The Journal of Steroid Biochemistry and Molecular Biology
Mechanisms of acquired resistance to endocrine therapy in hormone-dependent breast cancer cells
Introduction
The estrogen receptor (ER) is expressed in approximately two thirds of breast carcinomas. Estrogen plays a crucial role in promoting growth of ER positive breast cancer cells because removal of the ovaries in premenopausal patients causes objective tumor regression. Current therapeutic strategies involve blockade of the mitogenic effect of estrogen on breast carcinoma by drugs that inhibit ER activity or reduce estrogen biosynthesis. Tamoxifen was the first agent of targeted therapy for hormone-dependent breast cancer and remains the most commonly used agent in the adjuvant setting and for treatment of advanced disease. In addition to tamoxifen, postmenopausal patients benefit from aromatase inhibitors that reduce the levels of estrogen that are critical for tumor growth. Recent clinical trials suggest that third generation aromatase inhibitors are superior to tamoxifen as first line therapy and are effective in patients with disease relapse after tamoxifen treatment [1], [2]. While patients with hormone-dependent breast cancer initially experience objective tumor regression, the disease ultimately relapses. Preclinical and clinical studies have revealed that up-regulation of growth factor signaling pathways is associated with failure of endocrine therapy and targeting the growth factor signaling pathway has become an emerging therapeutic strategy for treatment of breast cancer.
In most cases, there is no loss of ERα when resistance to endocrine therapy develops. The receptor is apparently functioning because a portion of patients with relapsing disease are responsive to secondary endocrine therapies [3], [4]. Increased ER functionality has been found in breast cancer cells that become resistant to manipulations blocking estrogen action [5], [6]. This has been largely attributed to enhanced transcriptional activity of ERα by cross-talk with up-regulated growth factor pathways [7], [8], [9]. However, the process of adaptation to endocrine therapy varies in hormone-dependent breast cancer cells and the precise mechanisms are far from fully understood.
To investigate the mechanisms underlying failure of endocrine therapy, our laboratory has established two in vitro models using MCF-7 cells. Long-term culture of MCF-7 cells in estrogen deprived medium (LTED) mimics aromatase inhibition in patients. Continued exposure of MCF-7 cells to tamoxifen represents a model of acquired resistance to antiestrogens (TAM-R). We found that MCF-7 cells react differently to these two endocrine manipulations. Here we report possible mechanisms that are involved in the process of adaptation to endocrine therapy and re-growth of hormone dependent breast cancer.
Section snippets
Materials
Farnesylthiosalicylic acid (FTS) was a gift from Drs. Yoel Kloog (Tel-Aviv University, Tel-Aviv, Israel) and Wayne Bardin (Thyreos, New York, NY). Estradiol was from Steraloids (Wilton, NH). LY 294002 and rapamycin were purchased from Sigma (St. Louis, MO). U0126 was obtained from Promega (Madison, WI). c-Src kinase inhibitor PP2 was from Calbiochem (San Diego, CA). EGF was obtained from Collaborative Biomedical Products (Bedford, MA). Sources of antibodies for Western analysis are as follows:
Results
We have demonstrated in previous studies that enhanced activity of growth factor pathways in LTED cells renders the cells hypersensitive to the mitogenic effect of estradiol and dual blockade of the MAPK and PI3K pathways reverses the hypersensitive phenotype [10]. We proposed that inhibition of signal transduction at a nodal point where multiple pathways converge would provide an effective strategy for treatment of breast cancer with acquired endocrine resistance. Farnesylthiosalicylic acid
Discussion
The present study demonstrated that different mechanisms are involved when breast cancer cells adapt themselves to escape from the manipulations blocking the estrogen receptor signaling. Long-term estrogen deprivation results in sustained activation of both the MAPK and the mTOR pathways in MCF-7 cells. Using FTS as a tool, we found that the mTOR pathway is a predominant pathway in mediation of proliferation of LTED cells. The specific mTOR inhibitor, rapamycin, effectively inhibited growth of
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