Faculty Bibliography

To gain insight into cellular factors regulating AR action that could promote castration resistant prostate cancer (CRPC), we performed a genome-wide RNAi screen for factors that promote ligand-independent AR transcriptional activity and integrated clinical databases for candidate genes that are positively associated with prostate cancer metastasis and recurrence. From this analysis, we identified Dynein Axonemal Heavy Chain 8 (DNAH8) as an AR regulator that displayed higher mRNA expression in metastatic than in primary tumors, and showed high expression in patients with poor prognosis. Axonemal dyneins function in cellular motility, but the function of DNAH8 in prostate cancer or other cell types has not been reported. DNAH8 is on chromosome 6q21.2, a cancer-associated amplicon, and is primarily expressed in prostate and testis. Its expression is higher in primary tumors compared to normal prostate, and is further increased in metastatic prostate cancers. Patients expressing high levels of DNAH8 have a greater risk of relapse and a poor prognosis after prostatectomy. Depletion of DNAH8 in prostate cancer cells suppressed AR transcriptional activity and proliferation. Androgen treatment increased DNAH8 mRNA expression, and AR bound the DNAH8 promoter sequence indicating DNAH8 is an AR target gene. Thus, DNAH8 is a new regulator of AR associated with metastatic tumors and poor prognosis.

Despite decades of effort to develop effective therapy and to identify promising new drugs, prostate cancer is lethal once it progresses to castration-resistant disease. Studies show mis-regulation of multiple pathways in castration-resistant prostate cancer (CRPC), reflecting the heterogeneity of the tumors and also hinting that targeting androgen receptor (AR) pathway alone might not be sufficient to treat CRPC. In this study, we present evidence that the Wnt/beta-catenin pathway might be activated in prostate cancer cells after androgen-deprivation to promote androgen-independent growth, partly through enhanced interaction of beta-catenin with TCF4. Androgen-independent prostate cancer cells were more prone to activate a Wnt-reporter, and inhibition of the Wnt/beta-catenin pathway increased sensitivity of these cells to the second-generation antiandrogen, enzalutamide. Combined treatment of enzalutamide and Wnt/beta-catenin inhibitor showed increased growth repression in both androgen-dependent and -independent prostate cancer cells, suggesting therapeutic potential for this approach.

Uterine carcinosarcoma (UCS) is a rare type of cancer and accounts for 5% of uterine malignancies. However, UCS patients suffer a high prevalence of chemo-resistance and a very poor prognosis compared to uterine cancer patients. URI is a chaperone with functions in transcription. We analyzed the somatic URI1 copy number variation in 57 post-menopausal non-metastatic UCS patients in comparison to 363 uterine corpus endometrial carcinomas. URI1 amplification was detected in 40% (23/57) of primary UCS and 5.5% (20/363) of uterine carcinomas. UCS patients with URI1 amplification exhibited 13% (3/23) tumor-free survival compared to 41% (14/34) in the absence of URI amplification (P=0.023). URI1 amplification (OR=6.54, P=0.027), weight (OR=1.068, P=0.024), hypertension (OR=3.35, P=0.044), and tumor stage (OR=2.358, P=0.018) associated with poor survival. Patients treated with hormone replacement therapy (OR=15.87, P=0.011) displayed enhanced overall survival. Combined radiation and chemotherapy improved patient survival (median survival=2043 days) compared to single (median survival=597 days) or no treatment (median survival=317 days, P=0.0016). Importantly, patients with URI1 amplification had poor response to adjuvant treatment compared to control group (P=0.013). Tumors with URI1 amplification displayed decreased transcription of genes encoding tumor suppressor and apoptotic regulators and increased expression of genes regulating oncogenesis, survival and metastasis. Overexpression of URI1 in a cultured cell model induced ATM expression and resistance to cisplatin. Our findings suggest that high prevalence in UCS may associate with poor prognosis and worse response to adjuvant treatment.

It is well documented that androgen receptor (AR), a steroid hormone receptor, is important for prostate cancer (PCa) growth. Conversely, however, there is increasing evidence that activation of AR by androgens can also lead to growth suppression in prostate cells. AR mediated transcription is regulated by a number of different transcriptional coactivators. Changes in expression level or cellular localization of specific coactivators may play a crucial role in this switch between proliferative and anti- proliferative processes regulated by AR target gene programs. In this review, we discuss the expression and function of several AR coactivators exhibiting growth suppressive function in PCa, including ARA70/ELE1/NCOA4, androgen receptor coactivator p44/MEP50/WDR77, TBLR1, and ART-27. In luciferase reporter assays, they all have been shown to activate AR mediated transcriptional activation. ARA70 exists in two forms, the full length nuclear ARA70alpha and internally spliced cytoplasmic ARA70beta. For p44 and TBLR1, we identified nuclear and cytoplasmic forms with distinct expression and function. In comparison of their expression (ARA70alpha, p44, TBLR1 and ART-27) in prostate, these coactivators are expressed in the nucleus of benign prostate epithelial cells while they are more predominantly expressed in cytoplasmic form (ARA70beta, cytoplasmic p44 and TBLR1) in PCa. Consistent with their nuclear expression in benign prostate, the nuclear form of these coactivators inhibit PCa growth targeting a subset of AR target genes. In contrast, the cytoplasmic versions of these proteins enhance PCa growth and invasion. Interestingly, first characterized as an AR coactivator in luciferase assays, ART-27 functions as corepressor for endogenous AR target genes. Importantly, the growth inhibitions by these nuclear proteins are androgen-dependent processes and the regulation of invasion is androgen-independent. Understanding the molecular switches involved in the transition from AR dependent growth promotion to growth suppression and dysregulation of these coactivator proteins promoting androgen-independent invasion may lead to identification of novel therapeutic targets for PCa.

Androgen receptor (AR), a steroid hormone receptor, is critical for prostate cancer growth. However, activation of AR by androgens can also lead to growth suppression and differentiation. Transcriptional cofactors play an important role in this switch between proliferative and anti-proliferative AR target gene programs. Transducin beta-like-related protein 1 (TBLR1), a core component of the nuclear receptor corepressor complex, shows both corepressor and coactivator activities on nuclear receptors, but little is known about its effects on AR and prostate cancer. We characterized TBLR1 as a coactivator of AR in prostate cancer cells and determined that the activation is dependent on both phosphorylation and 19S proteosome. We showed that TBLR1 physically interacts with AR and directly occupies the androgen-response elements of the affected AR target genes in an androgen-dependent manner. TBLR1 is primarily localized in the nucleus in benign prostate cells and nuclear expression is significantly reduced in prostate cancer cells in culture. Similarly, in human tumor samples, the expression of TBLR1 in the nucleus is significantly reduced in the malignant glands compared with the surrounding benign prostatic glands (P<0.005). Stable ectopic expression of nuclear TBLR1 leads to androgen-dependent growth suppression of prostate cancer cells in vitro and in vivo by selective activation of androgen-regulated genes associated with differentiation (e.g. KRT18) and growth suppression (e.g. NKX3-1), but not cell proliferation of the prostate cancer. Understanding the molecular switches involved in the transition from AR-dependent growth promotion to AR-dependent growth suppression will lead to more successful treatments for prostate cancer.

Chronic inflammation has been linked to cancer initiation and progression in a variety of tissues, yet the impact of acute and chronic inflammatory signaling on androgen receptor function has not been widely studied. In this report, we examine the impact of the inflammation-linked cytokine, interleukin-1beta on androgen receptor function in prostate cancer cells. We demonstrate that acute interleukin-1beta treatment inhibits the transcription of the androgen receptor gene itself, resulting in the reduction of androgen receptor protein levels. Interestingly, in cells subjected to chronic interleukin-1beta stimulation, the transcription of the androgen receptor gene is restored within a few cell passages and the cells acquire the ability to grow in the presence of the anti-androgen, bicalutamide. Importantly, the changes that accompany this loss of androgen receptor regulation and gain of anti-androgen resistance are stably heritable since once established, the phenotype is maintained even in the absence of exogenously added interleukin-1beta. Further, bicalutamide resistance correlates with increased transcription of androgen receptor target genes and histone H3K4 dimethylation at M-phase gene enhancers. Overall, our studies demonstrate a novel route to anti-androgen resistance upon exposure to an inflammatory cytokine and provide a new tool to further understand how anti-androgen resistance emerges under chronic inflammation.

Androgen receptor (AR) plays an important role in the tumorigenesis and progression of prostate cancer (PCa), and is the primary therapeutic target for PCa treatment. AR activity can be regulated via phosphorylation at multiple phosphorylation sites within the protein. Modifications by phosphorylation alter AR function, including its cellular localization, stability and transcriptional activity, ultimately leading to changes in cancer cell biology and disease progression. Here we present a brief overview of AR phosphorylation sites in PCa, focusing on functional roles of phospho-AR (p-AR) species, relevance in PCa disease progression, and potential as biomarkers and/or therapeutic targets through the use of kinase inhibitors. Additionally, recent evidence has shown the important role of AR activity in the cancer associated stroma on PCa growth and progression. The phosphorylation status of epithelial and stromal AR may be distinct; however, the current data available on stromal AR phosphorylation is limited. Further research will determine global view on the synergistic effects of phosphorylation across multiple AR sites in both epithelial and stromal cells and validate whether together they can be used as prognostic markers and/or effective therapeutic targets for PCa.

Prostate cancer (PCa) is the second leading cause of cancer-related death in American men and many prostate cancer patients develop skeletal metastasis. Current treatment modalities for metastatic prostate cancer are mostly palliative with poor prognosis. Epidemiological studies indicated that patients receiving the diabetic drug metformin have lower prostate cancer risk and better prognosis, suggesting that metformin may have anti-neoplastic effects. The mechanism by which metformin acts as chemopreventive agent to impede prostate cancer initiation and progression is unknown. The amplification of c-MYC oncogene plays a key role in early prostate epithelia cell transformation and prostate cancer growth. The purpose of this study is to investigate the effect of metformin on c-myc expression and prostate cancer progression. Our results demonstrated that: (1) In Hi-Myc mice murine prostate neoplasia and tumor model, metformin attenuated the development of prostate intraepithelial neoplasia (PIN, the pre-cancerous lesion of prostate) and PCa lesions. (2) Metformin reduced c-myc protein levels in vivo and in vitro. In Myc-CaP mouse prostate cancer cells, metformin decreased c-myc protein levels by 50% through protein degradation and inhibition of de novo protein synthesis. (3) Metformin selectively inhibited the growth of prostate cancer cells by stimulating cell cycle arrest and apoptosis without affecting the growth of normal prostatic epithelial cells (RWPE-1). (4) Metformin reduced androgen receptor and proliferation marker Ki-67 levels in Hi-Myc mouse prostate glands. Our novel findings suggest that by downregulating c-myc, metformin may act as a chemopreventive agent to restrict prostatic neoplasia initiation and transformation.

Androgen receptor (AR) is the major therapeutic target in aggressive prostate cancer. However, targeting AR alone can result in drug resistance and disease recurrence. Therefore, simultaneous targeting of multiple pathways could in principle be an effective approach to treating prostate cancer. Here we provide proof-of-concept that a small-molecule inhibitor of nuclear beta-catenin activity (called C3) can inhibit both the AR and beta-catenin-signaling pathways that are often misregulated in prostate cancer. Treatment with C3 ablated prostate cancer cell growth by disruption of both beta-catenin/T-cell factor and beta-catenin/AR protein interaction, reflecting the fact that T-cell factor and AR have overlapping binding sites on beta-catenin. Given that AR interacts with, and is transcriptionally regulated by beta-catenin, C3 treatment also resulted in decreased occupancy of beta-catenin on the AR promoter and diminished AR and AR/beta-catenin target gene expression. Interestingly, C3 treatment resulted in decreased AR binding to target genes accompanied by decreased recruitment of an AR and beta-catenin cofactor, coactivator-associated arginine methyltransferase 1 (CARM1), providing insight into the unrecognized function of beta-catenin in prostate cancer. Importantly, C3 inhibited tumor growth in an in vivo xenograft model and blocked renewal of bicalutamide-resistant sphere-forming cells, indicating the therapeutic potential of this approach.