CD49f and CD61 identify Her2/neu-induced mammary tumor-initiating cells that are potentially derived from luminal progenitors and maintained by the integrin–TGFβ signaling

Abstract

Human epidermal growth factor receptor 2 (HER2)/Neu is overexpressed in 20–30% of breast cancers and associated with aggressive phenotypes and poor prognosis. For deciphering the role of HER2/Neu in breast cancer, mouse mammary tumor virus (MMTV)-Her2/neu transgenic mice that develop mammary tumors resembling human HER2-subtype breast cancer have been established. Several recent studies have revealed that HER2/Neu is overexpressed in and regulates self renewal of breast tumor-initiating cells (TICs). However, in the MMTV-Her2/neu transgenic mouse model, the identity of TICs remains elusive, despite previous studies showing supportive evidence for existence of TICs in Her2/neu-induced mammary tumors. Through systematic screening and characterization, we identified that surface markers CD49f, CD61 and ESA were aberrantly overexpressed in Her2-overexpressing mammary tumor cells. Analysis of these markers and CD24 detected anomalous expansion of the luminal progenitor population in preneoplastic mammary glands of Her2/neu transgenic mice, indicating that aberrant luminal progenitors originated in Her2-induced mammary tumors. The combined markers, CD49f and CD61, further delineated the CD49fhighCD61high-sorted fraction as a TIC-enriched population, which displayed increased tumorsphere formation ability, enhanced tumorigenicity both in vitro and in vivo and drug resistance to pacitaxel and doxorubicin. Moreover, the TIC-enriched population manifested increased transforming growth factor-β (TGFβ) signaling and exhibited gene expression signatures of stemness, TGFβ signaling and epithelial-to-mesenchymal transition. Our findings that self-renewal and clonogenicity of TICs were suppressed by pharmacologically inhibiting the TGFβ signaling further indicate that the TGFβ pathway is vital for maintenance of the TIC population. Finally, we showed that the integrin-β3 (CD61) signaling pathway was required for sustaining active TGFβ signaling and self-renewal of TICs. We for the first time developed a technique to highly enrich TICs from mammary tumors of Her2/neu transgenic mice, unraveled their properties and identified the cooperative integrin-β3–TGFβ signaling axis as a potential therapeutic target for HER2-induced TICs.