Fig. 6.

Pharmacological inhibition of NOS2 ameliorated the efficacy of PD-1 blockade in various tumor models. Concomitant blockade and inhibition of PD-1 and NOS2 with L-NAME in established MCA205WT a and MC38 b tumors. Tumor growth kinetics (left and middle panels) and survival curves (right panels) are depicted for each group. L-NAME treatment (1 g/L) was started one day prior to anti-PD-1 infusion and was maintained until the end of the experiment. The graphs depict tumor growth kinetics of a pool of 2–3 independent experiments encompassing 5–9 mice per group and per experiment. c Flow cytometry analyses of TILs after 4 injections focusing on the proportion of FOXP3⁺ cells among CD4⁺ T cells and the ratio of CD8⁺ T cells/CD4⁺ FOXP3⁺ Treg cells are depicted. Each dot represents 1 animal and graphs depict the pool of 3 independent experiments. Means ± SEM are represented. Statistical analyses were performed using ANOVA statistical tests and pairwise comparisons with Bonferroni adjustment. d Effect of L-NAME in anti-PD-1-treated MCA205WT tumors inoculated into WT or Nos2^−/− mice. The graphs depict a pool of 2 independent experiments including 5–7 mice/group and per experiment. Statistical analyses were performed using the specific software detailed in the Material and Methods. *p < 0.05, **p < 0.01, ***p < 0.001, n.s.: not significant