Abstract 260: Depletion of Ezh2 in the intestinal adenoma induced by JAK2V617F mutation leads to the malignant neoplasm

Abstract

Abstract Background The increased JAK2 and EZH2 plays a critical role in the colorectal or colon cancer (CRC) progression. Targeting aberrant JAK2 expression caused by JAK2 gain-of-function (GOF) mutation, JAK2V617F and deregulated EZH2 kinase activity, JAK2 and/or EZH2 inhibitors have been used to treat the advanced CRCs. However, it is unclear whether the aberrant JAK2 can directly induce CRC or intestinal epithelial (IEC) malignancy. We hypothesize that the overexpressed JAK2 driven by JAK2V617F GOF mutation increases IEC neoplasia in which depletion of EZH2 can lead to IEC malignant neoplasm. Materials and Methods Colorectal samples were collected from 20 cases of CRC patients. Immunohistochemistry staining (IH), confocal immunofluorescence staining (IF) and bulk RNAseq were used to determine LGR5, Ki67, JAK2, EZH2, mutant β-catenin and JAK2V617F GOF mutation. Crypt-derived organoids or tumoroids were used to determine the JAK2 and EZH2 in the proliferative LGR5 cells (Ki67hiLGR5+) and were treated with JAK2 and/or EZH2 inhibitors. The mice with JAK2 and/or EZH2 deficiency in IECs were generated by crossing VilCreER mice, floxed-Jak2 and/or floxed-Ezh2 mice to determine the intestinal and colonic neoplasia. Lgr5, Ki67, Cdx2 Jak2, Ezh2, and mutant β-catenin IH or IF, organoids culture and peritoneal xenograft were used to determine the malignant neoplasm. The JAK2V617F-transfected HT29 or Caco-2 cells were used to determine the migration induced by JAK2V617F GOF mutation, then treated with EZH2 inhibitors to determine the effects of EZH2 inhibition on JAK2V617F GOF mutation-induced migration. The spheres were formed with Caco-cells with JAK2V617F GOF mutation and were used to observe the effects of JAK2V617F GOF mutation on IEC neoplasia formation. Results JAK2 and/or EZH2 overexpression are positively correlated with the probability of CRC initiation. JAK2 and EZH2 are highly expressed in the CRC tumoroids. Inhibition of EZH2 increases the CRC tumoroids growth and spheroid formation, and reduces budding numbers while reducing the proliferation of organoids from healthy crypts. Jak2 over-expression in IECs increases intestinal microadenoma, and depletion of EZH2 in the adenoma increases mutant β-catenin and severity of intestinal adenoma, suggesting that depletion of EZH2 enhances the malignancy of overexpressed Jak2-derived IEC neoplasia. Consistently, JAK2V617F GOF mutation increases HT29 or Caco-2 cell migration. Conclusion JAK2V617F GOF mutation can drive IEC neoplastic proliferation and adenoma, depletion of EZH2 in the adenoma leads to IEC malignant neoplasm. Our research suggests that loss of Ezh2 may synergistically cooperate with the Jak2V617F mutation in the pathogenesis of IEC malignant neoplasm. Citation Format: Nardana Esmaeili, Ahmed Bakheet, Wen Gao, Haifeng Li, Dan Cai, Xiaonan Han. Depletion of Ezh2 in the intestinal adenoma induced by JAK2V617F mutation leads to the malignant neoplasm [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 260.