Investigating the role of DNA damage in CD8+ T-cell differentiation

The human T-cell compartment consists of distinct subsets, each with a specific role in mounting an adaptive immune response. Within the tumor microenvironment (TME), multiple stressors contribute to the development of a dysfunctional differentiation state, namely T-cell exhaustion. Exhausted T cells (TEX) are characterized by diminished effector functions, higher expression of inhibitory receptors, altered cellular metabolism, and impaired proliferative potential. Chronic exposure to tumor antigens represents a major mechanism involved in the acquisition of exhaustion traits which, together with chronic induction of proliferation and response to harmful stimuli such as increased reactive oxygen species (ROS), hypoxia, and replication stress result in DNA damage accumulation. We hypothesized that DNA damage regulates T-cell differentiation and function. We analyzed publicly available transcriptomic datasets from eight different tumor types, revealing that CD39+ tumor-reactive CD8+ T cells are highly enriched in DNA damage response (DDR) signatures. Indeed, CD39+ tumor-infiltrating lymphocytes (TILs) accumulated single- and double-strand DNA breaks when compared with bystander tumor-infiltrating or circulating CD8+ T cells from the same non-small cell lung cancer (NSCLC) patient. Among the factors potentially involved in DNA repair, we identified 2′-deoxynucleoside 5′-monophosphate N-glycosidase (DNPH1), a nucleotide sanitizer that protects cells by limiting the genomic incorporation of toxic nucleotides, ensuring correct cell-cycle progression. An in vitro system, optimized ad hoc to recapitulate T-cell exhaustion, revealed that CD8+ TEX largely depend on DNPH1 for proliferation and survival. In particular, DNPH1 inhibition in vitro resulted in the loss of genes related to DNA repair and genome integrity signatures. We propose DNPH1 as a novel factor required for the maintenance of TEX.