Immunosuppressive Myeloid Cells Induce Nitric Oxide–Dependent DNA Damage and p53 Pathway Activation in CD8⁺ T Cells

Abstract

Tumor-infiltrating myeloid-derived suppressor cells (MDSC) are associated with poor survival outcomes in many human cancers. MDSCs inhibit T cell–mediated tumor immunity in part because they strongly inhibit T-cell function. However, whether MDSCs inhibit early or later steps of T-cell activation is not well established. Here we show that MDSCs inhibited proliferation and induced apoptosis of CD8⁺ T cells even in the presence of dendritic cells (DC) presenting a high-affinity cognate peptide. This inhibitory effect was also observed with delayed addition of MDSCs to cocultures, consistent with functional data showing that T cells expressed multiple early activation markers even in the presence of MDSCs. Single-cell RNA-sequencing analysis of CD8⁺ T cells demonstrated a p53 transcriptional signature in CD8⁺ T cells cocultured with MDSCs and DCs. Confocal microscopy showed induction of DNA damage and nuclear accumulation of activated p53 protein in a substantial fraction of these T cells. DNA damage in T cells was dependent on the iNOS enzyme and subsequent nitric oxide release by MDSCs. Small molecule–mediated inhibition of iNOS or inactivation of the Nos2 gene in MDSCs markedly diminished DNA damage in CD8⁺ T cells. DNA damage in CD8⁺ T cells was also observed in KPC pancreatic tumors but was reduced in tumors implanted into Nos2-deficient mice compared with wild-type mice. These data demonstrate that MDSCs do not block early steps of T-cell activation but rather induce DNA damage and p53 pathway activation in CD8⁺ T cells through an iNOS-dependent pathway.