Differentiation and Expansion of Tumor-Infiltrating T Cell Clonotypes Occurs in the Spleen Following Immune Checkpoint Blockade

Abstract

Immune checkpoint blockade (ICB) enhances tumor-reactive T cell responses against cancer, leading to long-term tumor control and survival in a fraction of patients. Given the increasingly recognized complexity of T cell differentiation that occurs in response to chronic antigen stimulation, it remains unclear precisely which T cell differentiation states are critical for the response to ICB, as well as the anatomic sites at which ICB-mediated reinvigoration of these T cells occurs. We used paired single-cell RNA and T cell receptor (TCR) sequencing to profile endogenous, tumor-reactive CD8+ T cells isolated from tumors, tumor-draining lymph nodes, and spleens of mice treated with ICB. We identified an intermediate-exhausted population of T cells in the spleen which underwent the greatest expansion in response to ICB and gave rise to the majority of tumor-infiltrating clonotypes. Increasing concentrations of antigen in the spleen perturbed the differentiation of this phenotype towards a divergent exhausted_KLR state, resulting in reduced numbers of tumor-infiltrating T cells and blunted ICB efficacy. Likewise, an analogous population of exhausted_KLR CD8+ T cells in matched human tumor and blood samples and exhibited diminished tumor-trafficking ability. These data demonstrate that the spleen is a critical anatomic site for coordinating the differentiation of tumor-infiltrating clonotypes and their expansion in response to ICB.