Abstract 5144: Epigenetic reprogramming by tumor-derived EZH2 gain of function mutants leads to aggressive 3D-cell morphologies in both epithelial and melanoma cells

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA In addition to numerous genetic changes underlying cellular transformation, cancer cells are also characterized by epigenetic changes that are likely to play important roles in disease progression. EZH2 is an epigenetic repressor that plays well-established roles in development. In addition to widespread overexpression in a variety of tumors, the discovery of gain of function (GOF) mutations of EZH2 in diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, and melanoma strongly suggests an important function for this histone methyltransferase in cancer. To ascertain the function of elevated EZH2 catalytic activity, we expressed either wild-type EZH2 (WT) or EZH2 GOF mutants in both non-tumorigenic (immortalized epithelial cells) and tumorigenic (melanoma cells) settings. In both systems, EZH2 GOF mutants greatly increased global levels of H3K27me3 and decreased H3K27me2 levels, similar to the epigenetic pattern seen in DLBCL cell lines with endogenous EZH2 GOF mutations. In epithelial cells, expression of an EZH2 GOF mutant caused striking changes in 3D-morphology and gene changes that are indicative of cells that have undergone an epithelial to mesenchymal transition. In the disease relevant melanoma cells, several distinct EZH2 GOF mutants (but not EZH2 WT) caused prominent branching morphology in 3D-culture. Interestingly, these GOF mutants did not affect 2D-cell morphology or proliferation of melanoma cells. Furthermore, catalytic inhibition of EZH2 GOF mutants with a commercially available tool compound attenuated the 3D-phenotype. Importantly, EZH2 inhibition in melanoma cells expressing an endogenous GOF mutation also caused similar changes in 3D-morphology. RNA-seq analysis revealed genes involved in processes such as cell adhesion and axonal guidance that were down-regulated by EZH2 GOF mutants. Finally, melanoma cells expressing ectopic EZH2 GOF mutants formed larger tumors than control cells in mouse xenograft studies. Collectively, these results suggest that EZH2 GOF mutants may alter the interaction of tumor cells with their microenvironment and in this way provide a selective advantage to such tumors. Citation Format: Robert A. Rollins, Anthony M. Barsotti, Michael Ryskin, Wenyan Zhong, Wei-Guo Zhang, Andreas Giannakou, Christine Loreth, Veronica Diesl, Maximillian T. Follettie, Jonathon Golas, Michelle Lee, Timothy Nichols, Conglin Fan, Gary Li, Stephen Dann, Paul A. Rejto, Kim T. Arndt, Dominique Verhelle. Epigenetic reprogramming by tumor-derived EZH2 gain of function mutants leads to aggressive 3D-cell morphologies in both epithelial and melanoma cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5144. doi:10.1158/1538-7445.AM2014-5144