Loss of amphiregulin reduces myoepithelial cell coverage of mammary ducts and alters breast tumor growth

Serena P H Mao,Scott I Abrams,Jeffrey E Segall,Wenjun Guo,Minji Park,Paraic A Kenny,Ramon M Cabrera,George S Karagiannis,John S Condeelis,Dietmar M W Zaiss,Maja H Oktay,John R Christin

doi:10.1186/s13058-018-1057-0

Abstract

BackgroundAmphiregulin (AREG), a ligand of the epidermal growth factor receptor, is not only essential for proper mammary ductal development, but also associated with breast cancer proliferation and growth. In the absence of AREG, mammary ductal growth is stunted and fails to expand. Furthermore, suppression of AREG expression in estrogen receptor-positive breast tumor cells inhibits in-vitro and in-vivo growth.MethodsWe crossed AREG-null (AREG−/−) mice with the murine luminal B breast cancer model, MMTV-PyMT (PyMT), to generate spontaneous breast tumors that lack AREG (AREG−/− PyMT). We evaluated tumor growth, cytokeratin-8 (K8)-positive luminal cells, cytokeratin-14 (K14)-positive myoepithelial cells, and expression of AREG, Ki67, and PyMT. Primary myoepithelial cells from nontumor-bearing AREG+/+ mice underwent fluorescence-activated cell sorting and were adapted to culture for in-vitro coculture studies with AT-3 cells, a cell line derived from C57Bl/6 PyMT mammary tumors.ResultsIntriguingly, PyMT-induced lesions progress more rapidly in AREG−/− mice than in AREG+/+ mice. Quantification of K8+ luminal and K14+ myoepithelial cells in non-PyMT AREG−/− mammary glands showed fewer K14+ cells and a thinner myoepithelial layer. Study of AT-3 cells indicated that coculture with myoepithelial cells or exposure to AREG, epidermal growth factor, or basic fibroblast growth factor can suppress PyMT expression. Late-stage AREG−/− PyMT tumors are significantly less solid in structure, with more areas of papillary and cystic growth. Papillary areas appear to be both less proliferative and less necrotic. In The Cancer Genome Atlas database, luminal-B invasive papillary carcinomas have lower AREG expression than luminal B invasive ductal carcinomas.ConclusionsOur study has revealed a previously unknown role of AREG in myoepithelial cell development and PyMT expression. AREG expression is essential for proper myoepithelial coverage of mammary ducts. Both AREG and myoepithelial cells can suppress PyMT expression. We find that lower AREG expression is associated with invasive papillary breast cancer in both the MMTV-PyMT model and human breast cancer.

Highlights

Amphiregulin (AREG), a ligand of the epidermal growth factor receptor, is essential for proper mammary ductal development, and associated with breast cancer proliferation and growth
Expansion and progression of tumorigenic lesions is accelerated in the absence of AREG We examined the role of AREG in breast cancer using the mammary tumor virus (MMTV)-PyMT (PyMT) model in AREG−/− mice
The lesions in AREG+/+ PyMT mice were found in distinct regions in the ductal tree while in AREG−/− PyMT mice much of the ductal tree appeared to convert into the growing lesion

Summary

Introduction

Amphiregulin (AREG), a ligand of the epidermal growth factor receptor, is essential for proper mammary ductal development, and associated with breast cancer proliferation and growth. When AREG-null epithelial cells are transplanted into a cleared mammary gland, regardless of EGFR status in the stroma, the resultant gland shows a lack of cytokeratin-14 (K14) protein, a marker for myoepithelial cells [13]. While it is unknown whether AREG supports development and maintenance of myoepithelial cells, some evidence suggests that under low EGFR signaling conditions, mammary stem cells (MaSCs) preferentially differentiate into luminal, not myoepithelial, cells [14]. It is possible that AREG is important for the expansion of the ductal tree, and for proper differentiation of epithelial progenitor cells into luminal and myoepithelial cells

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