Fig. 1

Anti-4-1BB induces tumor regression in CD73 deficient mice. WT and CD73^−/− mice were injected s.c. with B16-SIY melanoma cells and treated with anti-4-1BB or control IgG. a Tumor size was measured every 2–4 days. b Survival curves of B16-SIY-bearing mice (5 mice per group). c B16-SIY tumors from treated WT and CD73^−/− mice were harvested 18 days after tumor challenge and analyzed by flow cytometry for accumulation of infiltrating CD3⁺TCRβ⁺, CD3⁺CD4⁺ and CD3⁺CD8⁺ T cells. d Absolute number of CD4⁺, CD4⁺Foxp3⁺, and CD8⁺ T cells per gram of tumors were also calculated. e Percentage of other main immune cell subsets as indicated of total CD45⁺ tumor infiltrates. f Percentage of different Treg-associated markers for CD4⁺Foxp3⁺ T cell infiltrates in the treated B16-SIY-bearing WT and CD73^−/− mice. g Ratio of CD4⁺Foxp3^- (effectors) to CD4⁺Foxp3⁺ cells (Tregs) and CD8⁺ to CD4⁺Foxp3⁺ cells. h Percentage of Ki67⁺ among tumor-infiltrating CD4⁺ or CD8⁺ T cells and (i) representative dot plots. j Percentage of IFN-γ⁺ in tumor-infiltrating CD8⁺ T cells, and (k) representative dot plots. l The ratio of CD8⁺IFN-γ⁺ to CD4⁺Foxp3⁺ Tregs was also calculated. *p < 0.05, **p < 0.01, ***p < 0.001. ANOVA analysis, log-rank test and unpaired Student’s two-tailed t test were used. Data (mean ± SEM) are representative of at least 2 independent experiments with 3–5 independently analyzed mice/group