Systematic lineage tracing reveals clonal progenitors and long-term persistence of tumor-specific T cells during immune checkpoint blockade

Paired single-cell RNA and T cell receptor sequencing (scRNA/TCR-seq) has allowed for enhanced resolution of clonal T cell dynamics in cancer. Here, we report scRNA/TCR-seq analysis of 187,650 T cells from 31 tissue regions, including tumor, adjacent normal tissues, and lymph nodes (LN), from patients who underwent resections for persistent or progressing lung cancers after immune checkpoint blockade (ICB). We found marked regional heterogeneity in CD8+ and CD4+ T cell phenotypes that was associated with heterogeneity in the presence of viable tumor cells. Regions with viable cancer cells were enriched for exhausted CD8+ T cells, regulatory CD4+ T cells (Treg), and follicular helper CD4+ T cells (TFH). Tracking individual T cell clonotypes across tumor regions and tissues, combined with neoantigen specificity assays, revealed that TFH and tumor-specific exhausted CD8+ T cells could be clonally linked to TCF7+ SELL+ progenitors in tumor draining LNs, and progressive exhaustion trajectories of CD8+ T, Treg, and TFH cells with proximity to the tumor microenvironment. Finally, longitudinal tracking of tumor-specific CD8+ and CD4+ T cell clones revealed persistence in the peripheral blood for years after ICB therapy; however, exhausted CD8+, Treg, and TFH cells had lower persistence relative to other subsets. Strikingly, the presence of clonally linked progenitors in the LN conferred greater longitudinal persistence. Altogether, this comprehensive scRNA/TCR-seq dataset with regional, clonal, and longitudinal resolution provides fundamental insights into the tissue distribution, differentiation trajectories, and persistence of the tumor-specific T cells that underlie clinical responses to ICB.