Differentiation of neoantigen-reactive CD8 T cells in tumors from patients with mismatch repair-proficient colorectal cancers 4297

Description While the phenotype of neoantigen-reactive CD8 T cells in solid tumors has been identified by us and others, little is known about their differentiation in mismatch repair-proficient (MMR-p) colorectal cancers (CRC). To address that, we analyzed paired adjacent normal colon (NC) and primary tumors from patients with MMR-p CRC. Flow cytometry analysis showed a higher T-cell infiltration in tumors, with an enrichment in CD8 T cells. Both, NC and tumor contained a population of CD39+CD103 + (DP) CD8 T cells, a phenotype reminiscent of tumor-reactive T cells. Yet, DP CD8 T cells from the NC lacked PD-1 and Ki-67. To uncover the differentiation of tumor-reactive DP CD8 T cells, we performed scRNA-seq and scTCR-seq on paired NC and tumor from five patients. Using gene expression and RNA velocity, our analysis revealed that tumor-infiltrating DP CD8 T cells (DP CD8 TILs) undergo a linear progression toward an exhausted state and acquire a cytotoxic potential. Neoantigen discovery on three patients identified neoantigens recognized by DP CD8 TILs. The same neoantigens were also recognized by memory CD8 T cells in the blood at time of surgery, albeit at very low frequencies. TCR sequencing of the neoantigen-reactive T cells confirmed their DP CD8 nature but highlighted variability in their differentiation status. This work demonstrates the existence of neoantigen-reactive T cells in the blood and tumor of patients with MMR-p CRC which could be harnessed to promote tumor control. Funding Sources Supported by the Providence Portland Medical Foundation. Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)