Characterization of a population of innate like T cells in mouse and human

T lymphocytes expressing a mature TCRαβ are classified in conventional T cells (CD4+ and CD8+) and unconventional T cells (UTCs), including invariant natural killer T cells (iNKT), mucosal-associated invariant T cells (MAIT) and intraepithelial lymphocytes (IELs). Our group recently identified a poorly defined population of UTCs referred to as double-negative αβ T cells (DNTαβ) and involved in resistance against sarcoma. DNTαβ express a TCRαβ but are not recognized by antigen-loaded CD1d tetramers and antigen-loaded MR1 tetramers, indicating that these cells are not iNKT or MAIT. Positive markers of DNTαβ have not been identified and little is known about their phenotype and biology in healthy condition or in pathologies. Therefore, our study aims to investigate the biology and immunobiology of DNT⍺β in mice and humans. We observed that DNT⍺β were present in different organs at different ages of healthy mice and were characterized by a unique set of NK cell-related molecules such as Ly49 receptors and CD94/NKG2A inhibitory complex. In addition, DNT⍺β expressed high levels of the maturation marker of T cells CD44. In line with this observation, gene set enrichment analysis identified a transcriptomic signature associated with immune response (i.e., Il1r1, Ifngr1) and lymphocyte activation (i.e., Tigit, Lag3 and Tim3) in DNT⍺β compared to conventional T cells and iNKT. We found that DNT⍺β expressed a polyclonal TCR repertoire. Moreover, using mice deficient for different antigens presenting molecules (i.e., CD1d-/-, MR1-/-, b2m-/- and MHC-II-/-) we provided evidence that DNTαβ undergo maturation independently of CD1d, MR1, MHC-I and MHC-II molecules, suggesting that DNTαβ differ from other unconventional T cell subsets such as iNKT, MAIT and from conventional CD4 and CD8 T cells. Interestingly, DNTαβ showed proliferation, production of effector molecules (i.e., interferon gamma (IFN-γ) and granzyme B (GZMB)) and cytotoxic activity against tumor cells. Finally, we identified a tumor-infiltrating population of DNTαβ in 3- methylcholanthrene (MCA)-induced sarcoma in mice and we obtained the first evidence for the presence and phenotype of DNTαβ in human sarcoma. Collectively, our results revealed specific features of a subset of UTCs, both in mouse and human, that could represent a potential new target for cancer therapy.