Fig. 2: CD39⁺PD-1⁺CD8⁺ T cells exhibit a proliferative exhausted phenotype.

a Schematic illustration of IMC processing of tumor samples with or without tertiary lymphoid structures (TLSs). b IMC processing example for a tumor with TLSs (patient ID: KU37). The IMC images are pseudo-colored as follows: Pan-cytokeratin (PanCK) (white), CD20 (green), CD3 (blue), SMA (red), and Collagen (pink). c Spatial distribution images of PD-1⁻CD8⁺ (orange plots) and PD-1⁺CD8⁺ (blue plots) T cells, derived from three representative IMC images, pseudo-colored as follows: E-cadherin (green) and Nuclei (dark blue). Tumor parenchyma in distribution images is outlined in red. d Each fraction (%) of the annotated immune profile among total cells from the tumor-ROIs for 27 patients at pre-treatment. e Proportion (%) of PD-1⁺CD8⁺ T cells among total cells in tumor ROIs, compared between responders (R, n = 9) vs. non-responders (NR, n = 13); two-sided Mann-Whitney U-test (P = 0.0045). Five NE cases are excluded. f Two representative UMAP plots of CD8⁺ tumor-infiltrating lymphocytes (TILs) (n = 555 cells each) within the tumor-ROIs, colored by the expression of each marker. The selected UMAP plots were chosen for their clear depiction of exhaustion patterns, which were observed in a subset of cases among a total of 11 independent ROIs. g Proportion (%) of cells expressing CD39 among PD-1⁺CD8⁺ T cells in paired pre-treatment PBMCs vs. TILs; two-sided Wilcoxon matched-pairs signed-rank test (n = 27, P < 0.0001). h Median mass intensity (MMI) of Ki67 and scaled expression of TOX in CD39⁺PD-1⁺CD8⁺ TILs vs. CD39⁻PD-1⁻CD8⁺ TILs; two-sided Wilcoxon matched-pairs signed test (each n = 25 independent tumor-ROIs, P < 0.0001 for both comparisons). Two cases with cold tumors lacking CD39⁺PD-1⁺CD8⁺ T cells are excluded. e, g, h Error bars indicate mean ± SEM. e, g n refers to patients. a was created in BioRender. Kenro, T. (2024) BioRender.com/s50j163. Source data are provided as a Source Data file.