Abstract 6108: In vivo CRISPR screens identified dual function of MEN1-MLL1 in regulating tumor-microenvironment interactions

Abstract

Abstract INTRODUCTION: CRISPR functional genomic screens have been widely adopted to identify essential genes and potential drug targets in cell line models. However, it is well known that cell line models studied in vitro do not fully capture the biology in patient tumors due to the lack of the tumor microenvironment. The primary objective of this study was to perform functional genomic screens in in vivo models to identify clinically relevant epigenetic vulnerabilities. METHODS: We designed an EpiDrug sgRNA library that target 357 epigenetic regulators in human, and applied this targeted sgRNA library for essentiality screen in 2D cell culture and 3D xenograft models. We subsequently selected targets that can only be identified in vivo for validation. We also investigated the underlying mechanism of the in vivo specific phenotype caused by target knockouts. Finally, we explored the clinical significance of treating selected target with a small molecule inhibitor. RESULTS: We identified MEN1 as the top hit that confers differential essentialities between in vitro and in vivo models. Knockout of MEN1 has no impact on cell proliferation in 2D cell culture but profoundly promotes tumor growth in human lung and colorectal adenocarcinoma cell line derived xenograft models. In syngeneic colon cancer model CT26, knockout of Men1 resulted in faster tumor growth in immune deficient mice, but reduced tumor growth in immune competent mice. Mechanistically, knockout of MEN1/Men1 can cause the redistribution of its interaction partner MLL1/Mll1, a histone methyl transferase, to repeat regions in chromatin and stimulate the expression of double stranded RNA. This resulted in viral mimicry response, which activates a group of genes involved in cytokine-cytokine receptor interaction. Single-cell RNA-seq and CyTOF analysis revealed that activation of the cytokines induces tumor promoting neutrophil and tumor suppressing CD8+ T cell infiltration in immunodeficient and immunocompetent mice, respectively. Using patient data from TCGA, we identified that the expression level of MEN1 is negatively correlated with neutrophil and CD8+ T cell tumor infiltration in most cancer types including lung and colorectal adenocarcinoma. A small molecular inhibitor targeting Men1-Mll1 interaction dramatically reduced CT26 tumor growth in immunocompetent mice, but its phenotype was reversed by CD8 neutralizing antibody. Finally, we demonstrated the strong synergy of the inhibitor with anti-PD-L1 antibody. CONCLUSION: Our study demonstrated the utility of in vivo CRISPR screen in identifying therapeutic targets that modulate tumor-microenvironment interactions, and identified MEN1 as a promising therapeutic targets alone or in combination with immunotherapy. Citation Format: Peiran Su, Yin Liu, Ming-Sound Tsao, Housheng H. He. In vivo CRISPR screens identified dual function of MEN1-MLL1 in regulating tumor-microenvironment interactions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6108.