Tasks / Specific Aims

Task 1. To conduct clinical trials to determine the efficacy and dose-response of the pro-apoptotic effects of estrogen (DES) in patients following the failure of two successive antihormonal therapies.

a) Confirm the efficacy of standard high-dose DES therapy and then determine a minimal dose to induce tumor regression.

b) Apply serum and tumor markers of estrogen-induced apoptosis for clinical monitoring (Tasks 2, 3 and 4).

Task 2. To elucidate the molecular mechanism of E2-induced survival and apoptosis in breast cancer cells resistant to either SERMs or long-term estrogen deprivation.

a) Complete a series of experiments using sets of well defined breast cancer models of E2-induced survival and apoptosis in vivo and in vitro. For each model module we will generate samples for the analyses under Task 3 (GU) and Task 4 (TGen).

b) Confirm and validate developing pathways of E2-induced breast cancer cell survival and apoptosis.

c) Collaborate with Task Teams completing Tasks 3 and 4 to define gene targets or protein products for validation in clinical samples.

d) Evaluate commercially available markers of apoptosis and circulating HER2 in serum from Task 1. Coded samples will validate their usefulness in monitoring patient responses to estrogen treatment.

Task 3. To decipher cellular signaling pathways of E2-induced survival and apoptosis using proteomics.

a) Identify protein signatures that are hallmarks of estrogen-induced apoptosis in breast cancer cells. Cell lines in which estrogen is a survival factor will be used for comparison (from Task 2).

b) Identify protein signatures in tumor xenografts that respond to estrogen administration in vivo by undergoing apoptosis. Xenograft models for which estrogen is a survival factor will be used for comparison (from Task 2).

c) Identify serum signature profiles that would be indicators of tumors that would undergo apoptosis after estrogen administration. The first analysis will be performed on mouse serum and profiles obtained in xenograft-bearing mice (Task 2). Results will be compared with serum profiles of responding and non–responding patients’ serum obtained from multi-center clinical trials (Task 1).

d) Determine which functional alterations in critical signaling molecules obtained from proteomic analysis identify target pathways for therapy of breast cancer patients with hormone-resistant disease (Task 1).

e) Mesh proteomics and mRNA analysis to identify targets for future drug development (Task 4).

Task 4. To analyze E2-induced survival and apoptotic pathways using gene array and siRNAs

a) Catalogue the transcriptional response of in vivo and in vitro models of endocrine resistance using array-based expression profiling (from Task 2).

b) Identify gene regulatory networks discriminatory for pathways indicative of differential responses to E2 in each laboratory model (from Task 2).

c) Interrogate molecular pathways contributing to endocrine resistance using ultra high throughput (UHT) RNA interference (cells from Task 2).