Androgen Production Research Articles

Impact of Chromium Picolinate on Leydig Cell Steroidogenesis and Antioxidant Balance Using an In Vitro Insulin Resistance Model.

Leydig cells (LCs) play a pivotal role in male fertility, producing testosterone. Chromium (III) picolinate (CrPic3), a contentious supplement with antidiabetic and antioxidant properties, raises concerns regarding male fertility. Using a rodent LC line, we investigated the cytotoxicity of increasing CrPic3 doses. An insulin resistance (IR) model was established using palmitate (PA), and LCs were further exposed to CrPic3 to assess its antioxidant/antidiabetic activities. An exometabolome analysis was performed using 1H-NMR. Mitochondrial function and oxidative stress were evaluated via immunoblot. Steroidogenesis was assessed by quantifying androstenedione through ELISA. Our results uncover the toxic effects of CrPic3 on LCs even at low doses under IR conditions. Furthermore, even under these IR conditions, CrPic3 fails to enhance glucose consumption but restores the expression of mitochondrial complexes CII and CIII, alleviating oxidative stress in LCs. While baseline androgen production remained unaffected, CrPic3 promoted androstenedione production in LCs in the presence of PA, suggesting that it promotes cholesterol conversion into androgenic intermediates in this context. This study highlights the need for caution with CrPic3 even at lower doses. It provides valuable insights into the intricate factors influencing LCs metabolism and antioxidant defenses, shedding light on potential benefits and risks of CrPic3, particularly in IR conditions.

Current Update on the Management and Treatment of Congenital Adrenal Hyperplasia

Congenital Adrenal Hyperplasia (CAH) comprises a set of inherited conditions marked by enzymatic deficiencies within the adrenal steroidogenesis pathway, resulting in compromised cortisol synthesis and, in certain instances, heightened androgen production. Congenital adrenal hyperplasia (CAH) arises from autosomal recessive genetic mutations affecting enzymes crucial for the adrenal glands’ biochemical synthesis of glucocorticoids, mineralocorticoids, or sex steroids from cholesterol. The majority of these conditions result in either excessive or deficient production of sex steroids, potentially influencing the development of primary or secondary sex characteristics in affected infants, children, or adults. The management and treatment of CAH are geared towards optimizing hormonal equilibrium, averting adrenal crises, and addressing associated comorbidities. This review presents an updated survey of contemporary approaches to CAH management, encompassing glucocorticoid and mineralocorticoid replacement therapies, along with strategies to alleviate androgen excess. Furthermore, it is required that we look into emerging treatments, including novel pharmaceutical interventions and advancements in genetic and reproductive technologies. The complexities and possibilities inherent in the comprehensive care of individuals with CAH, involving collaboration among endocrinologists, geneticists, and reproductive specialists, are also examined. This review presents an updated survey of contemporary approaches to CAH management, encompassing glucocorticoid and mineralocorticoid replacement therapies, along with strategies to alleviate androgen excess. By scrutinizing recent advancements, this abstract provides valuable insights into the dynamic landscape of CAH management, emphasizing the imperative for sustained research efforts to augment the well-being of affected individuals.

Antiproliferative activity of the novel CYP17A1 inhibitor alsevirone

BACKGROUND: Prostate cancer is the most frequently diagnosed type of cancer in men in developed countries. It is dependent upon androgens and could be effectively combated by androgen deprivation therapy. Reduction of androgen synthesis can be accomplished through the inhibition of the enzyme 17α-hydroxylase/17.20-lyase (CYP17A1), which catalyzes two sequential reactions in the production of androgens. Steroid derivatives modified with nitrogen-containing heterocycles attract attention as antineoplastic agents for prostate cancer treatment due to their inhibitory potential against CYP17A1.
 AIM: Evaluate cytotoxic activity and antitumor effects of the synthesized alsevirone in comparison with abiraterone.
 METHODS: Cytotoxicity was evaluated using MTT test. Anticancer effect was researched in vivo in prostate cancer xenograft models 22Rv1 and DU145 in Balb/c nude mice. Testosterone concentration was determined using an enzyme-linked immunosorbent assay in blood serum of BDF1 mice.
 RESULTS: Alsevirone demonstrated cytotoxic activity in prostate cancer cells: DU145 (23.8±1.2 µM vs 151.4±23.7 µM for abiraterone), 22Rv1 (35.9±5.6 µM vs 109.9±35.2 µM for abiraterone) and LNCaP (22.9±0.5 µM vs 28.8±1.6 µM for abiraterone). Testosterone concentration in blood serum of BDF1 mice reduced by 80% after 10-day treatment. Inhibition of the tumors’ growth in 22Rv1 xenograft model was statistically significant when using alsevirone in comparison with the control group: average tumor volume was 171.6±50.1 mm3 (р=0.022) vs 424.2±70.3 mm3 in control, with tumor growth inhibition index of 59%.
 CONCLUSIONS: Alsevirone has a higher cytotoxic potential against prostate cancer cells (DU145, LNCaP and 22Rv1) compared to abiraterone. Alsevirone demonstrated the ability to reduce the concentration of testosterone in the blood serum of BDF1 mice, and statistically significant antitumor activity in 22Rv1 xenograft models.

Integrated Studies on Male Reproductive Toxicity of Bis(2-ethylhexyl)-tetrabromophthalate: in Silico, in Vitro, ex Vivo, and in Vivo.

Bis(2-ethylhexyl)tetrabromophthalate (TBPH) has been widely detected in the environment and organisms; thus, its toxic effects on male reproduction were systematically studied. First, we found that TBPH can stably bind to the androgen receptor (AR) based on in silico molecular docking results and observed an antagonistic activity, but not agonistic activity, on the AR signaling pathway using a constructed AR-GRIP1 yeast assay. Subsequently, we validated the adverse effects on male germ cells by observing inhibited androgen production and proliferation in Leydig cells upon in vitro exposure and affected general motility and motive tracks of zebrafish sperm upon ex vivo exposure. Finally, the in vivo reproductive toxicity was demonstrated in male zebrafish by reduced mating behavior in F0 generation when paired with unexposed females and abnormal development of their offspring. In addition, reduced sperm motility and impaired germ cells in male zebrafish were also observed, which may be related to the disturbed homeostasis of sex hormones. Notably, the specifically suppressed AR in the brain provides further evidence for the antagonistic effects as above-mentioned. These results confirmed that TBPH affected male reproduction through a classical nuclear receptor-mediated pathway, which would be helpful for assessing the ecological and health risks of TBPH.

Inhibition of aldo-keto reductase 1C3 overcomes gemcitabine/cisplatin resistance in bladder cancer

Systemic chemotherapy with gemcitabine and cisplatin (GC) has been used for the treatment of bladder cancer in which androgen receptor (AR) signaling is suggested to play a critical role. However, its efficacy is often limited, and the prognosis of patients who develop resistance is extremely poor. Aldo-keto reductase 1C3 (AKR1C3), which is responsible for the production of a potent androgen, 5α-dihydrotestosterone (DHT), by the reduction of 5α-androstane-3α,17β-dione (5α-Adione), has been attracting attention as a therapeutic target for prostate cancer that shows androgen-dependent growth. By contrast, the role of AKR1C3 in bladder cancer remains unclear. In this study, we examined the effect of an AKR1C3 inhibitor on androgen-dependent proliferation and GC sensitivity in bladder cancer cells. 5α-Adione treatment induced the expression of AR and its downstream factor ETS-domain transcription factor (ELK1) in both T24 cells and newly established GC-resistant T24GC cells, while it did not alter AKR1C3 expression. AKR1C3 inhibitor 2j significantly suppressed 5α-Adione-induced AR and ELK1 upregulation, as did an AR antagonist apalutamide. Moreover, the combination of GC and 2j in T24GC significantly induced apoptotic cell death, suggesting that 2j could enhance GC sensitivity. Immunohistochemical staining in surgical specimens further revealed that strong expression of AKR1C3 was associated with significantly higher risks of tumor progression and cancer-specific mortality in patients with muscle-invasive bladder cancer. These results suggest that AKR1C3 inhibitors as adjunctive agents enhance the efficacy of GC therapy for bladder cancer.

Steroidogenesis and androgen/estrogen signaling pathways are altered in in vitro matured testicular tissues of prepubertal mice

Children undergoing cancer treatments are at risk for impaired fertility. Cryopreserved prepubertal testicular biopsies could theoretically be later matured in vitro to produce spermatozoa for assisted reproductive technology. A complete in vitro spermatogenesis has been obtained from mouse prepubertal testicular tissue, although with low efficiency. Steroid hormones are essential for the progression of spermatogenesis, the aim of this study was to investigate steroidogenesis and steroid signaling in organotypic cultures. Histological, RT-qPCR, western blot analyses, and steroid hormone measurements were performed on in vitro cultured mouse prepubertal testicular tissues and age-matched in vivo controls. Despite a conserved density of Leydig cells after 30 days of culture (D30), transcript levels of adult Leydig cells and steroidogenic markers were decreased. Increased amounts of progesterone and estradiol and reduced androstenedione levels were observed at D30, together with decreased transcript levels of steroid metabolizing genes and steroid target genes. hCG was insufficient to facilitate Leydig cell differentiation, restore steroidogenesis, and improve sperm yield. In conclusion, this study reports the failure of adult Leydig cell development and altered steroid production and signaling in tissue cultures. The organotypic culture system will need to be further improved before it can be translated into clinics for childhood cancer survivors.

Testicular regression and recrudescence in the bat Eptesicus furinalis: Morpho-physiological variations and hormonal signaling pathways.

Males of the bat Eptesicus furinalis show at least one process of testicular regression, in which the testes regress and temporarily interrupt the production of sperm, during its annual reproductive cycle. As the process of spermatogenesis is under hormonal control, mainly of pituitary and androgen hormones, our aim was to analyze the morphological variations and the hormonal control of the testes of E. furinalis during the four phases of its reproductive cycle. Testes of 18 adult males, divided into four sample groups (active, regressing, regressed, and recrudescence phases), were submitted to morphological, morphometric, and immunohistochemical analyzes. The results demonstrate that the processes of testicular regression and recrudescence of E. furinalis are under the control of pituitary, androgen and estrogen hormones. The regulation is exerted mainly through the activation and cross signaling of AR and FSHR in Sertoli cells and of LHR in Leydig cells. The testicular regression appears to be activated by an inhibition/reduction of AR expression in Sertoli cells, which inhibits the proliferation and differentiation of new spermatogonia and causes the deactivation of spermatogenesis. Conversely, the testicular recrudescence occurs by the increasing of the expression of LHR in Leydig cells, and AR and FSHR in Sertoli cells, which reactivates the testicular production of androgens and estrogens, the proliferation of spermatogonia and restarts the spermatogenesis.

Elevated luteinizing hormone receptor signaling or selenium treatment leads to comparable changes in adrenal cortex histology and androgen-AR/ZIP9 signaling

The importance and regulation of adrenal androgen production and signaling are not completely understood and are scarcely studied. In addition, there is still a search for appropriate animal models and experimental systems for the investigation of adrenal physiology and disease. Therefore, the main objective of the study was to evaluate the effect of luteinizing hormone (LH) signaling and selenium (Se2+) exposure on androgen adrenal signaling via canonical androgen receptor (AR), and membrane androgen receptor acting as zinc transporter (zinc- and iron-like protein 9; ZIP9). For herein evaluations, adrenals isolated from transgenic mice with elevated LH receptor signaling (KiLHRD582G) and adrenals obtained from rabbits used for ex vivo adenal cortex culture and exposure to Se2+ were utilized. Tissues were assessed for morphological, morphometric, and Western blot analyses and testosterone and zinc level measurements.Comparison of adrenal cortex histology and morphometric analysis in KiLHRD582G mice and Se2+-treated rabbits revealed cell hypertrophy. No changes in the expression of proliferating cell nuclear antigen (PCNA) were found. In addition, AR expression was decreased (p < 0.001) in both KiLHRD582G mouse and Se2+-treated rabbit adrenal cortex while expression of ZIP9 showed diverse changes. Its expression was increased (P < 0.001) in KiLHRD582G mice and decreased (P < 0.001) in Se2+-treated rabbits but only at the dose 10 ug/100 mg/ tissue. Moreover, increased testosterone levels (P < 0.05) and zinc levels were detected in the adrenal cortex of KiLHRD582G mice whereas in rabbit adrenal cortex treated with Se2+, the effect was the opposite (P < 0.001).

Abstract B034: Characterization of a dual steroid 3β-,17β-oxidoreductase in the gut bacterium Eggerthella lenta

Abstract As of 2023 there are over 200,000 new cases and over 30,000 deaths caused by prostate cancer with African-American (AA) men having a higher incidence then other ethnicities according to the American Cancer Society. Although numerous therapeutic methods are used to treat prostate cancer such as hormone therapy, radiation therapy, chemotherapy, and chemical castration, prostate cancer often becomes metastatic resulting in death with the five year survival rate for AA men presenting with castration resistant prostate cancer (CRPC) being ~ 32%. Androgen production is known to be the main driver of CRPC progression. Researchers are baffled as to why this progression is taking place after patients have received a combination of these therapeutic methods to stop androgen production and metabolism. Eggerthella lenta has a long history as an important bile acid and steroid metabolizing gut microbial species. We and others have previously published on the extensive oxidation and epimerization of bile acid hydoxyl groups. Previous work demonstrated that E. lenta may be involved in the conversion of 17-keto steroids to testosterone. By cloning, expressing, and purifying genes predicted to encode pyridine nucleotide-dependent oxidoreductases from E. lenta DSM 2243, we identified a recombinant enzyme with dual 3beta, 17beta-hydroxysteroid dehydrogenase activity. We performed kinetic analysis on both 5alpha-reduced bile acids ("allo"-bile acids) and 5alpha-reduced steroids. There is a paucity of work at present on gut bacteria that metabolize the 17beta-hydroxy group on steroids, so this observation demonstrates that a unique enzyme in the gut microbiome can alter host steroids. These findings are consistent with the possibility that gut or urinary tract bacteria may contribute to CRPC. Citation Format: Briawna J. Binion, Rex Gaskins, Jason Ridlon. Characterization of a dual steroid 3β-,17β-oxidoreductase in the gut bacterium Eggerthella lenta [abstract]. In: Proceedings of the 16th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2023 Sep 29-Oct 2;Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2023;32(12 Suppl):Abstract nr B034.

EFFECT OF HYPOXIC SECRETOME MESENCHYMAL STEM CELLS ON TGF-β EXPRESSION, ESTROGEN HORMONE LEVELS, AND NUMBER OF FOLLICLES (IN-VIVO EXPERIMENTAL STUDY ON FEMALE WISTAR RATS POLYCYSTIC OVARY SYNDROME-LIKE MODELS)

PCOS develops when the ovaries are stimulated to produce more androgen hormones so that the development of follicle count is disrupted. Elevated insulin levels contribute directly to abnormalities in the pituitary hypothalamus of the ovaries which have a direct impact on the occurrence of PCOS by hyperinsulinemia mechanism that increases the frequency of GnRH (Gonadotropin Releasing Hormone) secretion, LH (Luteinizing Hormone) production exceeds FSH, increased ovarian androgen production, decreased follicle count maturation, and increased TGF-β (transforming growth factor β) expression. This study aimed to determine the effect of SH-MSCs administration on TGF-β expression, estrogen hormone levels, and follicle count in female Wistar strain rats with PCOS-like. This study is an in vivo experimental study using a post-test-only control group design. The subjects of this study were female Wistar strain rats divided into 4 treatments consisting of 1 group of healthy mice, 2 negative control groups, 3 treatment groups 1, and treatment group 2 with each different dose. Then the data was analyzed using the One Way Annova Test to determine the influence of each group.One Way ANOVA test showed the results of TGF-β expression (p = 0.000), then the results of estrogen hormone levels (p = 0.000), then the Kruskall Wallis test showed the number of cystic follicles (p = 0.045) in all groups there was a significant difference (p &lt; 0.05) except cystic follicle.There is an effect of SH-MSCs secretome administration on decreasing TGF-β expression and caustic follicle count. There is an effect of SH-MSC administration on increasing estrogen hormone levels and corpus luteum count.

The Effect of BMI and Lipid Metabolism on Pregnancy Outcomes of PCOS Patients Treated with ICSI_IVF.

Polycystic ovary syndrome is a highly prevalent and important cause of infertility. This complication demonstrates relationships with obesity, increased androgen production, and insulin resistance. The typical pattern of dyslipidemia in women with PCOS is a decrease in high-density lipoprotein and an increase in triglycerides. On the other hand, fat people are at a higher risk of hyperandrogenism, insulin resistance, hypercholesterolemia, hypertriglyceridemia, and infertility compared to ordinary people. In this cross-sectional study, 150 patients under 40 years suffering from PCOS based on the Rotterdam criteria, who were candidates for in vitro fertilization (IVF), were examined at Yas Hospital (Tehran, Iran) from April to September 2022. In addition to demographic information, lipid profile, body mass index, obstetric, fasting blood sugar (FBS), beta HCG (negative or positive), and ultrasound were also checked and pregnancy status was checked six weeks later. The rate of clinical and chemical pregnancy in women with a normal BMI was about two times that of women with a BMI above 25kg/m2. In addition, the pregnancy rate in women with dyslipidemia was significantly lower than that in ordinary women. The amount of FBS was substantially lower in people with positive pregnancies. The probability of pregnancy in PCOS women with a BMI is higher than normal women, and dyslipidemia is lower than that in ordinary people. Therefore, enhancing BMI and dyslipidemia in these women before starting infertility treatment is recommended.

289 Postnatal Obesity Amplifies the Effect of Prenatal Testosterone Excess in Reducing the Progesterone Negative Feedback Sensitivity in First-Generation Ewes

Abstract Polycystic ovary syndrome (PCOS) is the leading cause of anovulatory infertility in women of reproductive age. Women with PCOS exhibit reduced sensitivity of the GnRH neuronal system to the inhibitory effects of both progesterone and estradiol. Additionally, obesity amplifies the severity of the PCOS phenotype. In sheep, testosterone (T) excess during gestational days (GD) 30-90 and 60-90 reduces neuroendocrine sensitivity to the progesterone negative feedback and recapitulates the PCOS phenotype. This study investigated the contribution of postnatal obesity in amplifying or mitigating the effects of mid-gestational T excess in disrupting the responsiveness to progesterone negative feedback in sheep. Suffolk ewes received T propionate (T; 100 mg i.m.) or corn oil (CO; vehicle) twice weekly from GD 60-90 (term = 147 d). At 5 mo of age, F1 T lambs were assigned randomly to either a maintenance (100% of NRC requirements) or overfed diet (130% of NRC requirements) and control lambs were fed the maintenance diet. At 22 mo of age, F1 ewes (n = 8/group) were synchronized with two injections of PGF2α 11 d apart. After the second PGF2α administration, blood samples were collected daily for 14 d and progesterone concentrations were determined via radioimmunoassay. Ewes that were confirmed to be cycling and in the mid-luteal phase based on progesterone concentrations (progesterone ≥ 1 ng/ml on day 10 and ≥ 0.5 ng/ml on days 8-9 and 11-12) were used for LH pulse characterization. On day 10 after PGF2α synchronization (mid-luteal phase), blood samples were collected every 15 min for 6 h to characterize LH pulsatile secretion via radioimmunoassay. Data were analyzed as one-way ANOVA with Tukey's HSD post-hoc analysis. Frequency of LH pulses was greater (P &amp;lt; 0.05) in T maintenance (3.75 ± 0.41 pulses/6h) and T overfed ewes (3.50 ± 0.34 pulses/6h) compared with controls (2.00 ± 0.37 pulses/6 h). Luteinizing hormone pulse amplitude was greater (P &amp;lt; 0.05) in T overfed (2.03 ± 0.15 ng/mL) compared with T maintenance (1.44 ± 0.17 ng/mL), but not to controls (1.74 ± 0.21 ng/mL). Also, average LH concentration was greater (P &amp;lt; 0.05) in T overfed (2.03 ± 0.24 ng/mL), but not T maintenance (1.31 ± 0.35 ng/mL), compared with controls (0.93 ± 0.30 ng/mLl). In conclusion, these findings demonstrate the activational role of postnatal obesity in amplifying prenatal T-induced alterations in tonic LH secretion during progesterone negative feedback. The activational effect of postnatal obesity in increasing LH secretion and consequently thecal androgen production is likely a contributor to the severity of hyperandrogenic status in women with PCOS. Research support: NIH-NICHD (R01HD099096).

Oocyte-derived growth differentiation factor 9 suppresses the expression of CYP17A1 and androgen production in human theca cells

ObjectiveTo investigate the direct effect of growth differentiation factor 9 (GDF9) on androgen production in human theca cells. DesignExperimental study SettingTertiary hospital-based research laboratory Patient(s)Women underwent IVF/intracytoplasmic sperm injection (ICSI) at our clinic were included in this study. Intervention(s)Primary cultured human theca cells from women undergoing IVF/ICSI treatment were treated with GDF9, Activin receptor-like kinase 5 (ALK5) inhibitor, and SMAD4 agonist. Main Outcome Measure(s)The expression of androgen synthesis-related gene StAR, CYP17A1, and LHCGR, levels of androstenedione and testosterone, phosphorylation of SMAD2/3 and the interaction between bone morphogenetic protein receptor II (BMPRII) and ALK5 were evaluated by RT-qPCR, Western blot, enzyme-linked immunosorbent assays, and co-immunoprecipitation assays respectively. ResultsGDF9 decreased StAR, CYP17A1, and LHCGR expression levels in human theca cells, which was prevented by the treatment with the ALK5 inhibitor, and suppressed production of androgen in human theca cells. GDF9 increased SMAD2/3 phosphorylation, and the ALK5 inhibitor also suppressed this effect. BMPRII and ALK5 bound to each other following GDF9 stimulation. The SMAD4 agonist kartogenin also decreased mRNA levels of StAR and CYP17A1, and protein levels of StAR in human theca cells. ConclusionGDF9 can activate the BMPRII-ALK5-SMAD2/3 signaling pathway, suppress CYP17A1 expression and androgen production in human theca cells.

Promising drug candidates for the treatment of polycystic ovary syndrome (PCOS) as alternatives to the classical medication metformin

Polycystic ovary syndrome (PCOS) is a prevalent hormonal disorder that affects women of reproductive age. It is characterized by abnormal production of androgens, typically present in small quantities in females. This study aimed to investigate the therapeutic potential of Irosustat (STX64), STX140, and compound 1G as new drug candidates for the treatment of letrozole-induced PCOS in female Wistar rats. 36 rats were divided into six groups of equal size. PCOS was induced in all groups, except the normal control group, by administering letrozole orally (1 mg/kg/day for 35 days). The onset of abnormal estrous cycle was confirmed by examining daily vaginal smears under a microscope. Subsequently, each rat group was assigned to a different treatment regimen, including one control group, one letrozole group, one metformin group (500 mg/kg/day) as a reference drug, and the other groups received a different drug candidate orally for 30 days. After treatment, blood collection was performed for biochemical measurements and determination of oxidative stress markers. The rats were dissected to separate ovaries and uterus for morphological, histological, and western blotting studies. Treatment with the drug candidates improved the ovaries and uterus weight measurements compared to the untreated PCOS group. The three tested drug candidates demonstrated promising improvements in lipid profile, blood glucose level, testosterone, progesterone, luteinizing hormone (LH), follicle-stimulating hormone (FSH), and estradiol levels. In addition, western blotting confirmed their promising effects on Akt, mTOR, and AMPK-α pathways. This study led to the discovery of three promising drug candidates for the management of PCOS as alternatives to metformin.

Semaphorin 3C promotes de novo steroidogenesis in prostate cancer cells.

Intratumoral androgen biosynthesis contributes to castration-resistant prostate cancer progression in patients treated with androgen deprivation therapy. The molecular mechanisms by which castration-resistant prostate cancer acquires the capacity for androgen biosynthesis to bypass androgen deprivation therapy are not entirely known. Here, we show that semaphorin 3C, a secreted signaling protein that is highly expressed in castration-resistant prostate cancer, can promote steroidogenesis by altering the expression profile of key steroidogenic enzymes. Semaphorin 3C not only upregulates enzymes required for androgen synthesis from dehydroepiandrosterone or de novo from cholesterol but also simultaneously downregulates enzymes involved in the androgen inactivation pathway. These changes in gene expression correlate with increased production of androgens induced by semaphorin 3C in prostate cancer model cells. Moreover, semaphorin 3C upregulates androgen synthesis in LNCaP cell-derived xenograft tumors, likely contributing to the enhanced in vivo tumor growth rate post castration. Furthermore, semaphorin 3C activates sterol regulatory element-binding protein, a transcription factor that upregulates enzymes involved in the synthesis of cholesterol, a sole precursor for de novo steroidogenesis. The ability of semaphorin 3C to promote intratumoral androgen synthesis may be a key mechanism contributing to the reactivation of the androgen receptor pathway in castration-resistant prostate cancer, conferring continued growth under androgen deprivation therapy. These findings identify semaphorin 3C as a potential therapeutic target for suppressing intratumoral steroidogenesis.

FRI435 A Grave Case Of Gynecomastia

Abstract Disclosure: S. Seltzer: None. A. illiams: None. T. Kader: None. Background: Gynecomastia is a common endocrinology referral. Our team saw a patient in the clinic with an unusual presentation. Clinical Case: A 28 year old male presented with worsening painful gynecomastia affecting one breast initially but eventually progressing to both breasts. His secondary concern was a 40lb weight loss and new onset tremor. He declined hypogonadal symptoms, palpitations, or new testicular growth. Exam revealed a resting pulse of 120 b.p.m., a palpable goiter, visible tremor, hyperdynamic heart sounds and bilateral tender gynecomastia. Labs revealed normal FSH, LH and hCG, high total testosterone (TT), thyroxine (T4) and triiodothyronine (T3) with positive thyroid receptor antibodies. Methimazole and beta blockade were started. He did develop an episode of atrial fibrillation requiring admission. Due to medication intolerance and patient preference the decision was made for total thyroidectomy. Three months post-surgery there was complete resolution of hyperthyroidism and gynecomastia.The differential for true gynecomastia is broad. The initial diagnostic approach includes a thorough history and physical exam followed by measuring serum testosterone (T), LH, hCG and estradiol (E2). Increased LH and low T indicates hypogonadism. Normal values may suggest an idiopathic cause. Elevated hCG warrants testicular imaging and if negative, chest and abdominal imaging to rule out extragonadal germ cell tumors. Elevated total T and LH should prompt measurement of TSH and T4 and if normal androgen resistance should be suspected. Elevated E2 and low/low normal LH should prompt testicular imaging and if normal an adrenal CT to rule out an adrenal neoplasm. If LH and T are low, prolactin should be measured as well as dedicated pituitary imaging.Any state which induces an increased E to T ratio may cause gynecomastia. The true pathophysiology in hyperthyroidism driving the increased sex hormone binding globulin (SHBG) production is unknown. It is postulated that this increase is driven by increased transcription and translation of hepatocyte nuclear factor-4ɑ secondary to elevated T41. SHBG more readily binds androgens than estrogens leading to a relative decrease in free T (FT). The lower FT levels result in negative feedback to the pituitary gland which causes increases in LH resulting in production of androgens, estradiol and peripheral aromatases. The increase in peripheral aromatization of testosterone leads to further elevation of the free estrogen levels. Conclusion: Hyperthyroidism should always be in the differential of gynecomastia as in our case the presentation may be atypical.

FRI376 Regulation Of Theca Cell Function By Salt-Inducible Kinases

Abstract Disclosure: M. Rodriguez Esquivel: None. E. Hayes: None. O. Lakomy: None. M. Hassan: None. R. Filzen: None. C.O. Stocco: Grant Recipient; Self; NIH R01HD097202. The most common identifiable factors of female infertility are ovulatory disorders. Ovulation is the pinnacle of folliculogenesis, a process that requires an interplay between the oocyte, the granulosa cells, and the theca cells (TCs). TCs are the only source of ovarian androgens. Androgens play vital roles in female fertility acting as substrates for estrogen production in granulosa cells and directly regulating ovarian function via the activation of the androgen receptor. However, abnormally elevated androgen levels reduce fertility in humans and animals, suggesting that a delicate equilibrium exists between the beneficial and harmful effects of androgens in the ovary. Therefore, uncovering novel mechanisms regulating androgen synthesis in TCs is of great significance. The capacity of TCs to synthesize androgens relies on the expression of the rate-limiting enzyme CYP17A1, which converts progestogens into androgens, and its stimulation by the luteinizing hormone (LH). Previously, our group showed that salt-inducible kinases (SIKs) regulate granulosa cell steroidogenesis. Here, we investigated whether SIKs play any role in the regulation of androgen production in TCs. SIK2 and SIK3 were detected by IHC in the TCs of rat ovaries. Western blot confirmed this finding using isolated rat TCs. Next, rat TCs in culture were treated with LH in the presence or absence of HG, a highly specific SIK inhibitor. SIK inhibition enhanced the stimulatory effect of LH on Cyp17a1 mRNA expression. HG alone also increased Cyp17a1 expression to levels higher than those found in the presence of LH. Similarly, HG alone increased testosterone and potentiated LH stimulation of testosterone production. SIK inhibition also enhanced the stimulatory effect of LH on the expression of StAR and Cyp11a1. Concentration-response experiments for the SIK inhibitor in the presence or absence of LH show that SIK inhibition stimulates Cyp17a1 in a concentration-dependent manner from 0.1 to 1 µM. Of note, at 3 µM, HG was less effective in inducing Cyp17a1 than at 1 µM, suggesting that higher concentrations of HG may have alternative unknown effects. Activation of adenyl cyclase with forskolin or emulation of increased intracellular cAMP with dbcAMP stimulated Cyp17a1 an effect that was also enhanced by the inhibition of SIK. Since the main target of cAMP is PKA, we used lentivirus to express constitutively active PKA (caPKA). Overexpression of caPKA was sufficient to stimulate testosterone production, an effect that was significantly augmented by SIK inhibition, suggesting that SIKs regulate factors downstream of PKA. This evidence shows that TCs express SIKs and reveal novel roles for SIKs in the regulation of TC function and androgen production. This information could contribute to uncovering therapeutic targets to treat hyperandrogenic diseases such as PCOS. Presentation: Friday, June 16, 2023

SAT024 Contribution of 11-Ketotestosterone and 11-Ketodihydrotestosterone to Castration-Resistant Prostate Cancer

Abstract Disclosure: H.S. Abu-Lebdeh: None. H.M. Heshmati: None. Background: Prostate cancer is among the most common malignancy in men and represents the second most common cause of cancer-related mortality among men in the US. Androgen-deprivation therapy with surgical or medical castration is widely used to treat hormone-sensitive prostate cancer. Disease progression despite androgen-deprivation therapy is a state defined as castration-resistant prostate cancer that is associated with poor survival rate. This review presents an update on the contribution of two adrenal androgens, 11-ketotestosterone and 11-ketodihydrotestosterone, to the castration-resistant prostate cancer. Methods: A systematic search of literature was conducted using the search terms prostate cancer, testosterone, dihydrotestosterone, adrenal androgens, 11-ketotestosterone, 11-ketodihydrotestosterone, androgen-deprivation therapy, castration, and castration-resistant prostate cancer. Results: In 2022, around 268,490 new cases of prostate cancer were diagnosed in the US with a mortality of 34,500 cases. Although androgen-deprivation therapy with castration is efficient at the beginning, most patients develop a state of castration resistance usually after 2-3 years as a result of the reactivation of androgen-receptor signaling despite low testosterone levels. The adrenal glands produce significant amounts of steroid hormones and metabolites. Among them, 11-ketotestosterone and 11-ketodihydrotestosterone are potent and efficacious androgens with potentially longer activity compared to testosterone and dihydrotestosterone because they are metabolized at a lower rate. Dihydrotestosterone derived from the metabolism of adrenal androgens (e.g., androstenedione and dehydroepiandrosterone) and especially 11-ketotestosterone and 11-ketodihydrotestosterone are important contributors to castration-resistant prostate cancer. In these cases, adrenal suppression with drugs causing a decrease in adrenal androgen production may improve the effectiveness of the androgen-deprivation therapy. Conclusion: In most patients with prostate cancer, androgen-deprivation therapy loses its effectiveness after 2-3 years leading to a state of castration-resistant prostate cancer. Two potent and highly efficacious androgen-receptor agonists of adrenal origin, 11-ketotestosterone and 11-ketodihydrotestosterone, are predominant androgens after castration and play an important role in the development and progression of castration-resistant prostate cancer. Their measurement in blood can be of clinical value during androgen-deprivation therapy. Concomitant treatment aimed to block the production of adrenal androgens is of potential interest in the management of these patients, but more studies are necessary to further investigate the benefits and adverse effects of this approach. Presentation: Saturday, June 17, 2023

SAT286 Effect of Tildacerfont On Gonadotropins And Testicular Steroidogenesis In Men With Classic 21-hydroxylase Deficiency

Abstract Disclosure: R. Charlton: Employee; Self; Spruce Biosciences, Inc. A. Khattab: Consulting Fee; Self; Antares Pharma, Inc. R.J. Auchus: Grant Recipient; Self; Spruce Biosciences, Inc. Research Investigator; Self; Spruce Biosciences, Inc. Impaired testicular function is a common comorbidity of classic 21-hydroxylase deficiency (21OHD). Testicular adrenal rest tumors (TART) can disrupt testicular architecture and cause primary hypogonadism. Both supraphysiologic glucocorticoid doses and adrenal-derived androgens and estrogens can suppress gonadotropins and cause secondary hypogonadism. We conducted a post-hoc analysis of gonadal axis function in men with 21OHD participating in SPR001-201 and SPR001-202, two phase 2 studies of tildacerfont, a once-daily oral corticotrophin-releasing factor receptor 1 (CRFR1) antagonist, designed to reduce ACTH and adrenal steroid production, including androstenedione (A4). Study 201 included 3 cohorts: Cohort A, 5 men who received tildacerfont in a 6-week dose escalation - 2 weeks at 200 mg QD, 2 weeks at 600 mg QD, and 2 weeks at 1000 mg QD without washout between doses; Cohort B, 1 man who received tildacerfont for 2 weeks at 200 mg BID; and Cohort C, 4 men who received tildacerfont for 2 weeks at 100 mg BID. Study 202 included 4 men who received tildacerfont 400 mg QD for 12 weeks. Participants continued their previously prescribed glucocorticoid ± mineralocorticoid replacement without dose adjustments. Geometric means were used to describe hormone levels due to the small participant numbers. Mean and median ages were 46/32y in Study 201 and 32/35y in Study 202 (range 18-54 in both studies). All participants reported race as white except one who identified as white and Asian. Four reported ethnicity as Hispanic or Latino. At baseline, testosterone (T) levels were normal except for four 201 participants that had T &amp;lt;300 ng/dL. At baseline, 6/11 (55%) had suppressed LH (including the four with low T), and 11/14 (79%) had A4/T ratios &amp;gt;1, indicating a predominantly adrenal origin of T. In Study 201, mean T levels increased in Study 201 from 375.5 ng/dL to 390.5 ng/dL at week 2 (n=9), 485.4 ng/dL at week 4 (n=4) and 420.7 ng/dL at week 6 (n=4). In Study 202, mean T levels were 448.4 ng/dL at baseline and 412.0 ng/dL at week 12. Mean LH levels increased in Study 201 from 0.68 IU/L to 1.59 IU/L at week 2 (n=10), 1.62 IU/L at week 4 (n=5) and 0.85 IU/L at week 6 (n=4) (normal 1.8-8.6IU/L). In Study 202, mean LH levels increased from 0.44 IU/L at baseline to 0.87 IU/L at week 12. Of the four men who had low T and suppressed LH (hypogonadotropic hypogonadism), LH rose in all 4 and normalized in 3/4 (75%). Mean A4/T ratios decreased across both studies. In Study 201, the mean declined from a baseline of 1.28 to 0.59 at week 2 (n=9), 0.87 at week 4 (n=4), and 1.03 at week 6 (n=4). In Study 202, the A4/T ratio decreased from baseline of 2.44 to 0.68 at week 12. All the hypogonadal men experienced reductions in A4/T ratios, and 3/4 (75%) achieved A4/T &amp;lt;1. This post-hoc analysis highlights that gonadal axis disruption is common in men with 21OHD and suggests that modulation of adrenal androgen production via CRFR1 antagonism may improve testicular function. Presentation: Saturday, June 17, 2023

FRI425 BIUXX

Abstract Disclosure: D.M. Macedo: None. J. Casanova: None. A. Guerra: None. R. Francisco: None. Title: Hyperandrogenism due to post-menopausal ovarian stromal hyperplasia. Introduction: Severe and rapidly progressive installation of hyperandrogenism in women after menopause should be carefully investigated. Stromal hyperplasia of the ovaries, which etiology is not completely understood, is a rare and benign cause of excess androgen production. In addition to hyperandrogenism, it is usually associated with hyperinsulinism, obesity, metabolic syndrome and acanthosis nigricans. Clinical case: A 60 year old overweight female patient with a history of type 2 diabetes mellitus, dyslipidemia, polycystic ovarian syndrome and menopause at 53 years old, reported the recent onset of hirsutism on the face and back, alopecia and increased libido. The laboratory evaluation revealed: total testosterone 89.6ng/dL (&amp;lt;35.9), free testosterone 5.13pg/mL (&amp;lt;4.2), sex hormone-binding globulin 22.9nmol/L (18-144), LH 21.3mIU/mL (14-52), FSH 27.4mIU/mL (26-135) and E2 &amp;lt;19.0pg/mL. The remaining androgens did not show alterations and hypercortisolism was excluded. Abdominal and pelvic ultrasound was performed and revealed enlarged ovaries with homogeneous echotexture and reduced echogenicity. A pelvic MRI was also performed and showed solid lesions in both ovaries, with moderate T1 and T2 hypointensity, suspicious of bilateral androgen-producing ovarian tumors. After multidisciplinary discussion, the patient underwent total hysterectomy with bilateral oophorectomy. Histopathology was compatible with bilateral stromal hyperplasia of the ovaries. Three months after surgery, the patient presented with a reduction of the facial and back hirsutism and slight improvement in alopecia. The laboratorial evaluation revealed a marked reduction in total testosterone (18.0ng/dL) and free testosterone (1.07pg/mL). Conclusions: New-onset hyperandrogenism in postmenopausal women is very uncommon and a diagnostic challenge. After excluding iatrogenic causes, hypercortisolism or adrenal tumor, the differential diagnosis between virilizing ovarian tumor and bilateral stromal hyperplasia is difficult to establish. Although the imaging distinction is possible, the definitive diagnosis is histopathologic and total hysterectomy with bilateral oophorectomy should be performed. In this case, although MRI was suspicious of bilateral ovarian tumors, the definitive diagnosis was stromal hyperplasia of the ovaries. Three months after surgery, the patient presented marked clinical and biochemical improvement. Presentation Date: Friday, June 16, 2023