Fig. 7: Caffeine enhances the effects of PD-1 antibody immunotherapy.

a Schematic depicting the treatment of caffeine (0.1% w/v, daily) or anti-PD-1 monoclonal antibody (PD-1 mAb, 10 mg/kg, ip, twice weekly) in subcutaneous CT26 tumor-bearing BALB/c mice. Representative images of tumor from caffeine-treated or PD-1 mAb groups (n = 8). Scale bars, 1 cm. Quantification of the tumor growth curve and tumor weight. b Waterfall plot on tumor volume changes (n = 8) and c Kaplan-Meier survival curves of subcutaneous CT26 tumor-bearing BALB/c mice treated with caffeine, PD-1 mAb, or their combination (n = 6). d Flow cytometry analysis of GZMB, IFN-γ, Ki67 of CD8⁺ T cells in tumor of subcutaneous CT26 tumor-bearing BALB/c mice treated with caffeine, PD-1 mAb, or their combination (n = 4). e Schematic depicting the treatment of caffeine or PD-1 mAb in subcutaneous MC38 tumor-bearing C57BL/6 mice. Representative images of tumor from caffeine-treated or PD-1 mAb groups (n = 6). Scale bars, 1 cm. Quantification of the tumor growth curve and tumor weight. f Waterfall plot on tumor volume changes (n = 6) and g Kaplan-Meier survival curves of subcutaneous MC38 tumor-bearing C57BL/6 mice treated with caffeine, PD-1 mAb, or their combination (n = 6). h Flow cytometry analysis of GZMB, IFN-γ, Ki67 of CD8⁺ T cells in tumor of subcutaneous MC38 tumor-bearing C57BL/6 mice treated with caffeine, PD-1 mAb, or their combination (n = 4). i Analysis of mRNA expression of COL12A1 associated with response to anti-PD-1 therapy in multiple cancers in public datasets, Zhao_2019_GBM, (PRJNA482620, n = 14), Vanallen_2015_Melanoma (PHS000452, PD/SD, n = 66, PR/CR, n = 39). j Kaplan-Meier survival analysis of patients treated with anti-PD-1 therapy in public datasets. k Immunohistochemistry staining and quantification of COL12A1 in pre-immunotherapy CRC samples from responders (n = 8) and non-responders (n = 13), scare bar, 50 μm. l Representative images of immunofluorescence co-staining for COL12A1 (red) and CD8 (green) in pre-immunotherapy CRC samples from responders and non-responders. Representative images from n = 6 independent experiments. Scare bar, 50μm. m Schematic diagram. Data error bars are mean ± SD. The statistical analyse of growth curve is Two-way ANOVA, survival curve is Log-rank test, the others are Student’s two-tailed unpaired t-test. The images of mice elements and the schematic diagram of graphical abstract were drawn by Figdraw. Source data are provided as a Source Data file.