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ErbB receptors and resistance to aromatase inhibitors in breast cancer
Background. A large body of experimental data suggests a causal association between overexpression and aberrant activity of the HER2/neu (erbB2) signaling pathway and antiestrogen resistance in human breast cancer. The HER2 receptor is the protein product of the HER2 proto-oncogene and a member of the EGF family of erbB receptor tyrosine kinases, which also includes HER3 and HER4. Upon binding of ligand to the EGF receptor (EGFR), HER3, or HER4, HER2 is recruited as the preferred partner of these ligand-bound receptors into a kinase-active, phosphorylated heterodimeric complex that activates several signaling pathways leading to enhanced proliferation and survival of tumor cells. HER2 is capable of transforming normal mammary epithelial cells and is overexpressed in a cohort of breast tumors where it is associated with a more metastatic behavior and poor patient prognosis. Patients with tumors that overexpress HER2 also exhibit statistically lower responses and or shorter duration of response to antiestrogen therapy. The EGFR is another member of this receptor family, initially discovered as the proto-oncogene of the mutant, constitutively active oncogenic v-erbB tyrosine kinase which induces avian erythroblastosis. Overexpression of EGFR and its ligands is seen in several human cancers and this is associated with increased tumor progression as well as antiestrogen resistance in breast cancer. Taken together, these data suggest that blockade of aberrant EGFR/HER2 signaling in human breast cancer cells may delay or reverse the emergence of resistance to antiestrogen therapies including aromatase inhibitors.
Objective/Hypothesis. Therefore, this proposal will test the hypothesis that overexpression of HER2 or its mouse homolog neu are causally associated with resistance to aromatase inhibitors in mammary cancer cells. A second objective will be the demonstration that chronic blockade of EGFR/HER2 signaling will prevent or delay the emergence of resistance to aromatase inhibitors. The following aims will be pursued:
Specific Aim 1: To determine whether blockade of EGFR/HER2 signaling prevents the emergence of acquired resistance to the aromatase inhibitor letrozole in human breast cancer cells.
Specific Aim 2: To determine if forced expression of HER2 results in de novo resistance to letrozole in MCF-7 cells stably transfected with aromatase gene (MCF-7/CA).
Specific Aim 3: To engineer MMTV/aromatase mice and determine if MMTV/aromatase x neu bigenic mice are resistant to the ability of letrozole to prevent aromatase-induced mammary ductal hyperplasias in MMTV/aromatase mice.
Study Design: 1. MCF-7/CA cells or xenografts stably expressing the aromatase gene will be grown in the presence of androstenedione (AD) „b letrozole. Cells or xenografts escaping letrozole-induced growth inhibition will be examined for levels of HER2, EGFR, and post-receptor signal transducers. Subsequently, letrozole-treated tumors will be treated concomitantly with the EGFR/HER2 inhibitor OSI-774 (Tarceva) to determine if blockade of erbB receptor signaling delays or prevents antiestrogen resistance.
2. MCF-7/CA cells with be stably transduced with pBabe-erbB2 (HER2) or pBabe retroviral vectors. Letrozole resistance will be documented by AD-induced reporter activity in cells transiently transfected with ERE-luciferase. MCF-7/CA/HER2 xenografts will be established in athymic mice and examined for sensitivity to letrozole „b OSI-774 by tumor size, BrdU incorporation, and TUNEL analysis.
3. MMTV/aromatase mice will be cross-bred with MMTV/neu transgenic mice. In bigenic mice we will determine whether a) neu accelerates aromatase-induced hyperplasias; b) neu and aromatase synergize in the induction of mammary tumors, and 3) bigenic tumors are resistant to letrozole compared to MMTV/aromatase mice, and d) pharmacological blockade of HER2 restores sensitivity of the bigenic tumors to letrozole.
Potential Outcomes and Benefits. This project may address for the first time if overexpression of HER2/neu (and/or EGFR) can confer resistance to aromatase inhibitors in breast cancer cells. Such results will lay the foundation for novel clinical trials with aromatase inhibitors in combination with ERGR/HER2 signaling inhibitors, currently in clinical development, in patients with hormone-dependent breast cancer.
Most breast cancers are estrogen-dependent and the majority of these tumors respond to an initial therapy with antiestrogens. However, breast cancers that initially respond to antiestrogens, almost invariably tend to become resistant to these hormonal therapies over time. Over the last few years, several studies have identified the EGF receptor (EGFR, erbB1) and HER2/neu (erbB2) as signaling molecules that can mediate early or late resistance to antiestrogens. Currently, several drugs that block the function of the EGFR and HER2 have become available, creating the exciting possibility of novel therapeutic combinations of EGFR/HER2 inhibitors with established antiestrogen approaches like aromatase inhibitors. However, formal proof that excessive signaling by the EGFR and/or HER2 mediates resistance to aromatase inhibitors is lacking. Therefore, in these studies we propose to determine if overexpression of EGFR and/or HER2 results in de novo or acquired resistance to the aromatase inhibitor letrozole, an approved and increasingly used for hormone-dependent breast cancers. The results from these studies will have significant clinical implications that may lead to novel treatment combinations of letrozole and other antiestrogens with inhibitors of EGFR/HER2. In turn, these approaches may increase the efficacy of current antiestrogen therapies and delay the emergence of resistance to aromatase inhibitors, thus improving the outcome and quality of life of patients with hormone-dependent breast cancer.