, we examined distinct exhaustion status of CD8 TILs based on the level of programmed cell death-1 (PD-1) and thymocyte selection-associated high mobility group box protein (TOX) expression in urothelial bladder cancer. We also evaluated the reinvigoration of exhausted CD8 TILs upon ex vivo treatment with inhibitory checkpoint blockers. TOX-expressing PD-1CD8 TILs had the highest expression of immune checkpoint receptors (ICRs), the most terminally exhausted features, and the highest tumor antigen reactivity among PD-1CD8 TILs. Bladder cancer patients with a high percentage of PD-1TOXCD8 TILs had more progressed T-cell exhaustion features and higher programmed death-ligand 1 expression in tumor tissues. TIGIT was the most frequent co-expressed ICR on PD-1CD8 TILs, and TIGIT blockade enhanced the PD-1 blockade-mediated cytokine production by CD8 TILs from bladder cancer patients. Our findings provide an improved understanding of the heterogeneous exhaustion status of CD8 TILs and additional immunotherapy strategies to improve outcomes of bladder cancer patients.