Molecular Mechanisms Of Prostate Cancer Progression

Author : Yezi Zhu
Publisher :
ISBN 13 : 9781321213430
Total Pages : pages
Book Rating : 4.2/5 (134 download)

DOWNLOAD NOW!

Download or read book Molecular Mechanisms of Prostate Cancer Progression and Treatment Resistance written by Yezi Zhu and published by . This book was released on 2014 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Prostate cancer is the most common type of cancer in American men and ranks second to lung cancer in cancer-related deaths. The growth and survival of primary prostate tumors requires physiological level of androgen. Thus hormone therapy has been used for treatment of primary prostate cancer. However, prostate cancer eventually progresses to a castration-resistant state. Understanding the molecular mechanisms leading to castration resistance is very critical for combat this lethal disease. The first part of this thesis will discuss the loss of Rho-GDP dissociation inhibitor [alpha] (RhoGDI[alpha]) as one of the molecular mechanisms leading to prostate cancer castration-resistant progression. My study demonstrated that RhoGDI[alpha] suppresses growth and survival of prostate cancer cells. In this part, I utilized the previously generated LNCaP-IL6+ cells which were able to grow in androgen-deprived condition. The protein expression profiles of LNCaP and LNCaP-IL6+ cells were compared using two-dimensional gel electrophoresis. Expression of RhoGDI[alpha] was reduced in LNCaP-IL6+ cells and was down-regulated in more aggressive prostate cancer cells compared to LNCaP cells. The effects of RhoGDI[alpha] in prostate cancer cells growth and survival were examined both in vitro and in vivo. Overexpression of RhoGDI[alpha] inhibited the growth and induced apoptosis of prostate cancer cells. RhoGDI[alpha] caused LNCaP-IL6+ cells reversal to androgen-sensitive state, and downregulation of RhoGDI[alpha] enhanced growth of androgen-sensitive LNCaP cells in androgen-deprived condition. In addition, RhoGDI[alpha] suppressed the tumorigenic ability and prostate-specific antigen (PSA) production of prostate tumor xenografts in vivo. The aberrant activation of androgen receptor (AR) is responsible for castration-resistant prostate cancer (CRPC) progression and regulation of AR-target genes such as PSA. My in vivo study showed RhoGDI[alpha] inhibited PSA production in the mice sera bearing tumors, indicating RhoGDI[alpha] inhibits AR signaling in prostate cancer cells. The effects of RhoGDI[alpha] on AR signaling will be further discussed. RhoGDI[alpha] was transiently or stably transfected into several prostate cancer cell lines including LNCaP, C4-2, CWR22Rv1 and DU145. The regulation of AR expression by RhoGDI[alpha] was analyzed by qRT-PCR and Western blot. Overexpression of RhoGDI[alpha] downregulated AR expression at both mRNA and protein levels. AR activity was measured by luciferase reporter assays and electrophoretic mobility shift analysis (EMSA). RhoGDI[alpha] was able to inhibit transactivation of AR target genes and prevent AR binding to androgen response element. Immunofluorescence assay was performed and overexpression of RhoGDI[alpha] prevented AR nuclear translocation induced by androgens. The interaction between RhoGDI[alpha] and AR was examined by co-immunoprecipitation assays. RhoGDI[alpha] was found to physically interact with the N-terminal domain of AR. Neuroendocrine differentiation (NED) is associated with castration-resistance of prostate cancer. It has been suggested as a marker of poor prognosis for prostate cancer. Paracrine interleukin-6 (IL-6) can mediate NED features in prostate cancer. The second part of this thesis will discuss the mechanism underlying IL-6-induced NED. RE1-silencing transcription factor (REST) is a main negative regulator of neurogenesis and represses expression of NED genes. I confirmed the IL-6-induced NED by cell morphological changes as well as the induction of NE markers such as neuron-specific enolase (NSE), chromogranin A (ChgA) and synaptophysin. The expression of REST was suppressed in IL-6-induced NED in LNCaP cells. To further study the impact of REST-mediated repression on NED in LNCaP cells, either wild-type REST or a dominant-positive form of REST, REST-VP16, in which both repressor domains of REST were replaced with the activation domain of the herpes simplex virus protein VP16, was introduced into LNCaP cells. Overexpression of exogenous wild type REST abrogated IL-6-induced NED in prostate cancer cells. Expression of the recombinant REST-VP16 fusion protein activated REST target genes and other neuronal differentiation genes and produced neuronal physiological properties. In addition, REST protein turnover was accelerated in IL-6 induced NE differentiated LNCaP cells via the ubiquitin-proteasome pathway, accompanied by a decrease in the expression of the deubiquitylase HAUSP, indicating that pathway(s) priming REST degradation may be involved in IL-6 induced NE differentiation. Docetaxel is the first-line standard treatment for CRPC. However, once tumors develop resistance to docetaxel, the treatment options are again limited. In the last part of this thesis, I will discuss the docetaxel resistance mechanisms and potential therapeutic strategies for docetaxel-resistant CRPC. I established a docetaxel resistant cell line, TaxR, by culturing C4-2B cells in docetaxel in a dose-escalation manner until cells were able to divide freely in 5 nM docetaxel. Global gene expression analysis by cDNA microarrays (approximately 28000 genes) was performed using mRNA from parental C4-2B and TaxR cells. ABCB1, which belongs to the ATP-binding cassette (ABC) transporter family, was identified among the top upregulated genes in TaxR cells. Overexpression of ABCB1 in TaxR cells has been validated by both real-time PCR and Western blot analysis. Downregulation of ABCB1 by specific shRNA or inhibiting ABCB1 activity by ABCB1 inhibitor elacridar restored docetaxel sensitivity in TaxR cells. Apigenin (4', 5, 7-trihydroxyflavone), a natural product belonging to the flavone family, downregulated ABCB1 protein expression in ubiquitin-proteasome pathway and overcame docetaxel resistance in TaxR cells. The effects of different anti-androgens like enzalutamide, abiraterone and bicalutamide on ABCB1 efflux activity were tested using rhodamine123 efflux assay. These antiandrogens inhibited ABCB1 efflux activity and reversed docetaxel resistance in TaxR cells in vitro. The reversal effect of bicalutamide was further confirmed in TaxR xenograft tumors, suggesting targeting ABCB1 could be a potential approach to resensitize docetaxel-resistant prostate cancer cells to docetaxel treatment.