Fig. 3

Correlation of TOX expression with the severity of CD8⁺ T cell exhaustion in human tumors. a–c Flow cytometric analysis of the tumor-infiltrating (TI) CD8⁺ T cells isolated from human non-small cell lung cancer (NSCLC) (n = 20) and head and neck squamous cell carcinoma (HNSCC) (n = 15). a Representative plots showing the co-expression of TOX and immune checkpoint (IC) molecules (PD-1, TIM-3, and TIGIT) in the TI CD8⁺ T cells. b Percentage of TOX⁺ cells in the two subpopulations of the TI CD8⁺ T cells (expressing or not expressing a specific IC molecule). Each line in the graph indicates the data derived from the same tumor tissue of each individual patient. c TOX protein levels in the three subsets of TI CD8⁺ T cells with different severities of exhaustion, i.e., PD-1⁻TIM-3⁻ (orange), PD-1⁺TIM-3⁻ (blue), and PD-1⁺TIM-3⁺ (red). Histogram represents TOX expression level in each subset of the TI CD8⁺ T cells. Percentage of TOX-expressing cells in each subset is described in the histogram, and mean fluorescence intensity (MFI) for TOX expression in each subset is indicated in parenthesis. A dashed line represents the boundary separating the TOX protein expression. Distribution of TOX-expressing subsets of TI CD8⁺ T cells across patients is summarized in grouped scattered plots. ns, not significant; *P < 0.05; **P < 0.01; ***P < 0.001. All statistical analyses were performed using unpaired Student’s t test