Enzyme 11β-hydroxysteroid Dehydrogenase Research Articles

Inappropriate Diet Exacerbates Metabolic Dysfunction-Associated Steatotic Liver Disease via Abdominal Obesity.

Metabolic dysfunction-associated steatotic liver disease (MASLD) represents a refined categorization of non-alcoholic fatty liver disease (NAFLD), highlighting the intricate relationship between hepatic steatosis and metabolic dysfunction. Abdominal obesity (AO), a key diagnostic criterion for metabolic dysfunction, predominantly results from inappropriate diet and unhealthy dietary habits. To comprehensively investigate which dietary factors contribute to MASLD through AO and to understand the underlying biological mechanisms, we initially conducted a systematic review of meta-analysis articles in the PubMed database from the past decade, summarizing dietary factors that affect AO. Subsequently, we conducted targeted searches in the PubMed database for these dietary factors and provided a narrative review of the mechanisms of how these dietary factors lead to AO and how AO exacerbates MASLD. A diet characterized by excessive intake of energy, carbohydrates, fructose, or ultra-processed foods (UPFs) is considered inappropriate. Inappropriate diet leads to the formation of MASLD and AO by enhancing pathways such as de novo lipid synthesis (DNL) in the liver, insulin resistance (IR), gut-liver dysfunction, and inflammation. Dietary interventions for inappropriate diets can effectively intervene in and improve MASLD and AO. The mechanism of inappropriate diet on abdominal fat deposition is through excessive energy or the activation of the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD-1) to increase endocortisol secretion. Then, the excessive accumulation of visceral fat facilitates a rapid and augmented flux of free fatty acids (FFAs) to the liver and initiates a series of deleterious effects, including oxidative stress (OS), endoplasmic reticulum stress (ERS), activation of protein kinase C (PKC) pathways, and inflammation. Additionally, FFAs may mediate excessive lipid deposition and hepatocellular damage through the action of hormones. These pathways to liver damage exacerbate MASLD and progression to metabolic dysfunction-associated steatohepatitis (MASH) and fibrosis. Furthermore, investigating other potential mechanisms by which AO may influence MASLD could offer new recommendations for the treatment guidelines of MASLD.

Unlocking Testosterone Production by Biotransformation: Engineering a Fungal Model of Aspergillus nidulans Strain Deficient in Steroid 11α-Hydroxylase Activity and Expressing 17β-Hydroxysteroid Dehydrogenase Enzyme as Proof of Concept.

Testosterone holds significant medical and economic importance, with the global market for testosterone replacement therapies valued at approximately USD 1.9 billion in 2023. This hormone is essential for the development and maintenance of male sexual characteristics as well as bone and muscle health. It plays a key role in conditions such as hypogonadism, muscle disorders, and andropause. However, the industrial production of testosterone often involves complex chemical processes that result in low yields, high costs, and environmental damage. Microbial biotransformation of steroids presents an eco-friendly alternative to traditional chemical synthesis. A knockout strain of Aspergillus nidulans deficient in steroid 11α-hydroxylase activity was developed, rendering it incapable of hydroxylating androstenedione, progesterone, and testosterone. In these strains, two newly identified CYP450 enzymes, CYP68L1 from A. nidulans and CYP68L8 from Aspergillus ochraceus, were expressed to confirm their roles as steroid 11α-hydroxylases of androstenedione, progesterone, and testosterone. The availability of these 11α-hydroxylases represents significant progress toward achieving efficient single-step steroid fermentation. Furthermore, the A. nidulans knockout strain serves as an effective model for studying the conversion of androstenedione to testosterone upon the expression of the enzyme 17β-hydroxysteroid dehydrogenase, due to its inability to hydroxylate testosterone.

HSD3B1, prostate cancer mortality and modifiable outcomes.

Androgen receptor stimulation by testosterone and dihydrotestosterone is crucial for prostate cancer progression. Despite the initial effectiveness of androgen deprivation therapy (ADT), castration-resistant prostate cancer eventually develops in most men. A common germline missense-encoding polymorphism in HSD3B1 increases extra-gonadal androgen biosynthesis from adrenal precursors owing to increased availability of the encoded enzyme 3β-hydroxysteroid dehydrogenase 1 (3βHSD1) - hence, it is called the adrenal-permissive enzyme. This mechanism explains the more rapid progression to castration-resistant prostate cancer in men who inherit this allele than in men without it via sustained androgen receptor activation despite ADT. Multiple clinical studies, including data derived from prospective phase III studies, have linked adrenal-permissive allele inheritance to inferior clinical responses to ADT and increased mortality, but reversal is possible with upfront adrenal androgen blockade. The adrenal-permissive allele exhibits divergent frequencies across various groups worldwide, which could contribute to differences in clinical outcomes among these populations. Large-scale data from the Million Veteran Program have shown homozygous HSD3B1 adrenal-permissive allele inheritance to be an independent biomarker of prostate cancer-specific mortality. Together, these observations support the integration of HSD3B1 into germline testing and clinical trials as it might help to identify groups at increased likelihood of benefiting from early, intensified, AR-targeting interventions. Lastly, 3βHSD1 is a promising target for pharmacological inhibition, which enables new strategies for systemic prostate cancer therapy.

Glucocorticoid excess alters metabolic rate and substrate utilisation via 11β-HSD1.

Systemic glucocorticoid excess causes several adverse metabolic conditions, most notably Cushing's syndrome. These effects are amplified by the intracellular enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). Here, we determined the less well-characterised effects of glucocorticoid excess, and the contribution of 11β-HSD1 amplification on metabolic rate in mice. Male and female C57BL/6J (wild type, WT) and 11β-HSD1 knockout (11β-HSD1 KO) mice were treated with high-dose corticosterone or a vehicle control for 3 weeks. Indirect calorimetry was conducted during the final week of treatment, with or without fasting, to determine the impact on metabolic rate. We found that corticosterone treatment elevated metabolic rate and promoted carbohydrate utilisation primarily in female WT mice, with effects more pronounced during the light phase. Corticosterone treatment also resulted in greater fat accumulation in female WT mice. Corticosterone induced hyperphagia was identified as a likely causal factor altering the respiratory exchange ratio (RER) but not energy expenditure (EE). Male and female 11β-HSD1 KO mice were protected against these effects. We identify novel metabolic consequences of sustained glucocorticoid excess, identify a key mechanism of hyperphagia, and demonstrate that 11β-HSD1 is required to manifest the full metabolic derangement.

Cortisone in saliva of pigs: validation of a new assay and changes after thermal stress

BackgroundCortisone is derived from cortisol through the action of the enzyme 11β-hydroxysteroid dehydrogenase type II, and it has gained importance in recent years as a biomarker of stress. This study aimed to develop and validate an assay for the measurement of cortisone in pig saliva and evaluate whether its concentration varies in stressful situations. For this purpose, a specific immunoassay was developed and validated analytically, and a study was performed to evaluate whether cortisone concentrations in saliva can vary under heat stress conditions.ResultsThe assay proved to be accurate, reliable, and sensitive for the measurement of cortisone in pig saliva. The limit of detection of the assay was set at 0.006 ng/ml, and the lower limit of quantification was 0.023 ng/ml. It also correlated significantly with the results obtained by LC‒MS/MS (P = 0.003; r = 0.64). In addition, the cortisone concentration in animals subjected to prolonged heat stress decreased significantly 15 days after treatment (P < 0.0001).ConclusionsAccording to these results, cortisone measured by this assay could be used as a tool for the non-invasive evaluation of thermal stress in pig saliva.

Deletion of Hsd11b1 suppresses caloric restriction-induced bone marrow adiposity in male but not female mice.

Bone marrow adipose tissue (BMAT) comprises >10% of total adipose mass in healthy humans. It increases in diverse conditions, including ageing, obesity, osteoporosis, glucocorticoid therapy, and notably, during caloric restriction (CR). BMAT potentially influences skeletal, metabolic, and immune functions, but the mechanisms of BMAT expansion remain poorly understood. Our hypothesis is that, during CR, excessive glucocorticoid activity drives BMAT expansion. The enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) amplifies glucocorticoid activity by catalysing intracellular regeneration of active glucocorticoids from inert 11-keto forms. Mice lacking 11β-HSD1 resist metabolic dysregulation and bone loss during exogenous glucocorticoid excess; thus, we hypothesised that 11β-HSD1 knockout mice would also resist excessive glucocorticoid action during CR, thereby restrining BMAT expansion and bone loss. To test this, we first confirmed that 11β-HSD1 is expressed in mouse and human bone marrow. We then investigated the effects of CR in male and female control and 11β-HSD1 knockout mice from 9 to 15 weeks of age. CR increased Hsd11b1 mRNA in adipose tissue and bone marrow. Deletion of Hsd11b1 did not alter bone or BMAT characteristics in mice fed a control diet and had little effect on tibial bone microarchitecture during CR. Notably, Hsd11b1 deletion attenuated the CR-induced increases in BMAT and prevented increases in bone marrow corticosterone in males but not females. This was not associated with suppression of glucocorticoid target genes in bone marrow. Instead, knockout males had increased progesterone in plasma and bone marrow. Together, our findings show that knockout of 11β-HSD1 prevents CR-induced BMAT expansion in a sex-specific manner and highlights progesterone as a potential new regulator of bone marrow adiposity.

Potent inhibition of human and rat 17β-hydroxysteroid dehydrogenase 1 by curcuminoids and the metabolites: 3D QSAR and in silico docking analysis

ABSTRACT Curcumin, an extensively utilized natural pigment in the food industry, has attracted considerable attention due to its potential therapeutic effects, such as anti-tumorigenic and anti-inflammatory activities. The enzyme 17β-Hydroxysteroid dehydrogenase 1 (17β-HSD1) holds a crucial position in oestradiol production and exhibits significant involvement in oestrogen-responsive breast cancers and endometriosis. This study investigated the inhibitory effects of curcuminoids, metabolites, and analogues on 17β-HSD1, a key enzyme in oestradiol synthesis. Screening 10 compounds, including demethoxycurcumin (IC50, 3.97 μM) and dihydrocurcumin (IC50, 5.84 μM), against human and rat 17β-HSD1 revealed varying inhibitory potencies. These compounds suppressed oestradiol secretion in human BeWo cells at ≥ 5–10 μM. 3D-Quantitative structure-activity relationship (3D-QSAR) and molecular docking analyses elucidated the interaction mechanisms. Docking studies and Gromacs simulations suggested competitive or mixed binding to the steroid or NADPH/steroid binding sites of 17β-HSD1. Predictive 3D-QSAR models highlighted the importance of hydrophobic regions and hydrogen bonding in inhibiting 17β-HSD1 activity. In conclusion, this study provides valuable insights into the inhibitory effects and mode of action of curcuminoids, metabolites, and analogues on 17β-HSD1, which may have implications in the field of hormone-related disorders.

Inhibition of the glucocorticoid-activating enzyme 11β-hydroxysteroid dehydrogenase type 1 drives concurrent 11-oxygenated androgen excess.

Aldo-keto reductase 1C3 (AKR1C3) is a key enzyme in the activation of both classic and 11-oxygenated androgens. In adipose tissue, AKR1C3 is co-expressed with 11β-hydroxysteroid dehydrogenase type 1 (HSD11B1), which catalyzes not only the local activation of glucocorticoids but also the inactivation of 11-oxygenated androgens, and thus has the potential to counteract AKR1C3. Using a combination of invitro assays and in silico modeling we show that HSD11B1 attenuates the biosynthesis of the potent 11-oxygenated androgen, 11-ketotestosterone (11KT), by AKR1C3. Employing exvivo incubations of human female adipose tissue samples we show that inhibition of HSD11B1 results in the increased peripheral biosynthesis of 11KT. Moreover, circulating 11KT increased 2-3 fold in individuals with type 2 diabetes after receiving the selective oral HSD11B1 inhibitor AZD4017 for 35 days, thus confirming that HSD11B1 inhibition results in systemic increases in 11KT concentrations. Our findings show that HSD11B1 protects against excess 11KT production by adipose tissue, a finding of particular significance when considering the evidence for adverse metabolic effects of androgens in women. Therefore, when targeting glucocorticoid activation by HSD11B1 inhibitor treatment in women, the consequently increased generation of 11KT may offset beneficial effects of decreased glucocorticoid activation.

Oncological aspects of ovarian endometriosis

Ovarian cancer is the second most common cause of death from gynaecological malignancies in the world, and in Ukraine it is the most serious type of gynecological cancer. Ovarian endometriomas significantly increase the risk of ovarian cancer, but their malignant transformation occurs in approximately 1% of cases.The article presents a literature review based on the scientific databases PubMed and Scopus for 2013–2023 on the incidence and frequency of ovarian malignant tumors on the background of ovarian endometriosis, carcinogenic mutations, immunological and hormonal disorders in ovarian endometriosis, which can cause its progression to ovarian cancer.Based on the analyzed scientific data, the connection between ovarian endometriosis and ovarian cancer is presented and all possible pathogenetic pathways through which ovarian endometriosis can lead to the formation of ovarian cancer are determined.According to the scientific literature, ovarian endometriosis can indeed lead to the formation of endometrioid and clear cell carcinomas, as well as other subtypes of malignant ovarian tumors. The risk of malignant changes in patients with ovarian endometriomas increases with age, the highest risk is observed in patients over 50 years of age. Despite this, some researchers believe that there are no time limits in the occurrence of malignant transformation of endometrioid ovarian cysts.Today, it is believed that atypical ovarian endometriosis, which is characterized by cytological atypia and architectural proliferation, is a precursor to ovarian cancer, and this condition that has the greatest risk for malignant process development is observed. Ovarian endometriomas contain a huge amount of heme and free iron, which leads to the appearance of an excess of free iron, and as a result, redox disorders occur, which cause carcinogenic mutations and destruction of cellular structures.Mutations in such genes as ARID1A, PIK3CA, AKT1, ERBB2 and PIK3R1, CTNNB1, KRAS, BRAF, PPP2R1A and occasionally in TP53 gene are involved in the occurrence of malignant changes in ovarian endometriomas. The same mutations are found in endometrioid foci of the ovaries and in endometrioid and clear cell carcinomas, which confirms the cancer development due to endometriosis. Disorders in the immune system in endometrioid lesions of the ovaries play a significant role in possible malignant transformation. The production of tumor necrosis factor, interleukin-1β, interleukin-6 increases, the function of natural killers decreases, and immunosuppression increases.Ovarian endometrioid cysts overexpress estradiol because they have increased amounts of the enzyme aromatase and lack the enzyme 17β-hydroxysteroid dehydrogenase type II, which is required to convert estradiol to estrone. Such changes lead to increased proliferative processes, which can also lead to the activation of oncogenic mutations.Thus, ovarian endometriosis significantly increases the risk of developing ovarian cancer, especially endometrioid and clear cell carcinomas. The mechanism of malignant transformation occurs precisely with the appearance of atypical endometriosis of the ovaries. The main pathogenetic pathways through which a malignant process can develop in ovarian endometriomas include: redox imbalance, which triggers a whole spectrum of oncogenic mutations, as well as immune disorders and exposure to high levels of estrogens. However, if patients with ovarian endometriomas are properly managed, the likelihood of ovarian cancer development is low.

P-08 Ectopic Cushing Syndrome in a patient with small cell lung cancer

Abstract Introduction Ectopic Cushing's syndrome (ECS) is a rare endocrine disorder caused by the autonomous secretion of ACTH from a tumor apart from the hypophysis. ECS is the second most common paraneoplastic syndrome associated with small cell lung cancer (SCLC). Due to the aggressive nature of the syndrome, ECS is difficult to diagnose. Most patients present electrolyte disturbances and muscle weakness rather than the typical clinical features of CS. Clinical Case A 70-year-old female patient hospitalized due to pleural effusion was referred for worsening hyperglycemia. She had no previous history of diabetes mellitus. She had a history of 90 pack-years of smoking and chronic obstructive pulmonary disease. The laboratory results were: fasting plasma glucose 216 mg/dl; creatinine, 0.81 mg/dl; sodium, 138 mmol/l; potassium, 3.1 mmol/l; HbA1c, 8.3%; Arterial blood gas analysis revealed metabolic alkalosis (pH 7.63, HCO3 53.6 meq/L). Due to resistant hypokalemia, the patient was evaluated for CS. The patient's ACTH level was 415.5 pg/ml and cortisol level was 63.44 μg/dl. Table 1 shows the tests perfomed for the diagnosis of CS. Pituitary MRG revealed no pathology. IPSS could not be performed as it is not available at our center. A suspicious mass in the right lung was observed on the thorax CT. The pathology of the cytology sample from the pleural effusion revealed SCLC. A chemotherapy regimen containing cisplatin was initiated. Intravenous potassium and acetazolamide treatment were administered for resistant hypokalemia. However, the patient died one month after diagnosis due to type 2 respiratory failure. Conclusion Ectopic secretion of ACTH accounts for 10–15% of CS. ECS is usually caused by mainly SCLC (45%), thymic carcinoma (15%), bronchial carcinoid (10%) and pancreas islet-cell carcinoma (10%). ECS is reported to occur in 1.6%–4.5% of patients with SCLC. IPSS is the most reliable examination for ECS. For ECS, surgery remains the optimal treatment. Some reports showed that metyrapone, ketoconazole and octreotide are effective but not widely used. In our case, the presence of resistant hypokalemia, metabolic alkalosis, and worsening hyperglycemia with the lack of classic clinical features of CS led us to suspect ECS. ECS has a higher incidence of hypokalemic alkalosis than other causes of Cushing's syndrome (&amp;gt;90% vs. 10%). Excessive amounts of adrenal steroids with the inhibition of the enzyme 11β-hydroxysteroid dehydrogenase may cause this pathology. This enzyme normally converts cortisol to cortisone avoiding the mineralocorticoid action of cortisol. In conclusion, although rare, in cases where conditions such as metabolic alkalosis, resistant hypokalemia, with the absence of classic clinical features of CS, the possibility of ectopic CS should be kept in mind.Table 1.Laboratory results of the patient

Halogenated bisphenol A derivatives potently inhibit human and rat 11β-hydroxysteroid dehydrogenase 1: Structure-activity relationship and molecular docking.

Chlorinated bisphenol A (BPA) derivatives are formed during chlorination process of drinking water, whereas bisphenol S (BPS) and brominated BPA and BPS (TBBPA and TBBPS) were synthesized for many industrial uses such as fire retardants. However, the effect of halogenated BPA and BPS derivatives on glucocorticoid metabolizing enzyme 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) remains unclear. The inhibitory effects of 6 BPA derivatives in the inhibition of human and rat 11β-HSD1 were investigated. The potencies for inhibition on human 11β-HSD1 were TBBPA (IC50, 3.87 μM) = monochloro BPA (MCBPA, 4.08 μM) = trichloro BPA (TrCBPA, 4.41 μM) > tetrachloro BPA (TCBPA, 9.75 μM) > TBBPS (>100 μM) = BPS (>100 μM), and those for rat 11β-HSD1 were TrCBPA (IC50, 2.76 μM) = MCBPA (3.75 μM) > TBBPA (39.58 μM) > TCBPA = TBBPS = BPS. All these BPA derivatives are mixed/competitive inhibitors of both human and rat enzymes. Molecular docking studies predict that MCBPA, TrCBPA, TCBPA, and TBBPA all bind to the active site of human 11β-HSD1, forming hydrogen bonds with catalytic residue Ser170 except TCBPA. Regression of the lowest binding energy with IC50 values revealed a significant inverse linear regression. In conclusion, halogenated BPA derivatives are mostly potent inhibitors of human and rat 11β-HSD1, and there is structure-dependent inhibition.

Metabolic Dysfunction in Idiopathic Intracranial Hypertension: Current Theories and Updates.

Idiopathic intracranial hypertension (IIH) is a disease characterized by increased intracranial pressure (ICP) without identifiable secondary causes. While the increased ICP is a critical diagnostic feature, the underlying pathophysiological mechanisms remain unclear. Previous theories have suggested cerebrospinal fluid (CSF) overproduction, impaired reabsorption, or circulatory obstruction as potential causes. Emerging evidence indicates that IIH may not be solely a central nervous system disorder but also a systemic metabolic disorder. Metabolic and hormonal dysregulation features, including hyperleptinemia, adipocyte leptin hypersecretion, increased insulin resistance, and androgen excess, have been noted in IIH. Furthermore, the targeted blockade of the cortisol-producing enzyme 11β-hydroxysteroid dehydrogenase type 1 has demonstrated therapeutic potential in treating IIH. Consequently, the role of metabolic dysfunction and hormonal imbalance in IIH warrants consideration. This review aims to provide a comprehensive update on current theories regarding the mechanisms of metabolic dysfunction in IIH.

Visceral fat sympathectomy ameliorates systemic and local stress response related to chronic sleep restriction.

Disturbance of sleep homeostasis encompasses health issues, including metabolic disorders like obesity, diabetes, and augmented stress vulnerability. Sleep and stress interact bidirectionally to influence the central nervous system and metabolism. Murine models demonstrate that decreased sleep time is associated with an increased systemic stress response, characterized by endocrinal imbalance, including the elevated activity of hypothalamic-pituitary-adrenal axis, augmented insulin, and reduced adiponectin, affecting peripheral organs physiology, mainly the white adipose tissue (WAT). Within peripheral organs, a local stress response can also be activated by promoting the formation of corticosterone. This local amplifying glucocorticoid signaling is favored through the activation of the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). In WAT, 11β-HSD1 activity is upregulated by the sympathetic nervous system, suggesting a link between sleep loss, augmented stress response, and a potential WAT metabolic disturbance. To gain more understanding about this relationship, metabolic and stress responses of WAT-sympathectomized rats were analyzed to identify the contribution of the autonomic nervous system to stress response-related metabolic disorders during chronic sleep restriction. Male Wistar rats under sleep restriction were allowed just 6 h of daily sleep over eight weeks. Results showed that rats under sleep restriction presented higher serum corticosterone, increased adipose tissue 11β-HSD1 activity, weight loss, decreased visceral fat, augmented adiponectin, lower leptin levels, glucose tolerance impairment, and mildly decreased daily body temperature. In contrast, sympathectomized rats under sleep restriction exhibited decreased stress response (lower serum corticosterone and 11β-HSD1 activity). In addition, they maintained weight loss, explained by a reduced visceral fat pad, leptin, and adiponectin, improved glucose management, and persisting decline in body temperature. These results suggest autonomic nervous system is partially responsible for the WAT-exacerbated stress response and its metabolic and physiological disturbances.

Rutin attenuated hexachlorobenzene-induced testicular injury via regulation of oxidative stress, steroidogenic enzymes and apoptotic process in male rats

Hexachlorobenzene (HCB) is a pesticide that is constantly produced as a by-product in many industrial processes. Rutin (RUT) is a flavonoid found in many plants with numerous biological and pharmacological properties. However, there is no scientifical information on the impact of RUT on HCB-induced reproductive toxicity in male rats. Thus, we investigated the effect of rutin on HCB-induced reproductive toxicity. Rats were divided into four groups consisting of control administered with corn oil, HCB alone (16 mg/kg), RUT alone (100 mg/kg) and HCB + RUT that were treated orally by gavage for 28 days. Hexachlorobenzene caused a significant reduction in epididymal sperm count, sperm progressive motility, viability, number of hypo-osmotic tail swelled spermatozoa and increased sperm morphological abnormalities (head, midpiece and tail). Furthermore, HCB decreased the activities of gamma glutamyl transferase, lactate dehydrogenase, sorbitol dehydrogenase, superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, levels of glutathione and total protein with increased malondialdehyde level. Additionally, HCB increased the levels of total cholesterol, low density lipoprotein, triglycerides, follicle stimulating hormone, luteinizing hormone but decreased high density lipoproteins, serum testosterone, steroidogenic enzymes (3β-hydroxysteroid dehydrogenase and 17β-hydroxysteroid dehydrogenase), B-cell lymphoma and increased pro-apoptotic markers (Bcl-2 associated protein (Bax), caspases-3 and-9) alongside histological and histomorphometrical abnormalities. Nevertheless, RUT prevented all the aberrations caused by HCB. Our result showed that HCB induced reproductive toxicity in rats and RUT protected against HCB-induced r reproductive toxicity.

The glucocorticoid-activating enzyme 11β-hydroxysteroid dehydrogenase type 1 catalyzes the activation of testosterone

Testosterone biosynthesis from its precursor androstenedione is thought to be exclusively catalysed by the 17β-hydroxysteroid dehydrogenases—HSD17B3 in testes, and AKR1C3 in the ovary, adrenal and peripheral tissues. Here we show for the first time that the glucocorticoid activating enzyme 11β-hydroxysteroid dehydrogenase type 1 (HSD11B1) can also catalyse the 17β-reduction of androstenedione to testosterone, using a combination of in vitro enzyme kinetic assays, mathematical modelling, and molecular docking analysis. Furthermore, we show that co-expression of HSD11B1 and AKR1C3 increases testosterone production several-fold compared to the rate observed with AKR1C3 only, and that HSD11B1 is likely to contribute significantly to testosterone production in peripheral tissues.

Nanomolar Benzothiazole-Based Inhibitors of 17β-HSD10 with Cellular Bioactivity.

Multifunctional mitochondrial enzyme 17β-hydroxysteroid dehydrogenase type 10 (17β-HSD10) is a potential drug target for the treatment of various pathologies. The most discussed is the pathology associated with Alzheimer's disease (AD), where 17β-HSD10 overexpression and its interaction with amyloid-β peptide contribute to mitochondrial dysfunction and neuronal stress. In this work, a series of new benzothiazole-derived 17β-HSD10 inhibitors were designed based on the structure-activity relationship analysis of formerly published inhibitors. A set of enzyme-based and cell-based methods were used to evaluate the inhibitory potency of new compounds, their interaction with the enzyme, and their cytotoxicity. Most compounds exhibited significantly a higher inhibitory potential compared to published benzothiazolyl ureas and good target engagement in a cellular environment accompanied by low cytotoxicity. The best hits displayed mixed-type inhibition with half maximal inhibitory concentration (IC50) values in the nanomolar range for the purified enzyme (3-7, 15) and/or low micromolar IC50 values in the cell-based assay (6, 13-16).

SAT276 Glucocorticoid Excess Elevates Metabolic Rate Via A 11b-HSD1 Dependent Mechanism In C57BL/6J Mice

Abstract Disclosure: S. Heaselgrave: None. S. Heising: None. S.A. Morgan: None. A. Kabli: None. M. Sagmeister: None. C. Doig: None. R.S. Hardy: None. N.M. Morton: None. K. Tsintzas: None. G.G. Lavery: None. Introduction: Glucocorticoids are vital metabolic regulators. However, glucocorticoid excess (GE) causes severe metabolic dysfunction, ultimately leading to Cushing’s Syndrome. This dysfunction is often dependent on the presence of the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). Whether GE also alters metabolic rate, and whether this is also dependent on 11β-HSD1, remains unclear and merits investigation in a mouse model of GE and 11β-HSD1 inhibition. Methods: Male and female wild type (WT) C57BL/6J and 11β-HSD1KO mice were treated with corticosterone or vehicle control ad libitum via drinking water for 3 weeks whilst being fed a standard chow diet (n=12). Mice were housed in a TSE Phenomaster system for the final week of treatment for comprehensive metabolic assessment. Results: Corticosterone treatment in WT mice resulted in a phenotype typical of GE, however female mice experienced greater fat accumulation and bodyweight gain compared to males. 11β-HSD1KO mice did not experience this phenotype. Corticosterone treatment elevated energy expenditure (EE) in female WT mice during the day alone (25±5.9%). Male WT mice did not experience a significant increase. Corticosterone did significantly elevate the respiratory exchange ratio (RER) towards 1 in both male (10.7±5.7%) and female (11.8±7.0%) WT mice during the day. RER remained elevated in female WT mice (7.6±4.8%) and moderately so in male WT mice (3.2±2.6%) at night, indicating a continued suppression of lipid oxidation in favour of carbohydrate oxidation. Activity, assessed in female WT mice only, was continually decreased by corticosterone treatment (-53.1±54.8%). Corticosterone treated WT mice became hyperphagic and polydipsic, continuously eating and drinking more than controls. Male and female 11β-HSD1KO mice treated with corticosterone did not experience these effects. Conclusion: These findings provide further insights into the metabolic consequences of GE as well as the dependency on 11β-HSD1 to facilitate them. Whilst elevations are seen in both male and female WT mice, they are greater in females. Activity is decreased meaning it is not responsible for the elevations. However, hyperphagia and the subsequent chemical energy cost of de novo lipogenesis might be responsible and merits further investigation. Presentation: Saturday, June 17, 2023

Physiologically relevant fluorescent assay for identification of 17β-hydroxysteroid dehydrogenase type 10 inhibitors.

Mitochondrial enzyme 17β-hydroxysteroid dehydrogenase type 10 (HSD10) is a potential molecular target for treatment of mitochondrial-related disorders such as Alzheimer's disease (AD). Its over-expression in AD brains is one of the critical factors disturbing the homeostasis of neuroprotective steroids and exacerbating amyloid beta (Aβ)-mediated mitochondrial toxicity and neuronal stress. This study was focused on revalidation of the most potent HSD10 inhibitors derived from benzothiazolyl urea scaffold using fluorescent-based enzymatic assay with physiologically relevant substrates of 17β-oestradiol and allopregnanolone. The oestradiol-based assay led to the identification of two nanomolar inhibitors (IC50 70 and 346 nM) differing from HSD10 hits revealed from the formerly used assay. Both identified inhibitors were found to be effective also in allopregnanolone-based assay with non-competitive or uncompetitive mode of action. In addition, both inhibitors were confirmed to penetrate the HEK293 cells and they were able to inhibit the HSD10 enzyme in the cellular environment. Both molecules seem to be potential lead structures for further research and development of HDS10 inhibitors.

Bisphenol A Analogues Inhibit Human and Rat 11β-Hydroxysteroid Dehydrogenase 1 Depending on Its Lipophilicity.

Bisphenol A (BPA) analogues substituted on the benzene ring are widely used in a variety of industrial and consumer materials. However, their effects on the glucocorticoid-metabolizing enzyme 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) remain unclear. The inhibitory effects of 6 BPA analogues on the inhibition of human and rat 11β-HSD1 were investigated. The potencies of inhibition on human 11β-HSD1 were bisphenol H (IC50, 0.75 µM) > bisphenol G (IC50, 5.06 µM) > diallyl bisphenol A (IC50, 13.36 µM) > dimethyl bisphenol A (IC50, 30.18 µM) > bisphenol A dimethyl ether (IC50, 33.08 µM) > tetramethyl bisphenol A (>100 µM). The inhibitory strength of these chemicals on rat 11β-HSD1 was much weaker than that on the human enzyme, ranging from 74.22 to 205.7 µM. All BPA analogues are mixed/competitive inhibitors of both human and rat enzymes. Molecular docking studies predict that bisphenol H and bisphenol G both bind to the active site of human 11β-HSD1, forming a hydrogen bond with catalytic residue Ser170. The bivariate correlation of IC50 values with LogP (lipophilicity), molecular weight, heavy atoms, and molecular volume revealed a significant inverse regression and the correlation of IC50 values with ΔG (low binding energy) revealed a positive regression. In conclusion, the lipophilicity, molecular weight, heavy atoms, molecular volume, and binding affinity of a BPA analogue determine the inhibitory strength of human and rat 11β-HSD isoforms.

3βHSD activity saturates at physiological substrate concentrations in intact cells.

Conversion of adrenally produced dehydroepiandrosterone (DHEA) to the potent androgen dihydrotestosterone (DHT) is an important mechanism by which prostate cancer reaches castration resistance. At the start of this pathway is a branch point at which DHEA can be converted to Δ4 -androstenedione by the enzyme 3β-hydroxysteroid dehydrogenase (3βHSD) or to Δ5 -androstenediol by 17βHSD. To better understand this process, we studied the kinetics of these reactions in cells. Prostate cancer cells (LNCaP cell line) were incubated with steroids (DHEA and Δ5 -androstenediol) over a range of concentrations and the steroid metabolism reaction products were measured by mass spectrometry or by high-performance liquid chromatography to determine reaction kinetics. To confirm the generalizability of results, experiments were also performed in JEG-3 placental choriocarcinoma cells. The two reactions displayed very different saturation profiles, with only the 3βHSD-catalyzed reaction beginning to saturate within a physiological substrate concentration range. Strikingly, incubating LNCaP cells with low (in the ~10 nM range) concentrations of DHEA resulted in a large majority of the DHEA undergoing 3βHSD-catalyzed conversion to Δ4 -androstenedione, whereas high concentrations of DHEA (in the 100s of nM range) resulted in most of the DHEA undergoing 17βHSD-catalyzed conversion to Δ5 -androstenediol. Contrary to expectations from previous studies that used purified enzyme, cellular metabolism of DHEA by 3βHSD begins to saturate in the physiological concentration range, suggesting that fluctuations in DHEA concentrations could be buffered at the downstream active androgen level.