TRLS-11 Uncovering Antigen-Specificity of Intratumoral Cd8+ T Cells in Non-Small Cell Lung Cancer Brain Metastases

Abstract While immune-modulating therapies which enhance antitumor T cell function have revolutionized cancer therapy, including for patients with brain metastases, the antigen specificity of intratumoral T cells remains poorly characterized. We performed droplet-based 5’ single-cell RNA sequencing and T cell receptor (TCR) sequencing to characterize the transcriptional profile and clonal repertoire of tumor-infiltrating T cells in resected tumors from patients with previously untreated non-small cell lung cancer (NSCLC) brain metastases (n=6). We identified a population of clonally-expanded, CXCL13+CD8+ T cells enriched for transcriptional signatures predictive of tumor reactivity. Using a high-throughput T cell antigen identification method developed by our group, we screened 164 TCRs from clonally-expanded and/or CXCL13+CD8+ T cells against 2,289 putative neoantigens in addition to 146 tumor-associated antigens. In total, we identified reactivities for 21% (n=35) of the TCRs screened. Consistent with recent studies, we observed that the transcriptional state of the T cell was more predictive of tumor reactivity than the degree of clonal expansion. Of TCR specificities identified, 25.7% (n=9) of TCRs were reactive to neoantigens derived from somatic single-nucleotide variants specific to each patient (private antigen). Intriguingly, 54.3% (n=19) of the TCRs were reactive to unmutated peptides expressed by many patients’ lung tumors (public antigens). In summary, we present a deep characterization of the antigenic reactivities of intratumoral T cells in NSCLC brain metastases. Our data are consistent with an emerging body of literature that has theorized that “missing reactivities” from neoantigen-centric studies arise from complex mechanisms of genetic and epigenetic dysregulation within tumors. Further characterization of these “public antigens” may provide the basis for novel antigen-directed T cell therapies.