Supplementary Figure 2: Prior PD-1 blockade abrogates vaccine-induced tumor-specific immune responses early during the course of treatment.

Tumor tissues were harvested 3 days after priming (D13) or three days after boosting (D20) from B16 melanoma-bearing mice. a-d. Numbers of total (a & c) and antigen-specific CD8⁺ T cells (b & d) at D20 (a-b) and at D13 (c-d). e-h. Frequencies of Annexin V⁺ total (e & g) and antigen-specific CD8⁺ T cells (f & h) in the TME at D20 (e-f) and at D13 (g-h) as indicated. i-l. Frequencies of CD40L⁺ (i and k) and IFN-γ⁺ (j & l) CD8⁺ T cells in the TME at D20 (i-j) and at D13 (k-l) as indicated. Flow cytometry data are the average of two independent experiments. Each dot corresponds to one mouse with the indicated number of mice per group given in parentheses. For comparison purposes, an unpaired, one-tailed Student’s t-test was used. The error bars indicate the s.e.m. ^NSnon-significant (a) *(lower) p=0.0228, *(middle) p=0.0277, *(upper) p=0.0286, **(left) p=0.0012, **(right) p=0.0014; (b) *(lower) p=0.0333, *(middle) p=0.042, *(upper) p=0.0472, ****p≤0.0001; (d) *(lower) p=0.018, *(upper) p=0.0207, **p=0.0072, ***p=0.0004, ****p≤0.0001; (e) *(lower) p=0.05, *(upper) p=0.03, **p=0.0041; (f) *(lower) p=0.0462, *(upper) p=0.0181, **p=0.0094; (g) *(lower) p=0.0157, *(upper) p=0.0469, ***p=0.0006; (h) *(lower) p=0.0265, *(upper) p=0.0152, **p=0.0062 (i) *(left) p=0.0384, *(middle) p=0.0399, *(right) p=0.0165; (j) *(lower) p=0.0414, *(upper) p=0.0260, **p=0.0013, ***p=0.0007; (k) *p=0.0343, **p=0.006, ****p≤0.0001; (l) *p=0.0156. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001.