Abstract 3176: Inhibitory activity of Globo-H and SSEA-4 on activated T-cells

Abstract

Abstract Background/Aim: Globo-H (GH) and stage-specific embryonic antigen-4 (SSEA-4) are globo-series glycosphingolipid (GSL) antigens highly expressed on a variety of epithelial cancers. GH and SSEA-4 are reported to play an important role in cancer cell development and survival and thus are promising targets for developing immunotherapeutic agents for cancer treatment. Shedding of GSLs by tumor cells and uptake by neighboring cells may generate a preferable environment for tumor progression and survival within the tumor microenvironment. GH shed from tumor cells was shown to promote angiogenesis and suppress immune responses in vitro. In addition, exogenous Globo-H ceramide (GHCer) inhibits proliferation of human PBMCs and murine CD4+ T-cells. Given the important role of sialic acid in regulating the immune system, SSEA-4, a polysaccharide with sialic acid at the terminal, may play an important role in the regulation of T-cell responses. This study aimed to investigate the GHCer and SSEA-4 ceramide (SSEA-4Cer) effects on T-cell activation. Methods: C57BL/6 mice bearing murine B16F10 melanoma were treated with GHCer by peritumoral injection and monitored for 26 days. T-cell tumor subpopulations were evaluated by FACS and immunohistochemical analysis. Presence of GHCer and SSEA-4Cer expression on human T cells after the treatment with GHCer and SSEA-4Cer was monitored by FACS analysis. Cell proliferation and apoptosis of T cells after treatment with GHCer, SSEA-4Cer, and glucoceramide (GlcCer) were assessed using ATP assay and annexin V staining, respectively. After treatment with GHCer/SSEA-4Cer/GlcCer, expression levels of CD25, cytokine IFN-γ, and Ki-67 in human PBMCs stimulated with anti-CD3/CD28 were measured by flow cytometry. Results: B16F10 tumor growth rate increased 2.54-fold (P<0.05, n=6) in GHCer-treated mice with a lower number of tumor-infiltrating CD3+ cells (0.45-fold; P<0.01, n=6) compared with control mice. Not surprisingly, addition of GHCer and SSEA-4Cer to human T cells was shown to increase GH and SSEA-4 levels on the surface of T cells. Furthermore, addition of GHCer and SSEA-4Cer to the activated T cells showed a 26.4% and 36.9% decrease in cell proliferation, respectively, while no induction of apoptosis was observed. In contrast, GHCer had no impact on cell viability and proliferation of resting T cells. Exogenous GHCer and SSEA-4Cer significantly reduced CD3+CD25+ and CD3+IFNγ+ T cells in anti-CD3/CD28-stimulated PBMCs. SSEA-4Cer exhibited higher activity in the suppression of CD8+CD25+ T cells in PBMCs than GHCer did. In addition, suppression of the proliferation of activated T-cells by SSEA-4Cer was abolished in the presence of NF-κB inhibitor. Conclusion: GHCer inhibits T-cell infiltration into the tumor in vivo and SSEA-4 exhibits immunosuppressive activity on activated T cells, suggesting that both GHCer and SSEA-4Cer may play a role in regulating immunologic responses in the tumor microenvironment. Citation Format: Tzer-Min Kuo, Chin-Chan Lee, Jiann-Shiun Lai, Chung-Chen Su, Ming-Tain Lai. Inhibitory activity of Globo-H and SSEA-4 on activated T-cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3176.