Acquisition of epithelial-mesenchymal transition phenotype in the tamoxifen-resistant breast cancer cell: a new role for G protein-coupled estrogen receptor in mediating tamoxifen resistance through cancer-associated fibroblast-derived fibronectin and β1-integrin signaling pathway in tumor cells

Abstract

IntroductionAcquired tamoxifen resistance remains the major obstacle to breast cancer endocrine therapy. β1-integrin was identified as one of the target genes of G protein-coupled estrogen receptor (GPER), a novel estrogen receptor recognized as an initiator of tamoxifen resistance. Here, we investigated the role of β1-integrin in GPER-mediated tamoxifen resistance in breast cancer.MethodsThe expression of β1-integrin and biomarkers of epithelial-mesenchymal transition were evaluated immunohistochemically in 53 specimens of metastases and paired primary tumors. The function of β1-integrin was investigated in tamoxifen-resistant (MCF-7R) subclones, derived from parental MCF-7 cells, and MCF-7R β1-integrin-silenced subclones in MTT and Transwell assays. Involved signaling pathways were identified using specific inhibitors and Western blotting analysis.ResultsGPER, β1-integrin and mesenchymal biomarkers (vimentin and fibronectin) expression in metastases increased compared to the corresponding primary tumors; a close expression pattern of β1-integrin and GPER were in metastases. Increased β1-integrin expression was also confirmed in MCF-7R cells compared with MCF-7 cells. This upregulation of β1-integrin was induced by agonists of GPER and blocked by both antagonist and knockdown of it in MCF-7R cells. Moreover, the epidermal growth factor receptor/extracellular regulated protein kinase (EGFR/ERK) signaling pathway was involved in this transcriptional regulation since specific inhibitors of these kinases also reduced the GPER-induced upregulation of β1-integrin. Interestingly, silencing of β1-integrin partially rescued the sensitivity of MCF-7R cells to tamoxifen and the α5β1-integrin subunit is probably responsible for this phenomenon. Importantly, the cell migration and epithelial-mesenchymal transition induced by cancer-associated fibroblasts, or the product of cancer-associated fibroblasts, fibronectin, were reduced by knockdown of β1-integrin in MCF-7R cells. In addition, the downstream kinases of β1-integrin including focal adhesion kinase, Src and AKT were activated in MCF-7R cells and may be involved in the interaction between cancer cells and cancer-associated fibroblasts.ConclusionsGPER/EGFR/ERK signaling upregulates β1-integrin expression and activates downstream kinases, which contributes to cancer-associated fibroblast-induced cell migration and epithelial-mesenchymal transition, in MCF-7R cells. GPER probably contributes to tamoxifen resistance via interaction with the tumor microenvironment in a β1-integrin-dependent pattern. Thus, β1-integrin may be a potential target to improve anti-hormone therapy responses in breast cancer patients.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-015-0579-y) contains supplementary material, which is available to authorized users.

The Widening Sphere of Influence of HOXB7 in Solid Tumors

“…Despite major advances in breast cancer treatment and management, and successful treatment of ER-positive breast cancer with tamoxifen and aromatase inhibitors, one third of women with estrogen receptorα (ER)-positive breast cancer suffer from recurrent disease within fifteen years (33). As EMT has been observed in tamoxifen (TAM)-resistant MCF-7 breast cancer cells (30,34,35) and given the association between HOXB7 and EMT in breast cancer (11), HOXB7 protein might possibly play a role in tamoxifen resistance and ensuing tumor progression. Evidence supporting this hypothesis was elucidated by Jin and collaborators through work that demonstrated that HOXB7 overexpression conferred TAM-resistance to ER+ breast cancer cells through the HOXB7-dependent transcriptional induction of EGFR (3,11).…”

Tumor microenvironment and cancer therapy resistance

“…In tamoxifen-resistant breast tumors, G protein-coupled estrogen receptor (GPER)/epidermal growth factor receptor (EGFR)/ extracellular regulated protein kinase (ERK) signaling enhances β1-integrin expression and activates downstream kinases, contributing to CAF-induced cell migration [17]. Moreover, downstream kinases of β1-integrin including focal adhesion kinase, Src and AKT are activated in resistant cells, potentially involved in the interaction between cancer cells and CAFs [17], highlighting the persistent tumor-stroma communication in a biologically dynamic TME.…”

“…Moreover, downstream kinases of β1-integrin including focal adhesion kinase, Src and AKT are activated in resistant cells, potentially involved in the interaction between cancer cells and CAFs [17], highlighting the persistent tumor-stroma communication in a biologically dynamic TME. More importantly, CAFs establish a synergistic relationship with cancer cells, contributing to their malignancy and therapeutic resistance.…”

Acquired Resistance of ER-Positive Breast Cancer to Endocrine Treatment Confers an Adaptive Sensitivity to TRAIL through Posttranslational Downregulation of c-FLIP

“…The development of endocrine therapy resistance in primary breast cancer is associated with poor prognosis and, in cell line models and clinical samples of endocrine resistance, a transition to more aggressive tumor phenotypes that have undergone partial EMT (14)(15)(16)(17). The cancer therapeutic TRAIL has been shown to preferentially induce cell death in breast cancer cell lines that exhibit a more mesenchymal phenotype (35).…”

“…Thus, the majority of patients with advanced disease and up to 30% of ERþve patients in the adjuvant setting will acquire resistance to the inhibitory effects of their endocrine treatment (12,13). The acquisition of resistance manifests with a partial epithelial-mesenchymal transition (EMT) and increased migratory and invasive activity both in anti-estrogen-resistant cell line models (14) and in clinical samples (14)(15)(16)(17). Furthermore, a subset of tumor-initiating cells with stem-like characteristics (cancer stem cells: CSCs) is enriched in tamoxifen-resistant breast cancers and may demonstrate resistance to endocrine therapies, thus contributing to disease recurrence and metastatic progression (18)(19)(20)(21).…”