Role Of Androgen Signaling Research Articles

Analysis of the responsiveness to antiandrogens in multiple breast cancer cell lines.

Antiandrogens were originally developed as therapeutic agents for prostate cancer but are also expected to be effective for breast cancer. However, the role of androgen signaling in breast cancer has long been controversial due to the limited number of experimental models. Our study aimed to comprehensively investigate the efficacy of antiandrogens on breast cancer. In the present study, a total of 18 breast cancer cell lines were treated with the agonist or antagonists of the androgen receptor (AR). Among the 18 cell lines tested, only T-47D cells proliferated in an androgen-dependent manner, while the other cell lines were almost irresponsive to AR stimulation. On the other hand, treatment with AR antagonists at relatively high doses suppressed the proliferation of not only T-47D cells but also some other cell lines including AR-low/negative cells. In addition, expression of the full-length AR and constitutively active AR splice variants, AR-V7 and ARV567es, was not correlated with sensitivity to AR antagonists. These data suggest that the antiproliferative effect of AR antagonists is AR-independent in some cases. Consistently, proliferation of AR-knockout BT-549 cells was inhibited by AR antagonists. Identification of biomarkers would be necessary to determine which breast cancer patients will benefit from these drugs.

Role of androgen signaling in androgen receptor-positive extramammary Paget's disease: Establishment of organoids and their biological analysis as a novel therapeutic target

BackgroundExtramammary Paget’s disease (EMPD) is a rare intraepithelial adenocarcinoma that mainly affects the anogenital and axillary regions. Although its etiology has not been fully elucidated, there is evidence that androgen receptors (AR) are expressed in most cases of EMPD. However, the role of androgen signaling in the pathogenesis of EMPD remains unclear. ObjectiveTo evaluate the role of androgen signaling in tumor growth of AR-positive EMPD. MethodsPatient-derived organoids were established and cultured from two AR-positive EMPD patients: one man and one woman. Cultured organoids were treated with androgen agonists and/or antagonists, then subjected to analysis of changes in organoid proliferation, as well as changes in androgen signaling pathway-specific genes. ResultsOrganoid cultures were established from each EMPD sample. These organoids were immunohistologically and genetically identical to the original tumor. For each organoid sample, viable cell number increased in response to androgen exposure. The mRNA level of Fkbp5, a known AR target gene, increased in a concentration-dependent manner in organoids exposed to the synthetic androgen R1881. Conversely, the AR inhibitor darolutamide suppressed the viable cell number in a concentration-dependent manner. The mRNA expression levels of MKI67 and Fkbp5 were also suppressed by darolutamide. ConclusionOur results indicate that androgen signaling is a key pathway involved in the growth of AR-positive EMPD. Therefore, androgen signaling inhibition may be a novel treatment option for EMPD patients who require systemic therapy.

OXER1 and RACK1-associated pathway: a promising drug target for breast cancer progression

Recent data indicate that receptor for activated C kinase 1 (RACK1) is a putative prognostic marker and drug target in breast cancer (BC). High RACK1 expression is negatively associated with overall survival, as it seems to promote BC progression. In tumors, RACK1 expression is controlled by a complex balance between glucocorticoids and androgens. Given the fact that androgens and androgenic derivatives can inhibit BC cell proliferation and migration, the role of androgen signaling in regulating RACK1 transcription in mammary tumors is of pivotal interest. Here, we provide evidence that nandrolone (19-nortosterone) inhibits BC cell proliferation and migration by antagonizing the PI3K/Akt/NF-κB signaling pathway, which eventually results in RACK1 downregulation. We also show that nandrolone impairs the PI3K/Akt/NF-κB signaling pathway and decreases RACK1 expression via binding to the membrane-bound receptor, oxoeicosanoid receptor 1 (OXER1). High levels of OXER1 are observed in several BC cell lines and correlate with RACK1 expression and poor prognosis. Our data provide evidence on the role played by the OXER1-dependent intracellular pathway in BC progression and shed light on the mechanisms underlying membrane-dependent androgen effects on RACK1 regulation. Besides the mechanistic relevance, the results of the study are of interest from a translational prospective. In fact, they identify a new and actionable pathway to be used for the design of innovative and rational therapeutic strategies in the context of the personalized treatment of BC. In addition, they draw attention on nandrolone-based compounds that lack hormonal activity as potential anti-tumor agents.

Activation of GPR56, a novel adhesion GPCR, is necessary for nuclear androgen receptor signaling in prostate cells.

The androgen receptor (AR) is activated in patients with castration resistant prostate cancer (CRPC) despite low circulating levels of androgen, suggesting that intracellular signaling pathways and non-androgenic factors may contribute to AR activation. Many G-protein coupled receptors (GPCR) and their ligands are also activated in these cells indicating that they may play a role in development of Prostate Cancer (PCa) and CRPC. Although a cross talk has been suggested between the two pathways, yet, the identity of GPCRs which may play a role in androgen signaling, is not established yet. By using blast analysis of 826 GPCRs, we identified a GPCR, GPCR 205, which exhibited maximum similarity with the ligand binding domain of the AR. We demonstrate that adhesion GPCR 205, also known as GPR56, can be activated by androgens to stimulate the Rho signaling pathway, a pathway that plays an important role in prostate tumor cell metastasis. Testosterone stimulation of GPR56 also activates the cAMP/ Protein kinase A (PKA) pathway, that is necessary for AR signaling. Knocking down the expression of GPR56 using siRNA, disrupts nuclear translocation of AR and transcription of prototypic AR target genes such as PSA. GPR56 expression is higher in all twenty-five prostate tumor patient's samples tested and cells expressing GPR56 exhibit increased proliferation. These findings provide new insights about androgen signaling and identify GPR56 as a possible therapeutic target in advanced prostate cancer patients.

BCOR-coupled H2A monoubiquitination represses a subset of androgen receptor target genes regulating prostate cancer proliferation.

We have identified BCL6 corepressor (BCOR) as a hormone-dependent interaction partner of androgen receptor (AR), a key transcription factor in the development of normal and cancerous prostate. BCOR is often mutated in cancers and hematological diseases and as a component of a non-canonical polycomb repressive complex 1 (ncPRC1.1) required for arranging many facets of cellular differentiation. However, its role in androgen signaling or prostate cancer cells remains unknown. Here, our genome-wide analyses reveal that BCOR is recruited in an androgen-dependent fashion to majority of AR-binding chromatin sites in castration-resistant prostate cancer (CRPC) cells. Interestingly, depletion of BCOR has a significant effect on the expression of androgen-repressed genes linked to regulation of cell proliferation, differentiation and development. At many of these genes, such as HOX genes, the depletion leads to a decrease in H2A K119 monoubiquitination and an increase in mRNA expression. Consistently, BCOR depletion impairs the proliferation and viability of CRPC cells, inducing their apoptosis. Collectively, our data indicate a key role for the BCOR-ncPRC1.1 complex in the corepression of an important subset of AR target genes and the regulation of prostate cancer cell proliferation.

Abstract 5225: Androgen deprivation upregulates SPINK1 expression and potentiates cellular plasticity in prostate cancer

Abstract The Serine Peptidase Inhibitor, Kazal type 1 (SPINK1) overexpression represents the second-largest prostate cancer (PCa) subtype associated with increased risk of recurrence and poor prognosis. Regardless of molecular subtype, androgen-deprivation therapy (ADT) remains the mainstay treatment for locally advanced and metastatic PCa patients. However, majority of the treated individuals eventually progress to castration-resistant stage and a subset of these patients develop ADT-induced neuroendocrine prostate cancer. Despite evidences of detrimental effects of ADT on PCa, possible role of androgen signaling in SPINK1-mediated prostate oncogenesis remains unexplored. Here, we show that androgen receptor (AR) functions as a direct transcriptional repressor of SPINK1, and blocking AR signaling relieves its repression, leading to upregulation of SPINK1. In agreement, we observe an inverse association between SPINK1 levels and AR expression across multiple patient cohorts, and in neuroendocrine differentiated LNCaP cells. We show that AR and its corepressor, the RE1-silencing transcription factor (REST), occupy SPINK1 promoter and inhibits its transcription. On the other hand, in the absence of AR, lineage reprogramming factor SOX2 in turn binds to SPINK1 promoter leading to its positive transcriptional regulation in androgen-deprived conditions with concomitant increase in neuroendocrine markers. Additionally, stable knockdown of SPINK1 results in reduced epithelial-mesenchymal transition, decreased stemness and drug resistance. Collectively, our findings provide a plausible explanation to the paradoxical clinical outcomes of ADT, arising due to increase in SPINK1 levels. Finally, we emphasize the need to take a well-informed decision prior to ADT and develop alternative therapeutic strategies for castrate-resistant PCa patients. Citation Format: Ritika Tiwari, Nishat Manzar, Vipul Bhatia, Anjali Yadav, Shannon Carskadon, Nilesh Gupta, Amina Zoubeidi, Nallasivam Palanisamy, Bushra Ateeq. Androgen deprivation upregulates SPINK1 expression and potentiates cellular plasticity in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5225.

Novel role for androgen signaling in pyelonephritis

Sex differences in urinary tract infection (UTI) susceptibility and severity are known, but have historically focused on anatomic differences between males and females. Until recently, experimental UTI has been limited to female animals due to ease of transurethral bladder catheterization. Olson and colleagues have developed a model of experimental UTI independent of sex that relies on direct bladder inoculation and thus permits investigation of sex differences in UTI susceptibility. They now build upon their prior work in this model by implicating androgens as drivers of tubular invasion by uropathogenic Escherichia coli, which form luminal bacterial communities preceding renal abscess formation.

Androgen Signaling in Esophageal Adenocarcinoma Cell Lines In Vitro

BackgroundWe showed previously that nuclear localization of the androgen receptor (AR) and expression of the androgen-responsive gene FK506-binding protein 5 (FKBP5) in esophageal adenocarcinoma (EAC) tissues were associated with decreased patient survival, suggesting a role for androgens in this cancer.AimTo investigate the effect of the AR ligand 5α-dihydrotestosterone (DHT) on AR-expressing EAC cell lines in vitro.Methods and ResultsIn tissue resection specimens from EAC patients, FKBP5 expression was positively associated with proliferation as measured by Ki-67 expression. We stably transduced AR into three AR-negative EAC cell lines, OE33, JH-EsoAd1, and OE19, to investigate androgen signaling in vitro. In the AR-expressing cell lines, 10 nM DHT, the concentration typically used to study AR signaling, induced changes in the expression of androgen-responsive genes and inhibited proliferation by inducing cell cycle arrest and senescence. At lower DHT concentrations near the half maximal inhibitory concentration (IC50), the AR-expressing cell lines proliferated and there were changes in the expression of androgen-responsive genes. In direct co-culture with cancer-associated fibroblast-like PShTert myofibroblasts, 10 nM DHT induced changes in the expression of androgen-responsive genes but did not inhibit proliferation.ConclusionsThis is the first study to show that EAC cell lines respond to androgen in vitro. Proliferation together with the expression of androgen-responsive genes was dependent on the concentration of DHT, or the presence of a permissive microenvironment, consistent with observations in the tissues. These findings are consistent with a role for androgen signaling in EAC.

Unexpected central role of the androgen receptor in the spontaneous regeneration of myelin.

Lost myelin can be replaced after injury or during demyelinating diseases in a regenerative process called remyelination. In the central nervous system (CNS), the myelin sheaths, which protect axons and allow the fast propagation of electrical impulses, are produced by oligodendrocytes. The abundance and widespread distribution of oligodendrocyte progenitors (OPs) within the adult CNS account for this remarkable regenerative potential. Here, we report a key role for the male gonad, testosterone, and androgen receptor (AR) in CNS remyelination. After lysolecithin-induced demyelination of the male mouse ventral spinal cord white matter, the recruitment of glial fibrillary acidic protein-expressing astrocytes was compromised in the absence of testes and testosterone signaling via AR. Concomitantly, the differentiation of OPs into oligodendrocytes forming myelin basic protein (MBP)+ and proteolipid protein-positive myelin was impaired. Instead, in the absence of astrocytes, axons were remyelinated by protein zero (P0)+ and peripheral myelin protein 22-kDa (PMP22)+ myelin, normally only produced by Schwann cells in the peripheral nervous system. Thus, testosterone favors astrocyte recruitment and spontaneous oligodendrocyte-mediated remyelination. This finding may have important implications for demyelinating diseases, psychiatric disorders, and cognitive aging. The testosterone dependency of CNS oligodendrocyte remyelination may have roots in the evolutionary history of the AR, because the receptor has evolved from an ancestral 3-ketosteroid receptor through gene duplication at the time when myelin appeared in jawed vertebrates.

One-Carbon Metabolism in Prostate Cancer: The Role of Androgen Signaling.

Cancer cell metabolism differs significantly from the metabolism of non-transformed cells. This altered metabolic reprogramming mediates changes in the uptake and use of nutrients that permit high rates of proliferation, growth, and survival. The androgen receptor (AR) plays an essential role in the establishment and progression of prostate cancer (PCa), and in the metabolic adaptation that takes place during this progression. In its role as a transcription factor, the AR directly affects the expression of several effectors and regulators of essential catabolic and biosynthetic pathways. Indirectly, as a modulator of the one-carbon metabolism, the AR can affect epigenetic processes, DNA metabolism, and redox balance, all of which are important factors in tumorigenesis. In this review, we focus on the role of AR-signaling on one-carbon metabolism in tumorigenesis. Clinical implications of one-carbon metabolism and AR-targeted therapies for PCa are discussed in this context.

Abstract 615: TRPM8 channel as a novel molecular target in androgen-regulated prostate cancer cells

Abstract TRPM8 channels are highly expressed in prostate cancer cells but the physiological and pathological functions of TRPM8 in these cells are not known. In addition to prostate cancer (PC) cells, TRPM8 is expressed in sensory neurons where it acts as a sensor of cold. Recently, trpm8 has been identified as both an important gene regulated by androgens and a critical regulator of Ca2+ homeostasis. The androgen and its receptor (AR) are pivotal in the biology of sex hormone-regulated malignancies, with PC the most affected tumor. From the chromatin-immunoprecipitation (ChIP) analysis, we observed that an androgen response element (ARE) mediates androgen regulation of TRPM8. Further, using immunoprecipitation and calcium-imaging experiments, we identified that TRPM8 Ca+2 channels are functionally associated with androgens (testosterone and 5α-dihydrotestosterone) in the prostate. We also identified that unlike prostate cancer cell lines (LNCAP, PC3 and RWPE2) and normal prostate tissues, prostate tumor tissues (grade 1-4) showed high levels of TRPM8 which was predominantly localized as the plasma membrane protein at the tumor periphery. Of note, the decreased stabilization of TRPM8 on the plasma membrane was associated with increased expression of AR protein. These results suggest an important role of androgen signaling in trpm8 gene transcription corroborating to cancer progression and clinical lethality in prostate cancer. Citation Format: Swapna Asuthkar, Kiran Velpula, Eleonora Zakharian. TRPM8 channel as a novel molecular target in androgen-regulated prostate cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 615. doi:10.1158/1538-7445.AM2014-615

CCAR1 promotes chromatin loading of androgen receptor (AR) transcription complex by stabilizing the association between AR and GATA2

Androgen receptor (AR), a ligand-dependent transcription factor, plays a critical role in prostate cancer onset and progression, and its transcriptional function is mediated largely by distinct nuclear receptor co-regulators. Here, we show that cell cycle and apoptosis regulator 1 (CCAR1) functions as an AR co-activator. CCAR1 interacted with and enhanced the transcriptional activity of AR. Depletion of CCAR1 caused reduction in androgen-dependent expression of a subset of AR target genes. We further showed that CCAR1 is required for recruitment of AR, MED1 and RNA polymerase II to the enhancers of AR target genes and for androgen-induced long-range prostate specific antigen enhancer–promoter interaction. The molecular mechanism underlying CCAR1 function in AR-mediated transcription involves CCAR1-mediated enhanced recruitment of GATA2, a pioneer factor for AR, to AR-binding sites. CCAR1 stabilized the interaction between AR and GATA2 by interacting directly with both proteins, thereby facilitating AR and GATA2 occupancy on the enhancers. Furthermore, CCAR1 depletion inhibited the growth, migration, invasion of prostate cancer cells and reduced the tumorigenicity of prostate cancer cells in vivo. Our results firmly established CCAR1 as an AR co-activator that plays a key role in AR transcription complex assembly and has an important physiological role in androgen signaling and prostate tumorigenesis.

Essential roles of androgen signaling in Wolffian duct stabilization and epididymal cell differentiation

The epididymis is a male accessory organ and functions for sperm maturation and storage under the control of androgen. The development of the epididymis is also androgen dependent. The Wolffian duct (WD), anlagen of the epididymis, is formed in both male and female embryos; however, it is stabilized only in male embryos by testicular androgen. Androgen drives subsequent differentiation of the WD into the epididymis. Although the essential roles of androgen in WD masculinization and epididymal function have been established, little is known about cellular events regulated precisely by androgen signaling during these processes. It is also unclear whether androgen signaling, especially in the epithelia, has further function for epididymal epithelial cell differentiation. In this study we examined the cellular death and proliferation controlled by androgen signaling via the androgen receptor (AR) in WD stabilization. Analyses using AR knockout mice revealed that androgen signaling inhibits epithelial cell death in this process. Analysis of AP2α-Cre;ARflox/Y mice, in which AR function is deleted in the WD epithelium, revealed that epithelial AR is not required for the WD stabilization but is required for epithelial cell differentiation in the epididymis. Specifically, loss of epithelial AR significantly reduced expression of p63 that is essential for differentiation of basal cells in the epididymal epithelium. We also interrogated the possibility of regulation of the p63 gene (Trp63) by AR in vitro and found that p63 is a likely direct target of AR regulation.

Androgen action via testicular peritubular myoid cells is essential for male fertility.

Androgens are essential for normal spermatogenesis and male fertility, but how androgens exert this effect remains uncertain. Androgen receptors (ARs) are expressed in several testicular cell types, but continuing uncertainty exists over which cell type mediates androgen control of spermatogenesis. Androgen signaling via Sertoli cells (SCs) is essential for complete spermatogenesis, but the role for androgen signaling via peritubular myoid (PTM) cells is contentious. To address this controversy, we generated PTM-specific AR-knockout (PTM-ARKO) mice in which gross reproductive development was normal, but all PTM-ARKO males were azoospermic and infertile. Testis weight was reduced beyond puberty, and in adulthood there was an 86% reduction in germ cells, compared with wild-type littermates. These changes were not explained by any deficits in testosterone, luteinizing hormone, or follicle-stimulating hormone concentrations. SC function was impaired in PTM-ARKO males, indicated by reduced seminiferous tubule fluid production and reduced expression of some androgen-dependent SC genes. Androgen action via PTM cells is therefore essential for normal testis function, spermatogenesis, and fertility in males. This study also provides the first direct evidence for the importance of androgen-driven stromal-epithelial interactions underpinning the regulation of spermatogenesis; PTM-ARKO mice will enable identification of the new molecular pathways involved.—Welsh, M., Saunders, P. T. K., Atanassova, N., Sharpe, R. M., Smith, L. B. Androgen action via testicular peritubular myoid cells is essential for male fertility.

Small glutamine-rich tetratricopeptide repeat-containing protein alpha (SGTA), a candidate gene for polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is a heterogenic, complex common genetic disease. Multiple pathways are involved in its pathogenesis, including the androgen signaling pathway and insulin signaling pathway. Small glutamine-rich tetratricopeptide repeat-containing protein alpha (SGTA) is a putative member of the androgen receptor-chaperone-co-chaperone complex, and may play a role in androgen signaling as a co-chaperone. Polymorphisms in the SGTA gene have not been evaluated for a role in PCOS. Women with and without PCOS (287 cases, 187 controls) were genotyped for three single nucleotide polymorphisms (SNPs) in SGTA. SNPs and haplotypes were determined and tested for association with PCOS and component traits of PCOS. For SNP rs1640262, homozygotes for the minor allele were protected against PCOS (P = 0.009). Haplotype 1 (G-A-T) was associated with increased risk of PCOS (P = 0.015). In women with PCOS, haplotype 2 (A-G-C) was associated with increased insulin resistance (P = 0.013), consequently resulting in increased insulin secretion (P = 0.014). This study presents genetic evidence suggesting a potential role of SGTA in the pathogenesis of PCOS. SGTA may provide a connection between multiple pathways in PCOS.

A Coregulatory Role for the Mediator Complex in Prostate Cancer Cell Proliferation and Gene Expression

Androgen receptor (AR) signaling pathways are important for the survival and proliferation of prostate cancer cells. Because AR activity is facilitated by distinct coregulatory factors and complexes, it is conceivable that some of these proteins might also play a role in promoting prostate oncogenesis. The multisubunit Mediator complex is an important coactivator for a broad range of regulatory transcriptional factors including AR, yet its role in prostate cancer is unclear. Here, we used RNA interference to knock down the expression of two integral Mediator components, MED1/TRAP220 and MED17, in prostate cancer cells. MED1/TRAP220 plays a particularly important role in androgen signaling in that it serves as a direct binding target for AR. We found that the knockdown of either subunit markedly decreases transcription from transiently transfected androgen-responsive reporter genes, as well as inhibits androgen-dependent expression of endogenous AR target genes. We show for the first time that loss of either MED1/TRAP220 or MED17 in prostate cancer cells significantly decreases both androgen-dependent and -independent cellular proliferation, inhibits cell cycle progression, and increases apoptosis. Furthermore, we show that MED1/TRAP220 is overexpressed in both AR-positive and -negative prostate cancer cells lines, as well as in 50% (10 of 20) of the clinically localized human prostate cancers we examined, thus suggesting that MED1/TRAP220 hyperactivity may have implications in prostate oncogenesis. In sum, our data suggest that Mediator plays an important coregulatory role in prostate cancer cell proliferation and survival, and therefore, may represent a new target for therapeutic intervention.