Abstract 2223: Rapamycin suppresses ErbB2-overexpressing mammary tumors through selective inhibition of luminal progenitor cells and tumor-initiating cells
Authors: Zhao Qingxia , Parris Amanda B. , Howard Erin W. , Zhao Ming , Guo Zhiying , Xing Ying , Ma Zhikun , Yang Xiaohe 
Published: July 2017

Abstract
Rapamycin is a potent inhibitor of the mTOR pathway and has demonstrated anti-cancer properties in various cancer models. Although rapamycin was reported to inhibit mammary carcinogenesis at multiple stages, including initiation, invasion, and metastasis, the underlying mechanisms remain unclear, especially in individual breast cancer subtypes. Specifically, ErbB2/Her2 amplification occurs in approximately 25% of invasive human breast cancers, and is characterized by Akt-mTOR pathway deregulation. Recent reports also suggest that cancer stem cells (CSCs) play a critical role in tumor development and metastasis. Therefore, we investigated the efficacy of rapamycin in the prevention of mammary tumor development in MMTV-ErbB2 transgenic mice by focusing on its effects on CSCs/tumor-initiating cells (TICs). In this study, MMTV-ErbB2 mice were inoculated with syngeneic 78617 cells, which are derived from MMTV-ErbB2 mammary tumors, followed by rapamycin treatment (1.5 mg/kg/day) for 12 days. We found that rapamycin elicited strong anti-tumorigenic effects on the syngeneic tumors, as indicated by substantially reduced tumor volume and weight. To determine the effect of rapamycin on mammary epithelial cells (MECs) in premalignant tissues, MMTV-ErbB2 mice were treated with low-dose rapamycin (1 mg/kg/day) between weeks 10 and 20 of age. Rapamycin significantly inhibited mammary morphogenesis at 20 weeks of age as compared to the control mice. As such, the branching density, ductal elongation, and proliferative index of the mammary glands from the rapamycin-treated mice were markedly decreased. Flow cytometric analysis of primary MECs demonstrated that rapamycin selectively inhibited the CD61highCD49fhigh subpopulation, which are enriched with luminal progenitor cells and TICs. 3D culture of FACS-sorted MEC subpopulations indicated that CD61highCD49fhigh cells are the major subpopulation that gives rise to 3D colonies. We further demonstrated that rapamycin significantly suppressed colony-forming cell (CFC) number and primary/secondary sphere formation of primary MECs and spontaneous MMTV-ErbB2 tumor cells, indicating rapamycin-induced inhibition of mammary progenitor cells and CSC self-renewal, respectively. Furthermore, molecular analysis of MECs demonstrated that rapamycin inhibited signaling associated with ER, ErbB2, and mTOR. In particular, rapamycin strikingly inhibited Wnt/β-catenin and TGFβ/Smad3 signaling. Taken together, our results demonstrate that rapamycin-mediated prevention of MMTV-ErbB2 mammary tumor development involves selective targeting of mammary progenitor cells and potential TICs in the premalignant mammary tissues through regulation of MEC stemness and multiple signaling pathways, which contributes to its potent anti-tumor effects on ErbB2-overexpressing breast cancer.
Citation Format: Qingxia Zhao, Amanda B. Parris, Erin W. Howard, Ming Zhao, Zhiying Guo, Ying Xing, Zhikun Ma, Xiaohe Yang. Rapamycin suppresses ErbB2-overexpressing mammary tumors through selective inhibition of luminal progenitor cells and tumor-initiating cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2223. doi:10.1158/1538-7445.AM2017-2223