In the majority of cases, advanced prostate cancer responds initially to androgen deprivation therapy by depletion of gonadal testosterone. The response is usually transient, and metastatic tumors almost invariably eventually progress as castration-resistant prostate cancer (CRPC). The development of CRPC is dependent upon the intratumoral generation of the potent androgen, dihydrotestosterone (<em>DHT</em>), from adrenal precursor steroids. Progression to CRPC is accompanied by increased expression of steroid-<em>5α</em>-reductase isoenzyme-1 (SRD5A1) over SRD5A2, which is otherwise the dominant isoenzyme expressed in the prostate. <em>DHT</em> synthesis in CRPC is widely assumed to require <em>5α</em>-reduction of testosterone as the obligate precursor, and the increased expression of SRD5A1 is thought to reflect its role in converting testosterone to <em>DHT</em>. Here, we show that the dominant route of <em>DHT</em> synthesis in CRPC bypasses testosterone, and instead requires <em>5α</em>-reduction of androstenedione by SRD5A1 to <em>5α</em>-androstanedione, which is then converted to <em>DHT</em>. This alternative pathway is operational and dominant in both human CRPC cell lines and fresh tissue obtained from human tumor metastases. Moreover, CRPC growth in mouse xenograft models is dependent upon this pathway, as well as expression of SRD5A1. These findings reframe the fundamental metabolic pathway that drives CRPC progression, and shed light on the development of new therapeutic strategies.

Prostate cancer resistance to castration occurs because tumours acquire the metabolic capability of converting precursor steroids to <em>5α</em>-dihydrotestosterone (<em>DHT</em>), promoting signalling by the androgen receptor and the development of castration-resistant prostate cancer. Essential for resistance, <em>DHT</em> synthesis from adrenal precursor steroids or possibly from de novo synthesis from cholesterol commonly requires enzymatic reactions by 3β-hydroxysteroid dehydrogenase (3βHSD), steroid-<em>5α</em>-reductase (SRD5A) and 17β-hydroxysteroid dehydrogenase (17βHSD) isoenzymes. Abiraterone, a steroidal 17α-hydroxylase/17,20-lyase (CYP17A1) inhibitor, blocks this synthetic process and prolongs survival. We hypothesized that abiraterone is converted by an enzyme to the more active Δ(4)-abiraterone (D4A), which blocks multiple steroidogenic enzymes and antagonizes the androgen receptor, providing an additional explanation for abiraterone's clinical activity. Here we show that abiraterone is converted to D4A in mice and patients with prostate cancer. D4A inhibits CYP17A1, 3βHSD and SRD5A, which are required for <em>DHT</em> synthesis. Furthermore, competitive androgen receptor antagonism by D4A is comparable to the potent antagonist enzalutamide. D4A also has more potent anti-tumour activity against xenograft tumours than abiraterone. Our findings suggest an additional explanation-conversion to a more active agent-for abiraterone's survival extension. We propose that direct treatment with D4A would be more clinically effective than abiraterone treatment.

The androgen receptor (AR) mediates the growth of benign and malignant prostate in response to dihydrotestosterone (<em>DHT</em>). In patients undergoing androgen deprivation therapy for prostate cancer, AR drives prostate cancer growth despite low circulating levels of testicular androgen and normal levels of adrenal androgen. In this report, we demonstrate the extent of AR transactivation in the presence of <em>5α</em>-androstane-3α,17β-diol (androstanediol) in prostate-derived cell lines parallels the bioconversion of androstanediol to <em>DHT</em>. AR transactivation in the presence of androstanediol in prostate cancer cell lines correlated mainly with mRNA and protein levels of 17β-hydroxysteroid dehydrogenase 6 (17β-HSD6), one of several enzymes required for the interconversion of androstanediol to <em>DHT</em> and the inactive metabolite androsterone. Levels of retinol dehydrogenase 5, and dehydrogenase/reductase short-chain dehydrogenase/reductase family member 9, which also convert androstanediol to <em>DHT</em>, were lower than 17β-HSD6 in prostate-derived cell lines and higher in the castration-recurrent human prostate cancer xenograft. Measurements of tissue androstanediol using mass spectrometry demonstrated androstanediol metabolism to <em>DHT</em> and androsterone. Administration of androstanediol dipropionate to castration-recurrent CWR22R tumor-bearing athymic castrated male mice produced a 28-fold increase in intratumoral <em>DHT</em> levels. AR transactivation in prostate cancer cells in the presence of androstanediol resulted from the cell-specific conversion of androstanediol to <em>DHT</em>, and androstanediol increased LAPC-4 cell growth. The ability to convert androstanediol to <em>DHT</em> provides a mechanism for optimal utilization of androgen precursors and catabolites for <em>DHT</em> synthesis.

The majority of prostate cancers (PCa) express high levels of androgen receptor (AR) and are dependent for their growth on testosterone produced by the testes, which is reduced in the prostate to the higher affinity ligand <em>5α</em>-dihydrotestosterone (<em>DHT</em>). PCa growth can be suppressed by androgen deprivation therapy, which involves removal of testicular androgens (surgical or medical castration) or treatment with an AR antagonist (or a combination of both), but patients invariably relapse with tumors that have been termed castration recurrent/resistant PCa (CRPC). Importantly, AR transcriptional activity becomes reactivated at this CRPC stage of the disease and remains essential for tumor growth. The objective of this review is to outline one clinically important mechanism contributing to this AR reactivation, which is increased intratumoral synthesis of testosterone and <em>DHT</em> from weak androgens produced by the adrenal glands and possibly de novo from cholesterol. Early studies showed that a substantial fraction of CRPC patients responded to adrenalectomy or medical suppression of adrenal androgen synthesis using agents such as ketoconazole (CYP17A1 inhibitor), and a recent phase III study of a more potent and selective CYP17A1 inhibitor (abiraterone) has demonstrated an improvement in survival. With the pending FDA approval of abiraterone for CRPC, defining the molecular mechanisms contributing to CYP17A1 inhibitor resistance/relapse and AR reactivation is now critical to build on these advances.

BACKGROUND

Steroid <em>5α</em>-reductase inhibitors are used to treat benign prostatic hyperplasia and androgenic alopecia, but the role of <em>5α</em>-dihydrotestosterone (<em>DHT</em>) in mediating testosterone's effects on muscle, sexual function, erythropoiesis, and other androgen-dependent processes remains poorly understood.

OBJECTIVE

To determine whether testosterone's effects on muscle mass, strength, sexual function, hematocrit level, prostate volume, sebum production, and lipid levels are attenuated when its conversion to <em>DHT</em> is blocked by dutasteride (an inhibitor of <em>5α</em>-reductase type 1 and 2).

METHODS

The <em>5α</em>-Reductase Trial was a randomized controlled trial of healthy men aged 18 to 50 years comparing placebo plus testosterone enthanate with dutasteride plus testosterone enanthate from May 2005 through June 2010.

METHODS

Eight treatment groups received 50, 125, 300, or 600 mg/wk of testosterone enanthate for 20 weeks plus placebo (4 groups) or 2.5 mg/d of dutasteride (4 groups).

METHODS

The primary outcome was change in fat-free mass; secondary outcomes: changes in fat mass, muscle strength, sexual function, prostate volume, sebum production, and hematocrit and lipid levels.

RESULTS

A total of 139 men were randomized; 102 completed the 20-week intervention. Men assigned to dutasteride were similar at baseline to those assigned to placebo. The mean fat-free mass gained by the dutasteride groups was 0.6 kg (95% CI, -0.1 to 1.2 kg) when receiving 50 mg/wk of testosterone enanthate, 2.6 kg (95% CI, 0.9 to 4.3 kg) for 125 mg/wk, 5.8 kg (95% CI, 4.8 to 6.9 kg) for 300 mg/wk, and 7.1 kg (95% CI, 6.0 to 8.2 kg) for 600 mg/wk. The mean fat-free mass gained by the placebo groups was 0.8 kg (95% CI, -0.1 to 1.7 kg) when receiving 50 mg/wk of testosterone enanthate, 3.5 kg (95% CI, 2.1 to 4.8 kg) for 125 mg/wk, 5.7 kg (95% CI, 4.8 to 6.5 kg) for 300 mg/wk, and 8.1 kg (95% CI, 6.7 to 9.5 kg) for 600 mg/wk. The dose-adjusted differences between the dutasteride and placebo groups for fat-free mass were not significant (P = .18). Changes in fat mass, muscle strength, sexual function, prostate volume, sebum production, and hematocrit and lipid levels did not differ between groups.

CONCLUSIONS

Changes in fat-free mass in response to graded testosterone doses did not differ in men in whom DHT was suppressed by dutasteride from those treated with placebo, indicating that conversion of testosterone to DHT is not essential for mediating its anabolic effects on muscle.

BACKGROUND

clinicaltrials.gov Identifier: NCT00493987.

BACKGROUND

Recent findings suggest that BPH has an inflammatory component. Clinical trials have documented that therapy with LHRH antagonist Cetrorelix causes a marked and prolonged improvement in LUTS in men with symptomatic BPH. We investigated the mechanism of action and effect of Cetrorelix in a rat model of BPH.

METHODS

Adult male Wistar rats were used. BPH was induced in rats by subcutaneous injections of TE 2 mg/day for 4 weeks. Control animals received injections of corn oil. After induction of BPH, rats received depot Cetrorelix pamoate at the doses of 0.625, 1.25, and 12.5 mg/kg on days 1 and 22 and TE-control rats received vehicle injections. Whole prostates were weighed and processed for RNA and protein. Real-time RT-PCR assays for numerous inflammatory cytokines and growth factors were performed. Quantitative analyses of prostatic LHRH receptor, LHRH, androgen receptor (AR) and <em>5α</em>-reductase 2 were done by real-time RT-PCR and immunoblotting; serum <em>DHT</em>, LH, PSA, and IGF-1 by immunoassays.

RESULTS

mRNA levels for inflammatory cytokines IFN-γ, IL-3, IL-4, IL-5, IL-6, IL-8, IL-13, IL-15, and IL-17 and for growth factors EGF, FGF-2, FGF-7, FGF-8, FGF-14, TGF-β1, and VEGF-A were significantly reduced by Cetrorelix 0.625 mg/kg (P < 0.05). Prostate weights were also significantly lowered by any dose of Cetrorelix.

CONCLUSIONS

This study suggests that Cetrorelix reduces various inflammatory cytokines and growth factors in rat prostate and, at doses which do not induce castration levels of testosterone, can lower prostate weights. Our findings shed light on the mechanism of action of LHRH antagonists in BPH.

BACKGROUND

17-Hydroxyprogesterone (17-OHP) can be converted to dihydrotestosterone (<em>DHT</em>) via an alternative "backdoor" route that bypasses the conventional intermediates androstenedione and testosterone. In this backdoor pathway, 17-OHP is converted to <em>5α</em>-pregnane-3α,17α-diol-20-one (pdiol), which is an excellent substrate for the 17,20 lyase activity of CYP17A1 to produce androsterone. OBJECTIVE AND HYPOTHESES: The objective of this study was to obtain evidence for the presence of the backdoor pathway in patients with 21-hydroxylase deficiency (21-OHD).

METHODS

We compared urinary steroid hormone profiles determined by gas chromatography-mass spectrometry of 142 untreated 21-OHD patients (age range, 1 d to 25.4 yr; 51 males) with 138 control subjects. The activity of the backdoor pathway was assessed using the ratios of the urinary concentrations of pdiol to those of the metabolites of the classic Δ4 and Δ5 pathways. In contrast to etiocholanolone, which originates almost exclusively from the classic pathways, androsterone may be derived additionally from the backdoor pathway. Therefore, the androsterone to etiocholanolone ratio can be used as an indicator for the presence of the backdoor pathway.

RESULTS

Untreated 21-OHD subjects showed increased urinary ratios of pdiol to the Δ4 and Δ5 pathway metabolites and a higher androsterone to etiocholanolone ratio.

CONCLUSIONS

The elevated ratios of pdiol to the Δ4 and Δ5 pathway metabolites as well as the higher androsterone to etiocholanolone ratio in patients with 21-OHD indicate postnatal activity of the backdoor pathway with maximum activity during early infancy. Our data provide new insights into the pathophysiology of androgen biosynthesis of 21-OHD.

Castration-resistant prostate cancer (CRPC) continues to pose a significant clinical challenge with new generation second-line hormonal therapies affording limited improvement in disease outcome. As the androgen receptor (AR) remains a critical driver in CRPC, understanding the determinants of its transcriptional activity is important for developing new AR-targeted therapies. FOXA1 is a key component of the AR transcriptional complex yet its role in prostate cancer progression and the relationship between AR and FOXA1 are not completely resolved. It is well established that FOXA1 levels are elevated in advanced prostate cancer and metastases. We mimicked these conditions by overexpressing FOXA1 in the androgen-responsive LNCaP prostate cancer cell line and observed a significant increase in AR genomic binding at novel regions that possess increased chromatin accessibility. High levels of FOXA1 resulted in increased proliferation at both sub-optimal and high <em>5α</em>-dihydrotestosterone (<em>DHT</em>) concentrations. Immunohistochemical staining for FOXA1 in a clinical prostate cancer cohort revealed that high FOXA1 expression is associated with shorter time to biochemical recurrence after radical prostatectomy (hazard ratio (HR) 5.0, 95% confidence interval (CI) 1.2-21.1, P=0.028), positive surgical margins and higher stage disease at diagnosis. The gene expression program that results from FOXA1 overexpression is enriched for PTEN, Wnt and other pathways typically represented in CRPC gene signatures. Together, these results suggest that in an androgen-depleted state, elevated levels of FOXA1 enhance AR binding at genomic regions not normally occupied by AR, which in turn facilitates prostate cancer cell growth.

Estrogen receptor β (ERβ) is activated in the prostate by <em>5α</em>-androstane-3β,17β-diol (3β-Adiol) where it exerts antiproliferative activity. The proliferative action of the androgen receptor is activated by <em>5α</em>-dihydrotestosterone (<em>DHT</em>). Thus, prostate growth is governed by the balance between androgen receptor and ERβ activation. 3β-Adiol is a high-affinity ligand and agonist of ERβ and is derived from <em>DHT</em> by 3-keto reductase/3β-hydroxysteroid dehydrogenase enzymes. Here, we demonstrate that, when it is expressed in living cells containing an estrogen response element-luciferase reporter, 17β-hydroxysteroid dehydrogenase type 6 (17βHSD6) converts the androgen <em>DHT</em> to the estrogen 3β-Adiol, and this leads to activation of the ERβ reporter. This conversion of <em>DHT</em> occurs at concentrations that are in the physiological range of this hormone in the prostate. Immunohistochemical analysis revealed that 17βHSD6 is expressed in ERβ-positive epithelial cells of the human prostate and that, in prostate cancers of Gleason grade higher than 3, both ERβ and 17βHSD6 are undetectable. Both proteins were present in benign prostatic hyperplasia samples. These observations reveal that formation of 3β-Adiol via 17βHSD6 from <em>DHT</em> is an important growth regulatory pathway that is lost in prostate cancer.

1. The central nervous systems (CNS) of males and females differ in the control mechanisms for the release of gonadotropins from the anterior pituitary gland as well as the capacity to display sex specific behaviors. 2. In guinea pigs and monkeys, these differences are organized through the actions of prenatal androgens secreted by the fetal testes. In both males and females androgen receptors have been identified within the brain during the period in development in which organization of the CNS occurs. Sex differences between the ratio of cytosolic and nuclear androgen receptors are due to the amount of endogenous androgen present in the circulation of the developing fetus. Thus, at least part of the biochemical machinery necessary for androgen action resides in the CNS during the period of sexual differentiation. 3. In addition to the physiological differences that have been observed, morphological differences that are androgen dependent have been found in the medial preoptic nucleus and the bed nucleus of the stria terminalis of the guinea pig. The location of these sex differences in brain morphology coincides roughly with the location of steroid binding neurons. 4. In some species the in situ conversion of testosterone to dihydrotestosterone (<em>DHT</em>) by the 5 alpha-reductases or to estradiol-17 beta by cytochrome P450 aromatase mediates testosterone's action. The gonadotropin surge mechanism of adult guinea pigs exposed to a <em>5a</em>-reductase inhibitor in utero during the critical period for sexual differentiation was unaffected in either males or females even though the development of the external organs of reproduction of males was feminized by the treatment. Likewise, the gonadotropin surge mechanism of subjects exposed to an aromatase inhibitor in utero during the critical period for sexual differentiation was unaffected by this treatment. 5. The mechanism controlling negative feedback, however, was affected in both males and females. Subjects that were exposed to an aromatase inhibitor while developing in utero could not respond to the negative feedback actions of estrogen on gonadotropin release in adulthood. 6. The surge mechanism for the control of gonadotropin secretion in nonhuman primates is not sexually differentiated as it is in rodents. Castrated male monkeys release surge amounts of LH in response to an estrogen challenge. Both infant and adult dimorphic behaviors of rhesus monkeys are organized by the prenatal actions of androgen.

High-affinity binding of dihydrotestosterone (<em>DHT</em>) to the androgen receptor (AR) initiates androgen-dependent gene activation, required for normal male sex development in utero, and contributes to prostate cancer development and progression in men. Under normal physiologic conditions, <em>DHT</em> is synthesized predominantly by <em>5α</em>-reduction of testosterone, the major circulating androgen produced by the testis. During androgen deprivation therapy, intratumoral androgen production is sufficient for AR activation and prostate cancer growth, even though circulating testicular androgen levels are low. Recent studies indicate that the metabolism of <em>5α</em>-androstane-3α, 17β-diol by 17β-hydroxysteroid dehydrogenase 6 in benign prostate and prostate cancer cells is a major biosynthetic pathway for intratumoral synthesis of <em>DHT</em>, which binds AR and initiates transactivation to promote prostate cancer growth during androgen deprivation therapy. Drugs that target the so-called backdoor pathway of <em>DHT</em> synthesis provide an opportunity to enhance clinical response to luteinizing-hormone-releasing hormone (LHRH) agonists or antagonists, AR antagonists, and inhibitors of <em>5α</em>-reductase enzymes (finasteride or dutasteride), and other steroid metabolism enzyme inhibitors (ketoconazole or the recently available abiraterone acetate).

BACKGROUND

Benign prostatic hypertrophy increases with age and can result in substantially decreased quality of life for older men. Surgery is often required to control symptoms. It has been hypothesized that long-term administration of a nonamplifiable pure androgen might decrease prostate growth, thereby decreasing or delaying the need for surgical intervention.

OBJECTIVE

To test the hypothesis that dihydrotestosterone (DHT), a nonamplifiable and nonaromatizable pure androgen, reduces late-life prostate growth in middle-aged men.

METHODS

Randomized, placebo-controlled, parallel-group trial. (Australian New Zealand Clinical Trials Registry number: ACTRN12605000358640) SETTING: Ambulatory care research center.

METHODS

Healthy men (n = 114) older than 50 years without known prostate disease.

METHODS

Transdermal DHT (70 mg) or placebo gel daily for 2 years.

METHODS

Prostate volume was measured by ultrasonography; bone mineral density (BMD) and body composition were measured by dual-energy x-ray absorptiometry; and blood samples and questionnaires were collected every 6 months, with data analyzed by mixed-model analysis for repeated measures.

RESULTS

Over 24 months, there was an increase in total (29% [95% CI, 23% to 34%]) and central (75% [CI, 64% to 86%]; P < 0.01) prostate volume and serum prostate-specific antigen level (15% [CI, 6% to 24%]) with time on study, but DHT had no effect (P>> 0.2). Dihydrotestosterone treatment decreased spinal BMD (1.4% [CI, 0.6% to 2.3%]; P < 0.001) at 24 months but not hip BMD (P>> 0.2) and increased serum aminoterminal propeptide of type I procollagen in the second year of the study compared with placebo. Dihydrotestosterone increased serum DHT levels and its metabolites (5α-androstane-3α,17β-diol and 5α-androstane-3β,17β-diol) and suppressed serum testosterone, estradiol, luteinizing hormone, and follicle-stimulating hormone levels. Dihydrotestosterone increased hemoglobin levels (7% [CI, 5% to 9%]), serum creatinine levels (9% [CI, 5% to 11%]), and lean mass (2.4% [CI, 1.6% to 3.1%) but decreased fat mass (5.2% [CI, 2.6% to 7.7%]) (P <0.001 for all). Protocol-specific discontinuations due to DHT were asymptomatic increased hematocrit (n = 8), which resolved after stopping treatment, and increased prostate-specific antigen levels (n = 3; none with prostate cancer) in the DHT group. No serious adverse effects due to DHT occurred.

CONCLUSIONS

Negative findings on prostate growth cannot exclude adverse effects on the natural history of prostate cancer.

CONCLUSIONS

Dihydrotestosterone treatment for 24 months has no beneficial or adverse effect on prostate growth but causes a decrease in spinal but not hip BMD. These findings have important implications for the wider use of nonsteroidal pure androgens in older men.

BACKGROUND

BHR Pharma.

Androgens play an important role in regulation of body fat distribution in humans. They exert direct effects on adipocyte differentiation in a depot-specific manner, via the androgen receptor (AR), leading to modulation of adipocyte size and fat compartment expansion. Androgens also impact directly on key adipocyte functions including insulin signalling, lipid metabolism, fatty acid uptake and adipokine production. Androgen excess and deficiency have implications for metabolic health in both males and females, and these metabolic effects may be mediated through adipose tissue via effects on fat distribution, adipocyte function and lipolysis. Research into the field of androgen metabolism in human and animal adipose tissue has produced inconsistent results; it is important to take into account the sex-, depot- and organism-specific effects of androgens in fat. In general, studies point towards a stimulatory effect on lipolysis, with impairment of adipocyte differentiation, insulin signalling and adipokine generation. Observed effects are frequently gender-specific. Adipose tissue is an important organ of pre-receptor androgen metabolism, through which local androgen availability is rigorously controlled. Adipose androgen exposure is tightly controlled by isoenzymes of AKR1C, <em>5α</em>-reductase and others, but regulation of the balance between generation and irreversible inactivation remains poorly understood. In particular, AKR1C2 and AKR1C3 are crucial in the regulation of local androgen bioavailability within adipose tissue. These isoforms control the balance between activation of androstenedione (A) to testosterone (T) by the 17β-hydroxysteroid dehydrogenase activity (17β-HSD) of AKR1C3, or inactivation of <em>5α</em>-dihydrotestosterone (<em>DHT</em>) to <em>5α</em>-androstane-3α,17β-diol by the 3α-hydroxysteroid dehydrogenase (3α-HSD) activity of AKR1C2. Most studies suggest that androgen inactivation is the predominant reaction in fat, particularly in the abdominal subcutaneous (SC) depot. Modulation of local adipose androgen availability may afford future therapeutic options to improve metabolic phenotype in disorders of androgen excess and deficiency.

Intratumoral synthesis of dihydrotestosterone (<em>DHT</em>) from precursors cannot completely explain the castration resistance of prostate cancer. We showed that <em>DHT</em> was intratumorally synthesized from the inactive androgen metabolites <em>5α</em>-androstane-3α/β,17β-diol (3α/β-diol) in prostate cancer cells via different pathways in a concentration-dependent manner. Additionally, long-term culture in androgen-deprived media increased transcriptomic expression of 17β-hydroxysteroid dehydrogenase type 6 (HSD17B6), a key enzyme of oxidative 3α-HSD that catalyzes the conversion of 3α-diol to <em>DHT</em> in prostate cancer cells. Correspondingly, the score for HSD17B6 in tissues of 42 prostate cancer patients undergoing androgen deprivation therapy (ADT) was about 2-fold higher than that in tissues of 100 untreated individuals. In men receiving ADT, patients showing biochemical progression had a higher HSD17B6 score than those without progression. These results suggested that 3α/β-diol also represent potential precursors of <em>DHT</em>, and the back conversion of <em>DHT</em> from androgen derivatives can be a promising target for combination hormone therapy.

BACKGROUND

Increased risk of schizophrenia in adolescent males indicates that a link between the development of dopamine-related psychopathology and testosterone-driven brain changes may exist. However, contradictions as to whether testosterone increases or decreases dopamine neurotransmission are found and most studies address this in adult animals. Testosterone-dependent actions in neurons are direct via activation of androgen receptors (AR) or indirect by conversion to 17β-estradiol and activation of estrogen receptors (ER). How midbrain dopamine neurons respond to sex steroids depends on the presence of sex steroid receptor(s) and the level of steroid conversion enzymes (aromatase and <em>5α</em>-reductase). We investigated whether gonadectomy and sex steroid replacement could influence dopamine levels by changing tyrosine hydroxylase (TH) protein and mRNA and/or dopamine breakdown enzyme mRNA levels [catechol-O-methyl transferase (COMT) and monoamine oxygenase (MAO) A and B] in the adolescent male rat substantia nigra. We hypothesized that adolescent testosterone would regulate sex steroid signaling through regulation of ER and AR mRNAs and through modulation of aromatase and <em>5α</em>-reductase mRNA levels.

RESULTS

We find ERα and AR in midbrain dopamine neurons in adolescent male rats, indicating that dopamine neurons are poised to respond to circulating sex steroids. We report that androgens (T and DHT) increase TH protein and increase COMT, MAOA and MAOB mRNAs in the adolescent male rat substantia nigra. We report that all three sex steroids increase AR mRNA. Differential action on ER pathways, with ERα mRNA down-regulation and ERβ mRNA up-regulation by testosterone was found. <em>5α</em> reductase-1 mRNA was increased by AR activation, and aromatase mRNA was decreased by gonadectomy.

CONCLUSIONS

We conclude that increased testosterone at adolescence can shift the balance of sex steroid signaling to favor androgenic responses through promoting conversion of T to DHT and increasing AR mRNA. Further, testosterone may increase local dopamine synthesis and metabolism, thereby changing dopamine regulation within the substantia nigra. We show that testosterone action through both AR and ERs modulates synthesis of sex steroid receptor by altering AR and ER mRNA levels in normal adolescent male substantia nigra. Increased sex steroids in the brain at adolescence may alter substantia nigra dopamine pathways, increasing vulnerability for the development of psychopathology.

BACKGROUND

Concern exists that androgen treatment might adversely impact prostate health in older men. Dihydrotestosterone (<em>DHT</em>), derived from local conversion of testosterone to <em>DHT</em> by <em>5α</em>-reductase enzymes, is the principal androgen within the prostate. Exogenous androgens raise serum <em>DHT</em> concentrations, but their effects on the prostate are not clear.

OBJECTIVE

To determine the impact of large increases in serum DHT concentrations on intraprostatic androgen concentrations and androgen action within the prostate.

METHODS

Double-blind, randomized, placebo-controlled.

METHODS

Single academic medical center.

METHODS

31 healthy men ages 35-55.

METHODS

Daily transdermal DHT or placebo gel.

METHODS

Serum and prostate tissue androgen concentrations and prostate epithelial cell gene expression after 4 wk of treatment.

RESULTS

Twenty-seven men completed all study procedures. Serum DHT levels increased nearly sevenfold, while testosterone levels decreased in men treated with daily transdermal DHT gel but were unchanged in the placebo-treated group (P < 0.01 between groups). In contrast, intraprostatic DHT and testosterone concentrations on d 28 were not different between groups (DHT: placebo = 2.8 ± 0.2 vs. DHT gel = 3.1 ± 0.5 ng/g; T: placebo = 0.6 ± 0.2 vs. DHT gel = 0.4 ± 0.1, mean ± se). Similarly, prostate volume, prostate-specific antigen, epithelial cell proliferation, and androgen-regulated gene expression were not different between groups.

CONCLUSIONS

Robust supraphysiologic increases in serum DHT, associated with decreased serum T, do not significantly alter intraprostatic levels of DHT, testosterone, or prostate epithelial cell androgen-regulated gene expression in healthy men. Changes in circulating androgen concentrations are not necessarily mimicked within the prostate microenvironment, a finding with implications for understanding the impact of androgen therapies in men.

The enzyme steroid <em>5α</em> reductase (S<em>5α</em> R) catalyzes the conversion of Δ⁴-3-ketosteroid precursors--such as testosterone, progesterone and androstenedione--into their <em>5α</em>-reduced metabolites. Although the current nomenclature assigns five enzymes to the S<em>5α</em> R family, only the types 1 and 2 appear to play an important role in steroidogenesis, mediating an overlapping set of reactions, albeit with distinct chemical characteristics and anatomical distribution. The discovery that the <em>5α</em>-reduced metabolite of testosterone, <em>5α</em>-dihydrotestosterone (<em>DHT</em>), is the most potent androgen and stimulates prostatic growth led to the development of S<em>5α</em> R inhibitors with high efficacy and tolerability. Two of these agents, finasteride and dutasteride, have received official approval for the treatment of benign prostatic hyperplasia and are being tested for prevention of prostate cancer. Finasteride is also approved for male-pattern alopecia and has been shown to induce very limited side effects. Over the last decade, converging lines of evidence have highlighted the role of <em>5α</em>-reduced steroids and their precursors in brain neurotransmission and behavioral regulation. Capitalizing on these premises, we and other groups have recently investigated the role of S<em>5α</em> R in neuropsychiatric disorders. Our preliminary data suggest that S5 R inhibitors may elicit therapeutic effects in a number of disorders associated to dopaminergic hyperreactivity, including psychotic disorders, Tourette syndrome and impulse control disorders. In the present article, we review emerging preclinical and clinical evidence related to these effects, and discuss some of the potential mechanisms underlying the role of S<em>5α</em> R in the pathophysiology of mental disorders.

The androgen receptor (AR) acts as a ligand-dependent transcriptional factor controlling development or progression of prostate cancer. Androgen ablation by castration is an effective therapy for prostate cancer, whereas eventually most of the tumors convert from a hormone-sensitive to a hormone-refractory disease state and grow even in a low androgen environment (e.g., 0.1nM <em>5α</em>-dihydrotestosterone (<em>DHT</em>)) like the castration-resistant stage. Androgen ablation results in hypoxia, and solid tumors possess hypoxic environments. Hypoxia-inducible factor (HIF)-1, which is composed of HIF-1α and HIF-1β/ARNT subunits, functions as a master transcription factor for hypoxia-inducible genes. Here, we report that hypoxia enhances AR transactivation in the presence of 0.05 and 0.1nM <em>DHT</em> in LNCaP prostate cancer cells. siRNA-mediated knockdown of HIF-1α inhibited hypoxia-enhanced AR transactivation. Its inhibition by HIF-1α siRNA was canceled by expression of a siRNA-resistant form of HIF-1α. HIF-1α siRNA repressed hypoxia-stimulated expression of the androgen-responsive NKX3.1 gene in the presence of 0.1nM <em>DHT</em>, but not in the absence of <em>DHT</em>. In hypoxia, HIF-1α siRNA-repressed AR transactivation was restored in mutants in which HIF-1α lacked DNA-binding activity. Furthermore, a dominant negative form of HIF-1α canceled hypoxia-enhanced AR transactivation, and HIF-1β/ARNT siRNAs had no influence on hypoxia-enhanced AR transactivation. These results indicate that hypoxia leads to HIF-1α-mediated AR transactivation independent of HIF-1 activity and that HIF-1β/ARNT is not necessarily required for the transactivation.

Sex disparities in inflammation have been reported, but the cellular and molecular basis for these discrepancies is unknown. Monocytes are central effector cells in immunity and possess high capacities to produce proinflammatory leukotrienes (LTs). Here, we investigated sex differences in the activation of 5-lipoxygenase (5-LO), the key enzyme in LT biosynthesis, in human peripheral monocytes. In cells from females, 5-LO product formation was 1.8-fold higher than in cells from males, as evaluated by HPLC. When female monocytes were resuspended in plasma from males, 5-LO products were significantly lower than in female plasma. Interestingly, <em>5α</em>-dihydrotestosterone (<em>5α</em>-<em>DHT</em>, 10 nM) repressed LT synthesis in female cells down to the levels observed in males, while estradiol (100 nM) was without effect, and progesterone (100 nM) caused only a slight inhibition. <em>5α</em>-<em>DHT</em> (10 nM) caused ERK phosphorylation and inhibition of phospholipase D (PLD), as evaluated by Western blot and measurement of PLD activity via radioenzymatic diacylglyceride (DAG) and nonradioactive choline assays. Accordingly, PLD activity and DAG formation were 1.4- to 1.8-fold lower in male vs. female monocytes connected to increased ERK phosphorylation. Our data indicate that ERK activation by androgens in monocytes represses PLD activity, resulting in impaired 5-LO product formation due to lack of activating DAGs.

Testosterone is the most abundant circulating androgen, and can be converted to dihydrotestosterone (<em>DHT</em>), a more potent androgen, by the <em>5α</em>-reductase enzymes in target tissues. Current treatments for prostate cancer consist of reducing androgen levels by chemical or surgical castration or pure antiandrogen therapy that directly targets the androgen receptor (AR). Although these therapies reduce tumor burden and AR activity, the cancer inevitably recurs within 18-30 months. An approach targeting the androgen-AR axis at different levels could, therefore, improve the efficacy of prostate cancer therapy. Inhibition of <em>5α</em>-reductase is one such approach; however, the two largest trials to investigate the use of the <em>5α</em>-reductase inhibitors (5ARIs) finasteride and dutasteride in patients with prostate cancer have shown that, although the incidence of cancer was reduced by 5ARI treatment, those cancers that were detected were more aggressive than in patients treated with placebo. Thus, the best practice for using these drugs to prevent and treat prostate cancer remains unclear.

Androgens have dramatic effects on neuronal structure and function in hippocampus. However, androgen depletion does not always lead to hippocampal impairment. To address this apparent paradox, we evaluated the hippocampus of adult male rats after gonadectomy (Gdx) or sham surgery. Surprisingly, Gdx rats showed increased synaptic transmission and long-term potentiation of the mossy fiber (MF) pathway. Gdx rats also exhibited increased excitability and MF sprouting. We then addressed the possible underlying mechanisms and found that Gdx induced a long-lasting upregulation of MF BDNF immunoreactivity. Antagonism of Trk receptors, which bind neurotrophins, such as BDNF, reversed the increase in MF transmission, excitability, and long-term potentiation in Gdx rats, but there were no effects of Trk antagonism in sham controls. To determine which androgens were responsible, the effects of testosterone metabolites <em>DHT</em> and <em>5α</em>-androstane-3α,17β-diol were examined. Exposure of slices to 50 nm <em>DHT</em> decreased the effects of Gdx on MF transmission, but 50 nm <em>5α</em>-androstane-3α,17β-diol had no effect. Remarkably, there was no effect of <em>DHT</em> in control males. The data suggest that a Trk- and androgen receptor-sensitive form of MF transmission and synaptic plasticity emerges after Gdx. We suggest that androgens may normally be important in area CA3 to prevent hyperexcitability and aberrant axon outgrowth but limit MF synaptic transmission and some forms of plasticity. The results also suggest a potential explanation for the maintenance of hippocampal-dependent cognitive function after androgen depletion: a reduction in androgens may lead to compensatory upregulation of MF transmission and plasticity.

The cellular response to circulating sex steroids is more than the sum of individual hormone actions, instead representing an interplay between activities of the evolutionarily related steroid hormone receptors. An example of this interaction is in breast cancer, where the risk of dying from estrogen receptor-α (ERα)-positive disease decreases approximately 4-fold when androgen receptor (AR) expression is high. In this study, we used chromatin immunoprecipitation sequencing (ChIP-seq) and microarray expression profiling to investigate the genomic and transcriptional cross talk between AR and ERα signaling in a luminal breast cancer cell line model, ZR-75-1. Expression profiling demonstrated reciprocal interference between <em>5α</em>-dihydrotestosterone (<em>DHT</em>)- and 17β-estradiol (E(2))-induced transcriptional programs. Specifically, regulation of 26% of E(2) and 15% of <em>DHT</em> target genes was significantly affected by cotreatment with the other hormone, in the majority of cases (78-83%) antagonistically. Pathway analysis suggested that <em>DHT</em> cotreatment, for example, depleted E(2)-regulated pathways in cell survival and proliferation. ChIP-seq identified substantial overlap between the steroid receptor cistromes in ZR-75-1 cells, with 10-13% of AR- and ERα-binding sites located within 10 kb of the other receptor. Enrichment of androgen response elements in ERα-binding sites and vice versa was revealed by motif analysis, and AR-binding sites were enriched about E(2)-responsive genes affected by <em>DHT</em> cotreatment. Targeted ChIP and expression analysis revealed locus-specific outcomes when AR and ERα bind to the same DNA region. This work provides the first cistrome data for two steroid receptors in the same cell, insight into the antagonistic interplay between estrogens and androgens in luminal breast cancer, and an important resource for future work aimed at evaluating interrelated steroid receptors in different cellular systems.