Abstract 1603: TNFR2+ regulatory T cells are key players in immune escape in gastric tumor microenvironment

Abstract BACKGROUND & AIMS: Regulatory T cells (Tregs) are potent immunosuppressive cells of the immune system. It is reported that the expression of TNFR2 identifies a subpopulation of Tregs with the maximally suppressive function. Thus, understanding the heterogeneity of human CD4+FOXP3+ Tregs and the function of TNFR2+ Tregs is of critical importance for moving Treg therapy into the clinics. METHODS: The expression phenotype of TNFR2+ Tregs from gastric cancer patients and normal controls were detected by flow cytometry. Further, we quantified the infiltrated Foxp3+ Tregs and TNFR2+ Tregs of gastric cancer patients using multicolor immunofluorescence assay. Moreover, to generate a deep transcriptional map of the Tregs in human gastric cancer, we profiled 9058 Tregs from three gastric carcinomas, as well as 16769 Tregs from matched peripheral blood mononuclear cells (PBMCs), using single-cell RNA-seq. RESULTS: In the peripheral blood of gastric cancer patients and healthy individuals, Tregs expressed markedly higher levels of TNFR2 than conventional T cells (Tconvs). The proportion of total FoxP3+ Tregs in the CD4+ cells and the percentage of TNFR2+ Tregs increased with tumor progression and lymphatic metastasis. Analysis of single-cell RNA sequencing data revealed that Tregs in gastric carcinomas microenvironment exhibited dissimilar phenotypes compared with Tregs in matched PBMCs. We observed lower phenotypic heterogeneity of tumor-derived Tregs. Tumor infiltrating Tregs expressed higher levels of immune checkpoint genes compared with peripheral blood Tregs, indicating they were in more activated states. Besides, we found the expression level of TNFR2 in two tumor-derived clusters was much higher compared with others. Treg biomarkers and immune checkpoint genes were also highly expressed in these two populations, suggesting that they were the main immunosuppressive Treg proportions in tumor microenvironment. Gene enrichment analysis indicated that the two Treg populations showed an overlap of gene expression signatures of Th1, Th2, and Th17 cells, which was consistent with the increasing evidence that Tregs can exhibit helper T cell-like phenotype to promote Tregs migrating and suppressing immune response. CONCLUSIONS: Our findings revealed that TNFR2 could be a promising molecular marker associated with poor prognosis of patients baring gastric cancer. We also established the heterogeneity at the single-cell level of CD4+FOXP3+ Tregs, explaining the remarkable complexity and functional diversity of human Tregs. Finally, our work provide important clues for exploring the mechanism by which TNFR2 regulates the phenotype and activity of Tregs in the tumor microenvironment. Citation Format: Liling Niu. TNFR2+ regulatory T cells are key players in immune escape in gastric tumor microenvironment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1603.