Anti-PD-1 Checkpoint Therapy Can Promote the Function and Survival of Regulatory T Cells

We have previously shown in a model of claudin-low breast cancer that regulatory T cells (T_regs) are increased in the tumor microenvironment (TME) and express high levels of PD-1. In mouse models and patients with triple-negative breast cancer, it is postulated that one cause for the lack of activity of anti-PD-1 therapy is the activation of PD-1-expressing T_regs in the TME. We hypothesized that the expression of PD-1 on T_regs would lead to enhanced suppressive function of T_regs and worsen antitumor immunity during PD-1 blockade. To evaluate this, we isolated T_regs from claudin-low tumors and functionally evaluated them ex vivo. We compared transcriptional profiles of T_regs isolated from tumor-bearing mice with or without anti-PD-1 therapy using RNA sequencing. We found several genes associated with survival and proliferation pathways; for example, Jun, Fos, and Bcl2 were significantly upregulated in T_regs exposed to anti-PD-1 treatment. Based on these data, we hypothesized that anti-PD-1 treatment on T_regs results in a prosurvival phenotype. Indeed, T_regs exposed to PD-1 blockade had significantly higher levels of Bcl-2 expression, and this led to increased protection from glucocorticoid-induced apoptosis. In addition, we found in vitro and in vivo that T_regs in the presence of anti-PD-1 proliferated more than control T_regs PD-1 blockade significantly increased the suppressive activity of T_regs at biologically relevant T_reg/T_naive cell ratios. Altogether, we show that this immunotherapy blockade increases proliferation, protection from apoptosis, and suppressive capabilities of T_regs, thus leading to enhanced immunosuppression in the TME.