137 Decreased cytotoxic T cells, decreased cytotoxic/regulatory T-cell ratio, and decreased TCR clonality are associated with increased numbers of primary cutaneous squamous cell carcinomas in solid organ transplant recipients

Abstract

Background: We developed a novel analytical method to define tumor infiltrating lymphocyte (TIL) phenotype in immune competent vs. immune suppressed cSCC patients based on data obtained from single cell sequencing and gene expression. Methods: CD8+ TILs (n = 34,399 for cSCC, n = 14,902 for TSCC) obtained from fresh tumor specimens from immunocompetent (n = 5) versus organ transplant recipients (OTRs; n=6) were subject to single-cell RNA profiling matched with T-cell receptor (TCR) sequencing via barcoding. Data were analyzed using iCellR, a custom R packaged we developed for single cell sequencing analysis. Antigens recognized by the top 10 clonotypes for each sample were assessed using McPAS. Results: Gene expression analysis showed TSCC had fewer cytotoxic cells (56% vs. 66%, p < 0.0001), fewer naïve cells (18% vs. 26% p < 0.0001), similar numbers of regulatory cells (8% vs. 5%) and similar numbers of exhausted T cells (9% vs. 10%). CD8+ TILs from TSCC exhibited more homogeneous gene expression compared with immunocompetent patients. TILs from both OTRs and immunocompetent patients showed clonality. However, fewer TCR clonotypes were observed in immune suppressed transplant patients (mean = 544 vs. 1140, p < 0.05). Many of the TCR sequences represent antigens previously reported in melanoma, other carcinomas and viral infections. The majority of TCR sequences for the top 10 clonotypes of each sample have known antigens, but up to 24% of these sequences recognize putative neoantigens. Solid organ transplant recipients in our study showed increased numbers of cSCC events over 12 months (6.2 vs. 1.2, p < 0.01). Conclusion: TSCC TILs tend to show decreased cytotoxic CD8+ TIL, decreased cytotoxic/regulatory T cell ratio and increased exhausted and naïve T cells and decreased cytotoxic T cells. This may contribute to the higher numbers of primary cSCC seen in OTRs.