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-1highCD8+ TILs had the highest expression of immune checkpoint receptors (ICRs), the most terminally exhausted features, and the highest tumor antigen reactivity among PD-1+CD8+ TILs. Bladder cancer patients with a high percentage of PD-1highTOX+CD8+ 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-1+CD8+ 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.

Han HS ，Jeong S ，Kim H ，Kim HD ，Kim AR ，Kwon M ，Park SH ，Woo CG ，Kim HK ，Lee KH ，Seo SP ，Kang HW ，Kim WT ，Kim WJ ，Yun SJ ，Shin EC ... -  《-》

TIGIT and PD1 Co-blockade Restores ex vivo Functions of Human Tumor-Infiltrating CD8(+) T Cells in Hepatocellular Carcinoma.

TIGIT is a co-inhibitory receptor, and its suitability as a target for cancer immunotherapy in HCC is unknown. PD1 blockade is clinically effective in about 20% of advanced HCC patients. Here we aim to determine whether co-blockade of TIGIT/PD1 has added value to restore functionality of HCC tumor-infiltrating T cells (TILs). Mononuclear leukocytes were isolated from tumors, paired tumor-free liver tissues (TFL) and peripheral blood of HCC patients, and used for flow cytometric phenotyping and functional assays. CD3/CD28 T-cell stimulation and antigen-specific assays were used to study the ex vivo effects of TIGIT/PD1 single or dual blockade on T-cell functions. TIGIT was enriched, whereas its co-stimulatory counterpart CD226 was down-regulated on PD1high CD8+ TILs. PD1high TIGIT+ CD8+ TILs co-expressed exhaustion markers TIM3 and LAG3 and demonstrated higher TOX expression. Furthermore, this subset showed decreased capacity to produce IFN-γ and TNF-α. Expression of TIGIT-ligand CD155 was up-regulated on tumor cells compared with hepatocytes in TFL. Whereas single PD1 blockade preferentially enhanced ex vivo functions of CD8+ TILs from tumors with PD1high CD8+ TILs (high PD1 expressers), co-blockade of TIGIT and PD1 improved proliferation and cytokine production of CD8+ TILs from tumors enriched for PD1int CD8+ TILs (low PD1 expressers). Importantly, ex vivo co-blockade of TIGIT/PD1 improved proliferation, cytokine production, and cytotoxicity of CD8+ TILs compared with single PD1 blockade. Ex vivo, co-blockade of TIGIT/PD1 improves functionality of CD8+ TILs that do not respond to single PD1 blockade. Therefore co-blockade of TIGIT/PD1 could be a promising immune therapeutic strategy for HCC patients.

Ge Z ，Zhou G ，Campos Carrascosa L ，Gausvik E ，Boor PPC ，Noordam L ，Doukas M ，Polak WG ，Terkivatan T ，Pan Q ，Takkenberg RB ，Verheij J ，Erdmann JI ，IJzermans JNM ，Peppelenbosch MP ，Kraan J ，Kwekkeboom J ，Sprengers D ... -  《Cellular and Molecular Gastroenterology and Hepatology》

Immune Checkpoint Inhibitor-induced Reinvigoration of Tumor-infiltrating CD8(+) T Cells is Determined by Their Differentiation Status in Glioblastoma.

Immune checkpoint inhibitors (ICI) are used for the treatment of various cancers, but clinical trials of anti-programmed cell death protein 1 (PD-1) with patients with recurrent glioblastoma (GBM) have failed to show clinical benefits. In this study, we examined the differentiation status of CD8+ tumor-infiltrating lymphocytes (TIL) from patients with primary GBM and their reinvigoration by ICIs to understand the nature of T-cell exhaustion in GBM. We isolated TILs from 98 patients with newly diagnosed GBM and examined the expression of immune checkpoint receptors and T-cell transcription factors using flow cytometry. TILs were ex vivo stimulated with anti-CD3 in the presence of anti-PD-1 and/or anti-cytotoxic T-lymphocyte antigen 4 (CTLA-4) and their proliferation assessed. CD8+ TILs had significantly increased expression of immune checkpoint receptors, including PD-1 and CTLA-4, compared with peripheral blood CD8+ T cells. Among CD8+ TILs, PD-1+ cells exhibited more terminally differentiated phenotypes (i.e., EomeshiT-betlo) than PD-1- cells. These data were confirmed by analyzing NY-ESO-1157-specific CD8+ TILs. Evaluating the proliferation of CD8+ TILs after ex vivo stimulation with anti-CD3 and anti-PD-1, we found that proliferation inversely correlated with the percentage of EomeshiT-betlo cells among PD-1+CD8+ TILs. When anti-CTLA-4 was used in combination with anti-PD-1, an additional increase in CD8+ TIL proliferation was observed in patients with low percentages of EomeshiT-betlo CD8+ TILs, who responded well to anti-PD-1 in ex vivo assays, but not in patients with high percentages of EomeshiT-betlo CD8+ TILs, who did not respond to anti-PD-1. In primary GBM, the differentiation status of CD8+ TILs determines their reinvigoration ability upon ICI treatment.

Park J ，Kwon M ，Kim KH ，Kim TS ，Hong SH ，Kim CG ，Kang SG ，Moon JH ，Kim EH ，Park SH ，Chang JH ，Shin EC ... -  《-》

Single-cell transcriptome analysis reveals TOX as a promoting factor for T cell exhaustion and a predictor for anti-PD-1 responses in human cancer.

Kim K ，Park S ，Park SY ，Kim G ，Park SM ，Cho JW ，Kim DH ，Park YM ，Koh YW ，Kim HR ，Ha SJ ，Lee I ... -  《Genome Medicine》

TIGIT and PD-1 impair tumor antigen-specific CD8⁺ T cells in melanoma patients.

Chauvin JM ，Pagliano O ，Fourcade J ，Sun Z ，Wang H ，Sander C ，Kirkwood JM ，Chen TH ，Maurer M ，Korman AJ ，Zarour HM ... -  《-》