Single-cell clonal lineage tracing identifies the transcriptional program controlling the cell fate decisions by neoantigen-specific CD8 + T cells

Abstract Neoantigen-specific T cells specifically recognize tumor cells and are critical for cancer immunotherapies. However, the transcriptional program controlling the cell fate decisions by neoantigen-specific T cells is incompletely understood. Here, using joint single-cell transcriptome and TCR profiling, we mapped the clonal expansion and differentiation of neoantigen-specific CD8 + T cells in the tumor and draining lymph node in mouse prostate cancer. Compared to other antitumor CD8 + T cells and bystanders, neoantigen-specific CD8 + tumor-infiltrating lymphocytes (TILs) upregulated gene signatures of T cell activation and exhaustion. In the tumor draining lymph node, we identified TCF1 + TOX - T SCM , TCF1 + TOX + T PEX , and TCF1 - TOX + effector-like T EX subsets among neoantigen-specific CD8 + T cells. Clonal tracing analysis of neoantigen-specific CD8 + T cells revealed greater clonal expansion in divergent clones and less expansion in clones biased towards T EX, T PEX , or T SCM . The T PEX subset had greater clonal diversity and likely represented the root of neoantigen-specific CD8 + T cell differentiation, whereas highly clonally expanded effector-like T EX cells were positioned at the branch point where neoantigen-specific clones exited the lymph node and differentiated into T EX TILs. Notably, T SCM differentiation of neoantigen-specific CD8 + clones in the lymph node negatively correlated with exhaustion and clonal expansion of the same clones in the tumor. In addition, the gene signature of neoantigen-specific clones biased toward tumor infiltration relative to lymph node residence predicted a poorer response to immune checkpoint inhibitor. Together, we identified the transcriptional program that controls the cell fate choices by neoantigen-specific CD8 + T cells and correlates with clinical outcomes in cancer patients.