Low TCR binding strength results in increased progenitor-like CD8+ tumor-infiltrating lymphocytes.

T cell receptor (TCR) binding strength to peptide-MHC antigen complex influences numerous T cell functions. However, the vast diversity of a polyclonal T cell repertoire for even a single antigen greatly increases the complexity of studying the impact TCR affinity has on T cell function. Here, we determined how TCR binding strength affected the protein and transcriptional profile of an endogenous, polyclonal T cell response to a known tumor-associated antigen (TAA) within the tumor microenvironment (TME). We confirmed that flow and CITE-Seq counts of MHC-tetramer labeling were reliable surrogates for the TCR-peptide-MHC steady-state binding affinity. We further demonstrated by single-cell RNA sequencing that tumor-infiltrating lymphocytes (TILs) with high and low binding affinity for a TAA can differentiate into cells with many antigen-specific transcriptional profiles within an established TME. However, more progenitor-like phenotypes were significantly biased towards lower affinity T cells, and proliferating phenotypes showed significant bias towards high-affinity TILs. Additionally, we found in a progressing TME that higher affinity T cells advanced more rapidly to terminal phases of T cell exhaustion and exhibited better tumor control. We confirmed the polyclonal TIL results using a TCR transgenic mouse possessing a single low-affinity TCR targeting the same TAA. These T cells maintained a progenitor-exhausted phenotype and exhibited impaired tumor control. We propose that high-affinity TCR interactions drive T cell fate decisions more rapidly than low-affinity interactions and that these cells differentiate faster. These findings illustrate divergent forms of T cell dysfunction based on TCR affinity which may impact TIL therapies and antitumor responses.