Abstract 3345: Enhancer of zeste homolog 2 (EZH2) promotes expansion of breast tumor initiation cells through activation of Notch1-jagged signaling

Abstract

Abstract EZH2 is an oncogene which has been recently shown to regulate the differentiation of tissue specific stem cells. However the role of EZH2 on mammary stem cells is unknown. Notch1 signaling has been implicated in cell-fate determination of human mammary stem cells. We hypothesized that EZH2 may contribute to initiation and expansion of mammary stem cells through activation of Notch1 signaling. Methods: We developed two cell culture models: an inducible EZH2 up regulation system in MCF10A benign mammary cells and a lentivirus-mediated shRNA interference targeting EZH2 in SUM149 breast cancer cells. Mammary stem cells were identified using ALDH1+ and CD44+/CD24- by FACS. The effect of EZH2 on the tumorigenic ability of ALDH-1 positive and negative SUM149 cells was studied in a xenograft mouse model. Results: We found preferential expression of EZH2 in the stem cell population of normal primary breast cells compared to non stem cells. In MCF10A cells, EZH2 overexpression, (EZ-OV), was predominantly increased in the ALDH-1+ population. When we tested the role of EZH2 in the stem cell population we found that EZH2-OV increased the stem cell population of MCF10A cells while EZH2-KD decreased it in SUM149 cells. Microarray analysis identified Notch1-Jagged1 signaling as the most significantly downregulated targets of EZH2 in ALDH+ vs. ALDH- SUM149 cells. Consistently, EZH2-OV in MCF10A and primary breast cells increased Notch1 mRNA specifically in the ALDH1+ population. EZH2-OV increased Notch1 protein level and promoter activity in MCF10A cells. Mechanistically, Notch1 inhibition by GSI reversed the effect of EZH2-OV in the stem cell population of MCF10A cells. Our in vivo studies demonstrated that EZH2 KD in SUM149 cells decreased the growth of ALDH-1+ cells in xenograft experiments. Consistently, EZH2 KD tumors exhibited decreased Notch1 and Jagged1 protein levels. Conclusion: Collectively, these results indicate that EZH2 regulates mammary stem cells and requires, at least in part the activation of Notch1 signaling. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3345. doi:10.1158/1538-7445.AM2011-3345