Publication Date

2018-03-14

Availability

Embargoed

Embargo Period

2020-03-13

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Cancer Biology (Medicine)

Date of Defense

2018-03-12

First Committee Member

Kerry Burnstein

Second Committee Member

Sanoj Punnen

Third Committee Member

Priyamvada Rai

Fourth Committee Member

Enrique Mesri

Fifth Committee Member

Nagi Ayad

Abstract

Castration-resistant prostate cancer (CRPC) progresses rapidly and is incurable. Constitutively active androgen receptor splice variants, such as AR-V7, represent a well-established mechanism of therapeutic resistance and disease progression. These AR variants lack the carboxy-terminal ligand binding domain, the target of all approved drugs against AR. Consequently, a significant challenge is to identify novel therapeutic strategies that are effective against AR variant-expressing prostate tumors. In this work, we evaluated the possibility of targeting AR-V7 in PC and CRPC through two independent approaches. The first approach consisted of disrupting the interaction between AR-V7 and its coactivators as an exploitable PC vulnerability. Using mutational and biochemical studies, we identified the domain in Vav3 (an established AR coactivator) necessary and sufficient to interact with AR-V7. Expressing this Vav3 domain in CRPC cells in vitro disrupted the interaction of Vav3 (as well as other coactivators) with AR-V7 and the subsequent enhancement of AR-V7 activity. Disrupting these interactions decreased CRPC cell proliferation, increased apoptosis, and decreased migration. Second, as an indirect approach to target cells that depend on AR-V7, we used a multi-pronged, unbiased systems biology analysis to identify downstream AR-V7-regulated hub genes that both drive cancer progression and feed back to enhance AR-V7 activity. Such genes likely encode prognostic markers as well as potential therapeutic targets acting within the AR-V7 network. Our systems biology approach utilized: 1) clinically relevant gene sets upregulated during human PC progression obtained by Weighted Gene-Co-expression Network Analysis (WGCNA); 2) an AR-V7 functional interactome from a high-throughput synthetic genetic array (SGA) screening in the yeast, S. pombe; and 3) genes regulated by AR-V7 in CRPC cells. We identified seven genes that were upregulated in human PC, functionally interacted with AR-V7, and were AR-V7 targets in PC. This gene set, composed of cell cycle and mitosis-regulating genes, comprised a signature that strongly correlated with patient Gleason score and predicted disease-free survival in large independent PC patient cohorts. Individually depleting the expression of these genes decreased ligand-independent AR transcriptional activity concomitant with reduced CRPC cell proliferation. Because CRPC may exhibit “addiction” to AR-V7 activity through dependency on the seven inter-related genes that we identified, pharmacologically targeting proteins encoded by two genes in this gene set led to potent synergistic inhibition of CRPC cell proliferation. This study utilized two different approaches to disrupt AR-V7 oncogenic signaling. First, we demonstrated the potential therapeutic utility of inhibiting constitutively active AR-V signaling by disrupting coactivator binding; second, by utilizing an unbiased and novel gene discovery strategy to identify clinically-relevant AR-V7 interaction networks, forming the basis for both prognostic use and rational, combinatorial therapy for CRPC.

Keywords

prostate cancer; systems-biology; androgen receptor; AR-V7; coactivators; VAV3

Available for download on Friday, March 13, 2020

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