Abstract A022: Potentiating the antitumor response of CD8+ T cells by targeting SQLE-mediated cholesterol metabolism reprogramming in hepatocellular carcinoma

Abstract Background: Accumulating evidence suggests that aberrant cholesterol metabolism plays a pivotal role in modulating immune function and tumor progression. Among various enzymes involved in cholesterol metabolism, squalene epoxidase (SQLE) stands out as a rate-limiting enzyme in cholesterol synthesis, and its aberrant expression has been found to have clinical relevance in several cancer types. However, the immunomodulation role of SQLE remains largely unexplored. Here our goal was to investigate the role of SQLE on the hepatocellular carcinoma (HCC) immune microenvironment and explore a new strategy that strengthens immunotherapy in HCC. Methods: Multiplex immunohistochemical staining and spectral flow cytometry were applied to describe the immune landscape of human and mouse HCC based on SQLE expression level. Additionally, human peripheral blood-derived CD8+T cells were co-cultured with SQLE-edited HCC cell lines to assess differences in cellular proliferation and immune function. By using metabolomics and transcriptomics analyses, we investigated the regulatory mechanisms of SQLE on CD8+ T cell immune function. Furthermore, we evaluated the combined anti-tumor strategy of SQLE inhibitor terbinafine with anti-PD1 treatment in an HCC mouse model that is refractory to PD1 monotherapy. Results: Our results describe a distinct HCC immune ecosystem mediated by SQLE and identify tumoral CD8+ T cells as a critical immune component closely associated with SQLE expression. Both terbinafine-inhibited and genetically silenced SQLE in HCC cells promoted CD8+ T cell proliferation and activation via enhanced T cell receptor (TCR) signaling. Through conditional screening, metabolomic profiling, and liquid chromatography-mass spectrometry (LC-MS) quantification, we discovered that oxysterol 27-hydroxycholesterol, produced by SQLE-expressing HCC cells, functions as a signaling metabolite that modulates CD8+ T cells by supporting cholesterol requirements during TCR signal activation. Significantly, the in vivo model underscores the necessity of tumor-infiltrating CD8+ T cells for the therapeutic efficacy of terbinafine and supports a promising therapeutic strategy combining terbinafine with anti-PD1. Conclusion: Our study indicates that targeting cancer SQLE can rescue CD8+ T cell immune function by overcoming cholesterol restrictions in the HCC tumor environment, and illustrates a promising strategy by targeting a metabolic vulnerability to strengthen the antitumor efficacy of immunotherapy in HCC. Citation Format: Shuang Qiao, Hao Zou, Jing Xu, Limin Zheng. Potentiating the antitumor response of CD8T cells by targeting SQLE-mediated cholesterol metabolism reprogramming in hepatocellular carcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr A022.