Androgen-regulated Gene Research Articles

Androgen-dependent sertoli cell tight junction remodeling is mediated by multiple tight junction components.

Sertoli cell tight junctions (SCTJs) of the seminiferous epithelium create a specialized microenvironment in the testis to aid differentiation of spermatocytes and spermatids from spermatogonial stem cells. SCTJs must be chronically broken and rebuilt with high fidelity to allow the transmigration of preleptotene spermatocytes from the basal to adluminal epithelial compartment. Impairment of androgen signaling in Sertoli cells perturbs SCTJ remodeling. Claudin (CLDN) 3, a tight junction component under androgen regulation, localizes to newly forming SCTJs and is absent in Sertoli cell androgen receptor knockout (SCARKO) mice. We show here that Cldn3-null mice do not phenocopy SCARKO mice: Cldn3(-/-) mice are fertile, show uninterrupted spermatogenesis, and exhibit fully functional SCTJs based on imaging and small molecule tracer analyses, suggesting that other androgen-regulated genes must contribute to the SCARKO phenotype. To further investigate the SCTJ phenotype observed in SCARKO mutants, we generated a new SCARKO model and extensively analyzed the expression of other tight junction components. In addition to Cldn3, we identified altered expression of several other SCTJ molecules, including down-regulation of Cldn13 and a noncanonical tight junction protein 2 isoform (Tjp2iso3). Chromatin immunoprecipitation was used to demonstrate direct androgen receptor binding to regions of these target genes. Furthermore, we demonstrated that CLDN13 is a constituent of SCTJs and that TJP2iso3 colocalizes with tricellulin, a constituent of tricellular junctions, underscoring the importance of androgen signaling in the regulation of both bicellular and tricellular Sertoli cell tight junctions.

Paracrine interactions between LNCaP prostate cancer cells and bioengineered bone in 3D in vitro culture reflect molecular changes during bone metastasis

As microenvironmental factors such as three-dimensionality and cell–matrix interactions are increasingly being acknowledged by cancer biologists, more complex 3D in vitro models are being developed to study tumorigenesis and cancer progression. To better understand the pathophysiology of bone metastasis, we have established and validated a 3D indirect co-culture model to investigate the paracrine interactions between prostate cancer (PCa) cells and human osteoblasts. Co-culture of the human PCa, LNCaP cells embedded within polyethylene glycol hydrogels with human osteoblasts in the form of a tissue engineered bone construct (TEB), resulted in reduced proliferation of LNCaP cells. LNCaP cells in both monoculture and co-culture were responsive to the androgen analog, R1881, as indicated by an increase in the expression (mRNA and/or protein induction) of androgen-regulated genes including prostate specific antigen and fatty acid synthase. Microarray gene expression analysis further revealed an up-regulation of bone markers and other genes associated with skeletal and vasculature development and a significant activation of transforming growth factor β1 downstream genes in LNCaP cells after co-culture with TEB. LNCaP cells co-cultured with TEB also unexpectedly showed similar changes in classical androgen-responsive genes under androgen-deprived conditions not seen in LNCaP monocultures. The molecular changes of LNCaP cells after co-culturing with TEBs suggest that osteoblasts exert a paracrine effect that may promote osteomimicry and modulate the expression of androgen-responsive genes in LNCaP cells. Taken together, we have presented a novel 3D in vitro model that allows the study of cellular and molecular changes occurring in PCa cells and osteoblasts that are relevant to metastatic colonization of bone. This unique in vitro model could also facilitate cancer biologists to dissect specific biological hypotheses via extensive genomic or proteomic assessments to further our understanding of the PCa-bone crosstalk.

ERG rearrangement and protein expression in the progression to castration-resistant prostate cancer

Approximately half of the prostate carcinomas are characterized by a chromosomal rearrangement fusing the androgen-regulated gene TMPRSS2 to the oncogenic ETS transcription factor ERG. Aim of this study was to comprehensively analyze the role and impact of the ERG rearrangement and protein expression on the progression to castration-resistant (CR) disease. We used a tissue microarray (TMA) constructed from 114 hormone naive (HN) and 117 CR PCs. We analyzed the ERG rearrangement status by fluorescence in situ hybridization and the expression profiles of ERG, androgen receptor (AR) and the proliferation marker Ki67 by immunohistochemistry. Nearly half of the PC tissue specimens (HN: 38%, CR: 46%) harbored a TMPRSS2-ERG gene fusion. HN PCs with positive translocation status showed increased tumor cell proliferation (P<0.05). As expected, TMPRSS2-ERG gene fusion was strongly associated with increased ERG protein expression in HN and CR PCs (both P<0.0001). Remarkably, the study revealed a subgroup (26%) of CR PCs with ERG rearrangement but without any detectable ERG protein expression. This subgroup showed significantly lower levels of AR protein expression and androgen-regulated serum PSA (both P<0.05). In this study, we identified a subgroup of ERG-rearranged CR PCs without detectable ERG protein expression. Our results suggest that this subgroup could represent CR PCs with a dispensed AR pathway. These tumors might represent a thus far unrecognized subset of patients with AR-independent CR PC who may not benefit from conventional therapy directed against the AR pathway.

Commentary on “Integrative genomic analyses reveal an androgen-driven somatic alteration landscape in early-onset prostate cancer.” Weischenfeldt J, Simon R, Feuerbach L, Schlangen K, Weichenhan D, Minner S, Wuttig D, Warnatz HJ, Stehr H, Rausch T, Jäger N, Gu L, Bogatyrova O, Stütz AM, Claus R, Eils J, Eils R, Gerhäuser C, Huang PH, Hutter B, Kabbe R, Lawerenz C, Radomski

Early-onset prostate cancer (EO-PCA) represents the earliest clinical manifestation of prostate cancer. To compare the genomic alteration landscapes of EO-PCA with “classical” (elderly-onset) PCA, we performed deep sequencing-based genomics analyses in 11 tumors diagnosed at young age, and pursued comparative assessments with seven elderly-onset PCA genomes. Remarkable age-related differences in structural rearrangement (SR) formation became evident, suggesting distinct disease pathomechanisms. Whereas EO-PCAs harbored a prevalence of balanced SRs, with a specific abundance of androgen-regulated ETS gene fusions including TMPRSS2:ERG, elderly-onset PCAs displayed primarily non-androgen-associated SRs. Data from a validation cohort of>10,000 patients showed age-dependent androgen receptor levels and a prevalence of SRs affecting androgen-regulated genes, further substantiating the activity of a characteristic “androgen-type” pathomechanism in EO-PCA.

DNA damage response (DDR) via NKX3.1 expression in prostate cells

It has been reported that NKX3.1 an androgen-regulated homeobox gene restricted to prostate and testicular tissues, encodes a homeobox protein, which transcriptionally regulates oxidative damage responses and enhances topoisomerase I re-ligation by a direct interaction with the ATM protein in prostate cells. In this study, we aimed to investigate the role of NKX3.1 in DNA double-strand break (DSB) repair. We demonstrate that the DNA damage induced by CPT-11 (irinotecan, a topo I inhibitor), doxorubicin (a topo II inhibitor), and H2O2 (a mediator of oxidative damage), but not by etoposide (another topo II inhibitor), is negatively influenced by NKX3.1 expression. We also examined γH2AX(S139) foci formation and observed that the overexpression of NKX3.1 resulted a remarkable decrease in the formation of γH2AX(S139) foci. Intriguingly, we observed in NKX3.1 silencing studies that the depletion of NKX3.1 correlated with a significant decrease in the levels of p-ATM(S1981) and γH2AX(S139). The data imply that the DNA damage response (DDR) can be altered, perhaps via a decrease in the topoisomerase I re-ligation function; this is consistent with the physical association of NKX3.1 with DDR mediators upon treatment of both PC-3 and LNCaP cells with CPT-11. Furthermore, the depletion of NKX3.1 resulted in a G1/S progression via the facilitation of an increase in E2F stabilization concurrent with the suppressed DDR. Thus, the topoisomerase I inhibitor-mediated DNA damage enhanced the physical association of NKX3.1 with γH2AX(S139) on the chromatin in LNCaP cells, whereas NKX3.1 in the soluble fraction was associated with p-ATM(S1981) and RAD50 in these cells. Overall, the data suggest that androgens and NKX3.1 expression regulate the progression of the cell cycle and concurrently activate the DDR. Therefore, androgen withdrawal may augment the development of an error-prone phenotype and, subsequently, the loss of DNA damage control during prostate cancer progression.

ETS fusion genes in prostate cancer

Prostate cancer is very common in elderly men in developed countries. Unravelling the molecular and biological processes that contribute to tumor development and progressive growth, including its heterogeneity, is a challenging task. The fusion of the genes ERG and TMPRSS2 is the most frequent genomic alteration in prostate cancer. ERG is an oncogene that encodes a member of the family of ETS transcription factors. At lower frequency, other members of this gene family are also rearranged and overexpressed in prostate cancer. TMPRSS2 is an androgen-regulated gene that is preferentially expressed in the prostate. Most of the less frequent ETS fusion partners are also androgen-regulated and prostate-specific. During the last few years, novel concepts of the process of gene fusion have emerged, and initial experimental results explaining the function of the ETS genes ERG and ETV1 in prostate cancer have been published. In this review, we focus on the most relevant ETS gene fusions and summarize the current knowledge of the role of ETS transcription factors in prostate cancer. Finally, we discuss the clinical relevance of TMRPSS2-ERG and other ETS gene fusions in prostate cancer.

Expression of androgen receptor target genes in skeletal muscle

We aimed to determine the mechanisms of the anabolic actions of androgens in skeletal muscle by investigating potential androgen receptor (AR)-regulated genes in in vitro and in vivo models. The expression of the myogenic regulatory factor myogenin was significantly decreased in skeletal muscle from testosterone-treated orchidectomized male mice compared to control orchidectomized males, and was increased in muscle from male AR knockout mice that lacked DNA binding activity (ARΔZF2) versus wildtype mice, demonstrating that myogenin is repressed by the androgen/AR pathway. The ubiquitin ligase Fbxo32 was repressed by 12 h dihydrotestosterone treatment in human skeletal muscle cell myoblasts, and c-Myc expression was decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle, and increased in ARΔZF2 muscle. The expression of a group of genes that regulate the transition from myoblast proliferation to differentiation, Tceal7, p57Kip2, Igf2 and calcineurin Aa, was increased in ARΔZF2 muscle, and the expression of all but p57Kip2 was also decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle. We conclude that in males, androgens act via the AR in part to promote peak muscle mass by maintaining myoblasts in the proliferative state and delaying the transition to differentiation during muscle growth and development, and by suppressing ubiquitin ligase-mediated atrophy pathways to preserve muscle mass in adult muscle.

Targeted Androgen Pathway Suppression in Localized Prostate Cancer: A Pilot Study

Ligand-mediated activation of the androgen receptor (AR) is critical for prostate cancer (PCa) survival and proliferation. The failure to completely ablate tissue androgens may limit suppression of PCa growth. We evaluated combinations of CYP17A and 5-α-reductase inhibitors for reducing prostate androgen levels, AR signaling, and PCa volumes. Thirty-five men with intermediate/high-risk clinically localized PCa were randomly assigned to goserelin combined with dutasteride (ZD), bicalutamide and dutasteride (ZBD), or bicalutamide, dutasteride, and ketoconazole (ZBDK) for 3 months before prostatectomy. Controls included patients receiving combined androgen blockade with luteinizing hormone-releasing hormone agonist and bicalutamide. The primary outcome measure was tissue dihydrotestosterone (DHT) concentration. Prostate DHT levels were substantially lower in all experimental arms (0.02 to 0.04 ng/g v 0.92 ng/g in controls; P < .001). The ZBDK group demonstrated the greatest percentage decline in serum testosterone, androsterone, and dehydroepiandrosterone sulfate (P < .05 for all). Staining for AR and the androgen-regulated genes prostate-specific antigen and TMPRSS2 was strongly suppressed in benign glands and moderately in malignant glands (P < .05 for all). Two patients had pathologic complete response, and nine had ≤ 0.2 cm(3) of residual tumor (defined as a near-complete response), with the largest numbers of complete and near-complete responses in the ZBDK group. Addition of androgen synthesis inhibitors lowers prostate androgens below that achieved with standard therapy, but significant AR signaling remains. Tissue-based analysis of steroids and AR signaling is critical to informing the search for optimal local and systemic control of high-risk prostate cancer.

TBLR1 as an androgen receptor (AR) coactivator selectively activates AR target genes to inhibit prostate cancer growth

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 β-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.

Androgen receptor genomic regulation.

The transcriptional activity of the androgen receptor (AR) is not only critical for the normal development and function of the prostate but also pivotal to the onset and progression of prostate cancer (PCa). The studies of AR transcriptional regulation were previously limited to a handful of AR-target genes. Owing to the development of various high-throughput genomic technologies, significant advances have been made in recent years. Here we discuss the discoveries of genome-wide androgen-regulated genes in PCa cell lines, animal models and tissues using expression microarray and sequencing, the mapping of genomic landscapes of AR using Combining Chromatin Immunoprecipitation (ChIP)-on-chip and ChIP-seq assays, the interplay of transcriptional cofactors in defining AR binding profiles, and the genomic regulation and AR reprogramming in advanced PCa.

Characterisation of the androgen regulation of glycine N-methyltransferase in prostate cancer cells

The development and growth of prostate cancer is dependent on androgens; thus, the identification of androgen-regulated genes in prostate cancer cells is vital for defining the mechanisms of prostate cancer development and progression and developing new markers and targets for prostate cancer treatment. Glycine N-methyltransferase (GNMT) is a S-adenosylmethionine-dependent methyltransferase that has been recently identified as a novel androgen-regulated gene in prostate cancer cells. Although the importance of this protein in prostate cancer progression has been extensively addressed, little is known about the mechanism of its androgen regulation. Here, we show that GNMT expression is stimulated by androgen in androgen receptor (AR) expressing cells and that the stimulation occurs at the mRNA and protein levels. We have identified an androgen response element within the first exon of the GNMT gene and demonstrated that AR binds to this element in vitro and in vivo. Together, these studies identify GNMT as a direct transcriptional target of the AR. As this is an evolutionarily conserved regulatory element, this highlights androgen regulation as an important feature of GNMT regulation.

Understanding the temporal sequence of genetic events that lead to prostate cancer progression and metastasis

Prostate cancer is the second most common cancer among men, with an estimated 903,000 newly diagnosed cases and 258,000 deaths per year worldwide (1). Although the prognosis is favorable in patients with localized disease, the 5-y survival rate drops to less than 30% for individuals with advanced, metastatic tumors (2). Although androgen deprivation therapy in the form of chemical or surgical castration is currently used to treat metastatic disease, response to treatment is usually temporary, and a significant percentage of patients invariably progress to castration-resistant prostate cancer (CRPC), which often arises due to micrometastases (3). The molecular mechanisms that contribute to tumor development, recurrence, and resistance to standard therapies are poorly understood, making it extremely difficult to predict which patients with treatable, early-stage prostate cancer will progress to lethal forms of the disease. Fortunately, global and computational analyses of the prostate cancer genome have provided us with part of the foundation needed to tackle this issue. One of the first genomewide studies was published in 2005 (4) and showed that ∼50% of all prostate tumors contain chromosomal rearrangements that involve the ETS family of transcription factors, primarily in the form of the TMPRSS2-ERG translocation, which fuse the ETS family member ERG with the androgen-regulated TMPRSS2 gene. This and other translocations involving ETS family genes are present even at early stages of the disease and are specific to prostate cancer (4). However, as the disease progresses and androgen receptor (AR) dependence is lost, these genetic lesions cooperate with signaling pathways, such those mediated by pRB, RAS/RAF, and phosphatidylinositol 3-kinase (PI3K)/PTEN/AKT, which are altered in a more “global” sense to promote metastasis (5). In PNAS, Aytes et al. (6) describe the temporal progression and the consequence of molecular activation of the PI3K and RAS signaling pathways during prostate cancer metastasis. Using a unique genetically engineered mouse model of prostate cancer that allows lineage tracing, the authors demonstrate that the spread of tumor cells is an early event that actually occurs well before the onset of metastasis. They further show that metastasis is temporally coincident with the expression of ETV4, a member of the PEA3 subfamily of ETS transcription factors that mediates late events associated with metastasis (Fig. 1).

Next-generation Sequencing of Advanced Prostate Cancer Treated with Androgen-deprivation Therapy

BackgroundAndrogen-deprivation therapy (ADT) is standard treatment for locally advanced or metastatic prostate cancer (PCa). Many patients develop castration resistance (castration-resistant PCa [CRPC]) after approximately 2–3 yr, with a poor prognosis. The molecular mechanisms underlying CRPC progression are unclear. ObjectiveTo undertake quantitative tumour transcriptome profiling prior to and following ADT to identify functionally important androgen-regulated pathways or genes that may be reactivated in CRPC. Design, setting, and participantsRNA sequencing (RNA-seq) was performed on tumour-rich, targeted prostatic biopsies from seven patients with locally advanced or metastatic PCa before and approximately 22 wk after ADT initiation. Differentially regulated genes were identified in treatment pairs and further investigated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) on cell lines and immunohistochemistry on a separate CRPC patient cohort. Functional assays were used to determine the effect of pathway modulation on cell phenotypes. Outcome measurements and statistical analysisWe searched for gene expression changes affecting key cell signalling pathways that may be targeted as proof of principle in a CRPC in vitro cell line model. Results and limitationsWe identified ADT-regulated signalling pathways, including the Wnt/β-catenin signalling pathway, and observed overexpression of β-catenin in a subset of CRPC by immunohistochemistry. We validated 6 of 12 (50%) pathway members by qRT-PCR on LNCaP/LNCaP-AI cell RNAs, of which 4 (67%) demonstrated expression changes consistent with RNA-seq data. We show that the tankyrase inhibitor XAV939 (which promotes β-catenin degradation) reduced androgen-independent LNCaP-AI cell line growth compared with androgen-responsive LNCaP cells via an accumulation of cell proportions in the G0/G1 phase and reduction in the S and G2/M phases. Our biopsy protocol did not account for tumour heterogeneity, and pathway inhibition was limited to pharmacologic approaches. ConclusionsRNA-seq of paired PCa samples revealed ADT-regulated signalling pathways. Proof-of-principle inhibition of the Wnt/β-catenin signalling pathway specifically delays androgen-independent PCa cell cycle progression and proliferation and warrants further investigation as a potential target for therapy for CRPC.

TCTP is an androgen-regulated gene implicated in prostate cancer.

TCTP has been implicated in a plethora of important cellular processes related to cell growth, cell cycle progression, malignant transformation and inhibition of apoptosis. In addition to these intracellular functions, TCTP has extracellular functions and plays an important role in immune cells. TCTP expression was previously shown to be deregulated in prostate cancer, but its function in prostate cancer cells is largely unknown. Here we show that TCTP expression is regulated by androgens in LNCaP prostate cancer cells in vitro as well as human prostate cancer xenografts in vivo. Knockdown of TCTP reduced colony formation and increased apoptosis in LNCaP cells, implicating it as an important factor for prostate cancer cell growth. Global gene expression profiling in TCTP knockdown LNCaP cells showed that several interferon regulated genes are regulated by TCTP, suggesting that it may have a role in regulating immune function in prostate cancer. In addition, recombinant TCTP treatment increased colony formation in LNCaP cells suggesting that secreted TCTP may function as a proliferative factor in prostate cancer. These results suggest that TCTP may have a role in prostate cancer development.

Novel 5′ Fusion Partners of ETV1 and ETV4 in Prostate Cancer

Gene fusions involving the erythroblast transformation-specific (ETS) transcription factors ERG, ETV1, ETV4, ETV5, and FLI1 are a common feature of prostate carcinomas (PCas). Themost common upstream fusion partner described is the androgenregulated prostate-specific gene TMPRSS2, most frequently with ERG, but additional 5′fusion partners have been described. We performed 5′ rapid amplification of cDNA ends in 18 PCas with ETV1, ETV4, or ETV5 outlier expression to identify the 5′ fusion partners. We also evaluated the exon-level expression profile of these ETS genes in 14 cases. We identified and confirmed by fluorescent in situ hybridization (FISH) and reverse transcription-polymerase chain reaction the two novel chimeric genes OR51E2-ETV1 and UBTF-ETV4 in two PCas. OR51E2 encodes a G-protein.coupled receptor that is overexpressed in PCas, whereas UBTF is a ubiquitously expressed gene encoding an HMG-box DNA-binding protein involved in ribosome biogenesis. We additionally describe two novel gene fusion combinations of previously described genes, namely, SLC45A3- ETV4 and HERVK17-ETV4. Finally, we found one PCa with TMPRSS2-ETV1, one with C15orf21-ETV1, one with EST14-ETV1, and two with 14q133-q21.1-ETV1. In nine PCas (eight ETV1 and one ETV5), exhibiting ETS outlier expression and genomic rearrangement detected by FISH, no 5′ fusion partner was found. Our findings contribute significantly to characterize the heterogeneous group of ETS gene fusions and indicate that all genes described as 5′ fusion partners with one ETS gene can most likely be rearranged with any of the other ETS genes involved in prostate carcinogenesis.

Differentially Expressed Androgen-Regulated Genes in Androgen-Sensitive Tissues Reveal Potential Biomarkers of Early Prostate Cancer

BackgroundSeveral data favor androgen receptor implication in prostate cancer initiation through the induction of several gene activation programs. The aim of the study is to identify potential biomarkers for early diagnosis of prostate cancer (PCa) among androgen-regulated genes (ARG) and to evaluate comparative expression of these genes in normal prostate and normal prostate-related androgen-sensitive tissues that do not (or rarely) give rise to cancer.MethodsARG were selected in non-neoplastic adult human prostatic epithelial RWPE-1 cells stably expressing an exogenous human androgen receptor, using RNA-microarrays and validation by qRT-PCR. Expression of 48 preselected genes was quantified in tissue samples (seminal vesicles, prostate transitional zones and prostate cancers, benign prostatic hypertrophy obtained from surgical specimens) using TaqMan® low-density arrays. The diagnostic performances of these potential biomarkers were compared to that of genes known to be associated with PCa (i.e. PCA3 and DLX1).Results and DiscussionBy crossing expression studies in 26 matched PCa and normal prostate transitional zone samples, and 35 matched seminal vesicle and PCa samples, 14 genes were identified. Similarly, 9 genes were overexpressed in 15 benign prostatic hypertrophy samples, as compared to PCa samples. Overall, we selected 8 genes of interest to evaluate their diagnostic performances in comparison with that of PCA3 and DLX1. Among them, 3 genes: CRYAB, KCNMA1 and SDPR, were overexpressed in all 3 reference non-cancerous tissues. The areas under ROC curves of these genes reached those of PCA3 (0.91) and DLX1 (0.94).ConclusionsWe identified ARG with reduced expression in PCa and with significant diagnostic values for discriminating between cancerous and non-cancerous prostatic tissues, similar that of PCA3. Given their expression pattern, they could be considered as potentially protective against prostate cancer. Moreover, they could be complementary to known genes overexpressed in PCa and included along with them in multiplex diagnostic tools.

Targeting Alternative Sites on the Androgen Receptor to Treat Castration-Resistant Prostate Cancer

Recurrent, metastatic prostate cancer continues to be a leading cause of cancer-death in men. The androgen receptor (AR) is a modular, ligand-inducible transcription factor that regulates the expression of genes that can drive the progression of this disease, and as a consequence, this receptor is a key therapeutic target for controlling prostate cancer. The current drugs designed to directly inhibit the AR are called anti-androgens, and all act by competing with androgens for binding to the androgen/ligand binding site. Unfortunately, with the inevitable progression of the cancer to castration resistance, many of these drugs become ineffective. However, there are numerous other regulatory sites on this protein that have not been exploited therapeutically. The regulation of AR activity involves a cascade of complex interactions with numerous chaperones, co-factors and co-regulatory proteins, leading ultimately to direct binding of AR dimers to specific DNA androgen response elements within the promoter and enhancers of androgen-regulated genes. As part of the family of nuclear receptors, the AR is organized into modular structural and functional domains with specialized roles in facilitating their inter-molecular interactions. These regions of the AR present attractive, yet largely unexploited, drug target sites for reducing or eliminating androgen signaling in prostate cancers. The design of small molecule inhibitors targeting these specific AR domains is only now being realized and is the culmination of decades of work, including crystallographic and biochemistry approaches to map the shape and accessibility of the AR surfaces and cavities. Here, we review the structure of the AR protein and describe recent advancements in inhibiting its activity with small molecules specifically designed to target areas distinct from the receptor’s androgen binding site. It is anticipated that these new classes of anti-AR drugs will provide an additional arsenal to treat castration-resistant prostate cancer.

GRTH: A Key to Understanding Androgen-Mediated Germ Cell Signaling

GRTH: A Key to Understanding Androgen-Mediated Germ Cell Signaling

A Peptide against Soluble Guanylyl Cyclase α1: A New Approach to Treating Prostate Cancer

Among the many identified androgen-regulated genes, sGCα1 (soluble guanylyl cyclase α1) appears to play a pivotal role in mediating the pro-cancer effects of androgens and androgen receptor. The classical role for sGCα1 is to heterodimerize with the sGCβ1 subunit, forming sGC, the enzyme that mediates nitric oxide signaling by catalyzing the synthesis of cyclic guanosine monophosphate. Our published data show that sGCα1 can drive prostate cancer cell proliferation independent of hormone and provide cancer cells a pro-survival function, via a novel mechanism for p53 inhibition, both of which are independent of sGCβ1, NO, and cGMP. All of these properties make sGCα1 an important novel target for prostate cancer therapy. Thus, peptides were designed targeting sGCα1 with the aim of disrupting this protein’s pro-cancer activities. One peptide (A-8R) was determined to be strongly cytotoxic to prostate cancer cells, rapidly inducing apoptosis. Cytotoxicity was observed in both hormone-dependent and, significantly, hormone-refractory prostate cancer cells, opening the possibility that this peptide can be used to treat the usually lethal castration-resistant prostate cancer. In mouse xenograft studies, Peptide A-8R was able to stop tumor growth of not only hormone-dependent cells, but most importantly from hormone-independent cells. In addition, the mechanism of Peptide A cytotoxicity is generation of reactive oxygen species, which recently have been recognized as a major mode of action of important cancer drugs. Thus, this paper provides strong evidence that targeting an important AR-regulated gene is a new paradigm for effective prostate cancer therapy.

Low systemic testosterone levels induce androgen maintenance in benign rat prostate tissue

Prostate cancer (PC) is both an age- and an androgen-dependent disease. Paradoxically, systemic levels of androgens decline with age as the risk of PC rises. While there is no correlation between systemic androgen levels and the risk of PC, systemic androgen levels do not reflect the levels of androgens in prostate tissue. In metastatic PC, changes in the androgen biosynthesis pathway during hormone therapy result in increased levels of androgens in cancer tissue and contribute to continued androgen receptor (AR) signaling. It is possible that similar changes occur in normal prostate tissue as androgen levels decline with age and that this contributes to tumorigenesis. In the present study, we sought to determine whether the rat prostate is able to maintain functional levels of androgens despite low serum testosterone levels. Rats were castrated and implanted with capsules to achieve castrate, normal, sub-physiological, and supra-physiological levels of testosterone. After 6 weeks of treatment, LC-MS/MS was used to quantify the levels of testosterone and dihydrotestosterone (DHT) in the serum and prostate tissue. Quantitative RT-PCR was used to quantify the expression of genes involved in the androgen/AR signaling axis. Despite significantly different levels of testosterone and DHT being present in the serum, testosterone and DHT concentrations in prostate tissue from different testosterone-treatment groups were very similar. Furthermore, the expression of androgen-regulated genes in the prostate was similar among all the testosterone-treatment groups, demonstrating that the rat prostate can maintain a functional level of androgens despite low serum testosterone levels. Low-testosterone treatment resulted in significant alterations in the expression of androgen biosynthesis genes, which may be related to maintaining functional androgen levels.