Androgen Receptor Signalling in Prostate Cancer: The Functional Consequences of Acetylation

The androgen receptor (AR) is a ligand activated transcription factor and member of the steroid hormone receptor (SHR) subfamily of nuclear receptors. In the early stages of prostate carcinogenesis, tumour growth is dependent on androgens, and AR directly mediates these effects by modulating gene expression. During transcriptional regulation, the AR recruits numerous cofactors with acetylation-modifying enzymatic activity, the best studied include p300/CBP and the p160/SRC family of coactivators. It is known that recruitment of histone acetyltransferases (HATs) and histone deacetylases (HDACs) is key in fine-tuning responses to androgens and is thus likely to play a role in prostate cancer progression. Further, these proteins can also modify the AR itself. The functional consequences of AR acetylation, the role of modifying enzymes in relation to AR transcriptional response, and prostate cancer will be discussed

Eighty years of targeting androgen receptor activity in prostate cancer: the fight goes on

Prostate cancer (PCa) is the most common cancer in men in the West, other than skin cancer, accounting for over a quarter of cancer diagnoses in US men. In a seminal paper from 1941, Huggins and Hodges demonstrated that prostate tumours and metastatic disease were sensitive to the presence or absence of androgenic hormones. The first hormonal therapy for PCa was thus castration. In the subsequent eighty years, targeting the androgen signalling axis, where possible using drugs rather than surgery, has been a mainstay in the treatment of advanced and metastatic disease. Androgens signal via the androgen receptor, a ligand-activated transcription factor, which is the direct target of many such drugs. In this review we discuss the role of the androgen receptor in PCa and how the combination of structural information and functional screenings is continuing to be used for the discovery of new drug to switch off the receptor or modify its function in cancer cells

A novel role for GSK3β as a modulator of Drosha microprocessor activity and MicroRNA biogenesis

Regulation of microRNA (miR) biogenesis is complex and stringently controlled. Here, we identify the kinase GSK3β as an important modulator of miR biogenesis at Microprocessor level. Repression of GSK3β activity reduces Drosha activity toward pri-miRs, leading to accumulation of unprocessed pri-miRs and reduction of pre-miRs and mature miRs without altering levels or cellular localisation of miR biogenesis proteins. Conversely, GSK3β activation increases Drosha activity and mature miR accumulation. GSK3β achieves this through promoting Drosha:cofactor and Drosha:pri-miR interactions: it binds to DGCR8 and p72 in the Microprocessor, an effect dependent upon presence of RNA. Indeed, GSK3β itself can immunoprecipitate pri-miRs, suggesting possible RNA-binding capacity. Kinase assays identify the mechanism for GSK3β-enhanced Drosha activity, which requires GSK3β nuclear localisation, as phosphorylation of Drosha at S300 and/or S302; confirmed by enhanced Drosha activity and association with cofactors, and increased abundance of mature miRs in the presence of phospho-mimic Drosha. Functional implications of GSK3β-enhanced miR biogenesis are illustrated by increased levels of GSK3β-upregulated miR targets following GSK3β inhibition. These data, the first to link GSK3β with the miR cascade in humans, highlight a novel pro-biogenesis role for GSK3β in increasing miR biogenesis as a component of the Microprocessor complex with wide-ranging functional consequences

Manipulating prohibitin levels provides evidence for an in vivo role in androgen regulation of prostate tumours

Current hormonal therapies for prostate cancer are effective initially, but inevitably tumours progress to an advanced, metastatic stage, often referred to as ‘androgen independent’. However, the androgen receptor (AR) signalling pathway is still key for their growth. It is speculated that tumours escape hormonal control via reduction in corepressor proteins. Manipulating such proteins is thus a potential therapeutic strategy to halt or even reverse tumour progression. We aimed to elucidate the effects of altering levels of the AR corepressor and androgen-target protein prohibitin (PHB) on prostate tumour growth. Prostate cancer cells incorporating an integrated androgen-responsive reporter gene and stably expressing vectors to inducibly overexpress or knockdown PHB were generated and used to assess effects on androgen signalling (by real time imaging) and tumour growth both in culture and in vivo. PHB overexpression inhibited AR activity and prostate-specific antigen (PSA) expression as well as androgen-dependent growth of cells, inducing rapid accumulation in G0/G1. Conversely, reduction in PHB increased AR activity, PSA expression, androgen-mediated growth and S-phase entry. In vivo, doxycycline-induced PHB regulation resulted in marked changes in AR activity, and showed significant effects upon tumour growth. Overexpression led to tumour growth arrest and protection from hormonal starvation, whereas RNAi knockdown resulted in accelerated tumour growth, even in castrated mice. This study provides proof of principle that i) reduction in PHB promotes both androgen-dependent and ‘androgen-independent’ tumour growth, and ii) altering AR activity via increasing levels or activity of corepressors is a valid therapeutic strategy for advanced prostate cancer

Roles of steroid receptors in the lung and COVID-19

COVID-19 symptoms and mortality are largely due to its devastating effects in the lungs. The disease is caused by the SARS (Severe Acute Respiratory Syndrome)-CoV-2 coronavirus, which requires host cell proteins such as ACE2 (angiotensin-converting enzyme 2) and TMPRSS2 (transmembrane serine protease 2) for infection of lung epithelia. The expression and function of the steroid hormone receptor family is important in many aspects that impact on COVID-19 effects in the lung – notably lung development and function, the immune system, and expression of TMPRSS2 and ACE2. This review provides a brief summary of current knowledge on the roles of the steroid hormone receptors [androgen receptor (AR), glucocorticoid receptor (GR), progesterone receptor (PR), mineralocorticoid receptor (MR) and oestrogen receptor (ER)] in the lung, their effects on host cell proteins that facilitate SARS-CoV-2 uptake, and provides a snapshot of current clinical trials investigating the use of steroid receptor (SR) ligands to treat COVID-19

Detection of YAP1 and AR-V7 mRNA for Prostate Cancer prognosis using an ISFET Lab-On-Chip platform

AbstractProstate cancer (PCa) is the second most common cause of male cancer-related death worldwide. The gold standard of treatment for advanced PCa is androgen deprivation therapy (ADT). However, eventual failure of ADT is common and leads to lethal metastatic castration resistant PCa (mCRPC). As such, the detection of relevant biomarkers in the blood for drug resistance in mCRPC patients could lead to personalized treatment options. mRNA detection is often limited by the low specificity of qPCR assays which are restricted to specialised laboratories. Here, we present a novel reversetranscription loop-mediated isothermal amplification (RT-LAMP) assay and have demonstrated its capability for sensitive detection of AR-V7 and YAP1 RNA (3×101 RNA copies per reaction). This work presents a foundation for the detection of circulating mRNA in PCa on a non-invasive Lab-on-chip (LoC) device for use at point-of-care. This technique was implemented onto a Lab-on-Chip platform integrating an array of chemical sensors (ion-sensitive field-effect transistors - ISFETs) for real-time detection of RNA. Detection of RNA presence was achieved through the translation of chemical signals into electrical readouts. Validation of this technique was conducted with rapid detection (&lt;15 min) of extracted RNA from prostate cancer cell lines 22Rv1s and DU145s.</jats:p

Antiandrogens Act as Selective Androgen Receptor Modulators at the Proteome Level in Prostate Cancer Cells*

Current therapies for prostate cancer include antiandrogens, inhibitory ligands of the androgen receptor, which repress androgen-stimulated growth. These include the selective androgen receptor modulators cyproterone acetate and hydroxyflutamide and the complete antagonist bicalutamide. Their activity is partly dictated by the presence of androgen receptor mutations, which are commonly detected in patients who relapse while receiving antiandrogens, i.e. in castrate-resistant prostate cancer. To characterize the early proteomic response to these antiandrogens we used the LNCaP prostate cancer cell line, which harbors the androgen receptor mutation most commonly detected in castrate-resistant tumors (T877A), analyzing alterations in the proteome, and comparing these to the effect of these therapeutics upon androgen receptor activity and cell proliferation. The majority are regulated post-transcriptionally, possibly via nongenomic androgen receptor signaling. Differences detected between the exposure groups demonstrate subtle changes in the biological response to each specific ligand, suggesting a spectrum of agonistic and antagonistic effects dependent on the ligand used. Analysis of the crystal structures of the AR in the presence of cyproterone acetate, hydroxyflutamide, and DHT identified important differences in the orientation of key residues located in the AF-2 and BF-3 protein interaction surfaces. This further implies that although there is commonality in the growth responses between androgens and those antiandrogens that stimulate growth in the presence of a mutation, there may also be influential differences in the growth pathways stimulated by the different ligands. This therefore has implications for prostate cancer treatment because tumors may respond differently dependent upon which mutation is present and which ligand is activating growth, also for the design of selective androgen receptor modulators, which aim to elicit differential proteomic responses dependent upon cellular context

The antiviral potential of the antiandrogen enzalutamide and the viral‐androgen signaling interplay in seasonal coronaviruses

The sex disparity in COVID-19 outcomes with males generally faring worse than females has been associated with the androgen-regulated expression of the protease TMPRSS2 and the cell receptor ACE2 in the lung and fueled interest in antiandrogens as potential antivirals. In this study, we explored enzalutamide, an antiandrogen used commonly to treat prostate cancer, as a potential antiviral against the human coronaviruses which cause seasonal respiratory infections (HCoV-NL63, -229E, and -OC43). Using lentivirus-pseudotyped and authentic HCoV, we report that enzalutamide reduced 229E and NL63 entry and infection in both TMPRSS2- and nonexpressing immortalized cells, suggesting a TMPRSS2-independent mechanism. However, no effect was observed against OC43. To decipher this distinction, we performed RNA-sequencing analysis on 229E- and OC43-infected primary human airway cells. Our results show a significant induction of androgen-responsive genes by 229E compared to OC43 at 24 and 72 h postinfection. The virus-mediated effect on AR-signaling was further confirmed with a consensus androgen response element-driven luciferase assay in androgen-depleted MRC-5 cells. Specifically, 229E induced luciferase-reporter activity in the presence and absence of the synthetic androgen mibolerone, while OC43 inhibited induction. These findings highlight a complex interplay between viral infections and androgen-signaling, offering insights for disparities in viral outcomes and antiviral interventions

Follicle-stimulating hormone promotes growth of human prostate cancer cell line-derived tumor xenografts

Chemical castration in prostate cancer can be achieved with gonadotropin-releasing hormone (GnRH) agonists or antagonists. Their effects differ by the initial flare of gonadotropin and testosterone secretion with agonists and the immediate pituitary-testicular suppression by antagonists. While both suppress luteinizing hormone (LH) and follicle-stimulating hormone (FSH) initially, a rebound in FSH levels occurs during agonist treatment. This rebound is potentially harmful, taken the expression of FSH receptors (R) in prostate cancer tissue. We herein assessed the role of FSH in promoting the growth of androgen-independent (PC-3, DU145) and androgen-dependent (VCaP) human prostate cancer cell line xenografts in nude mice. Gonadotropins were suppressed with the GnRH antagonist degarelix, and effects of add-back human recombinant FSH were assessed on tumor growth. All tumors expressed GnRHR and FSHR, and degarelix treatment suppressed their growth. FSH supplementation reversed the degarelix-evoked suppression of PC-3 tumors, both in preventive (degarelix and FSH treatment started upon cell inoculation) and therapeutic (treatments initiated 3 weeks after cell inoculation) setting. A less marked, though significant FSH effect occurred in DU145, but not in VCaP xenografts. FSHR expression in the xenografts supports direct FSH stimulation of tumor growth. Testosterone supplementation, to maintain the VCaP xenografts, apparently masked the FSH effect on their growth. Treatment with the LH analogue hCG did not affect PC-3 tumor growth despite their expression of luteinizing hormone/choriongonadotropin receptor. In conclusion, FSH, but not LH, may directly stimulate the growth of androgen-independent prostate cancer, suggesting that persistent FSH suppression upon GnRH antagonist treatment offers a therapeutic advantage over agonist