Breast cancer is the second most common cause of cancer-related deaths for women in the United States, and the mortality for those 20% of patients with recurrences and/or metastases is nearly 100%. EZH2 (Enhancer of Zeste Homolog 2), a Polycomb group protein that functions in transcriptional memory, is upregulated during progression from ductal carcinoma in situ to invasive carcinoma and distant metastasis. EZH2 protein is overexpressed in 55% of invasive breast carcinomas and women with tumors expressing high EZH2 have a worse disease free and overall survival than women with tumors expressing low EZH2. We hypothesize that EZH2 overexpression may give rise to breast cancers that are on a highly aggressive path from the outset and may enhance metastasis. To test our hypothesis, we have investigated the role of EZH2 in cell morphology, invasion and motility in vitro, and metastasis in vivo. Stable shRNA EZH2 knockdown was carried out in 3 breast cancer cell lines (MDA-MB-231, CAL51, SUM149) which normally express high levels of EZH2 and have motile and invasive abilities. EZH2 knockdown in all cell lines tested decreased their invasiveness using a modified Boyden chamber assay. Random motion motility assays using live cell imaging tracking on MDA-MB-231 vector and shEZH2 cells revealed that EZH2 knockdown significantly reduces the random motion average cell velocity of these cells. Concomitant with the effect of EZH2 knockdown in decreasing invasion and velocity of movement of breast cancer cells, we determined that EZH2 knockdown in MDA-MB-231 and SUM149 cells results in a morphological change and protein expression pattern consistent with a mesenchymal to epithelial transition (MET). Recent experiments have demonstrated that EZH2 knockdown in breast cancer cell lines decreases the levels of activated, phosphorylated p38 mitogen activated protein kinase (MAPK), a protein shown to regulate breast cancer cell motility and metastasis. The in vivo relevance is highlighted by our data showing that EZH2 knockdown in MDA-MB-231 cells significantly decreased the number of lung metastases per mouse, decreased the average size of lung metastases and decreased the proliferative index of the metastatic deposit as measured by decreased Ki67 immunostaining, when compared to controls. Decreased phosphorylated p38 staining has also been verified in the mouse xenograft lung metastases. Taken together, these data support a role for EZH2 in establishing rapid cell motion, invasion and metastasis in breast cancers. Experiments to further delineate the underlying mechanism and to determine the role of p38 in EZH2-mediated functions are underway.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5309. doi:1538-7445.AM2012-5309