ABSTRACT
Background Functional states of T cells are heterogeneous in tumor microenvironment. Immune checkpoint inhibitor (ICI) can reinvigorate only stem-like exhausted T cells, suggesting that impeding progress of exhaustion will improve immunotherapy efficacy. Transcription factors promoting T-cell exhaustion could be potential targets for delaying the process, improving efficacy of ICI.
Methods Analyzing single-cell transcriptome data derived from melanoma and non-small cell lung cancer (NSCLC), we divided tumor-infiltrating CD8+ T-cell population by PDCD1 (also known as PD-1) expression level into PDCD1-high cells and PDCD1-low cells and identified differentially expressed genes as candidate factors promoting intratumoral T-cell exhaustion. Co-expression of candidate genes with immune checkpoint molecules among tumor-infiltrating CD8+ T cells was confirmed by single-cell trajectory analysis and flow-cytometry analysis. Loss-of-function effect of the candidate regulator was examined by cell-based knockdown assay. Clinical effect of the candidate regulator was evaluated based on overall survival and anti-PD-1 responses.
Results We identified TOX among the most differentially expressed transcription factors between PDCD1-high subset and PDCD1-low subset of tumor-infiltrating CD8+ T cells in melanoma and NSCLC. TOX expression level tend to increase as CD8+ T cells become more exhausted. Flow-cytometry analysis of tumor-infiltrating T cells showed correlation of TOX expression with severity of intratumoral T-cell exhaustion. TOX knockdown resulted in downregulation of PD-1 and TIM-3, suggesting that TOX promotes intratumoral T-cell exhaustion by inducing immune checkpoint molecules. Finally, we found that TOX expression in tumor-infiltrating T cells was predictive for overall survival and anti-PD-1 efficacy in melanoma and NSCLC.
Conclusions We found TOX to promote intratumoral CD8+ T-cell exhaustion via positive regulation of PD-1—TIM-3 cooperation, suggesting that inhibition of TOX potentially impede T-cell exhaustion and improve efficacy of ICI. In addition, TOX expression in tumor-infiltrating T cells would be a useful feature for patient stratification in anti-tumor treatments including anti-PD-1 immunotherapy.
