Inhibition Of 11β-HSD1 Activity Research Articles

Novel 2-(Adamantan-1-ylamino)Thiazol-4(5H)-One Derivatives and Their Inhibitory Activity towards 11β-HSD1-Synthesis, Molecular Docking and In Vitro Studies.

A common mechanism in which glucocorticoids participate is suggested in the pathogenesis of such metabolic diseases as obesity, metabolic syndrome, or Cushing’s syndrome. The enzyme involved in the control of the availability of cortisol, the active form of the glucocorticoid for the glucocorticoid receptor, is 11β-HSD1. Inhibition of 11β-HSD1 activity may bring beneficial results for the alleviation of the course of metabolic diseases such as metabolic syndrome, Cushing’s syndrome or type 2 diabetes. In this work, we obtained 10 novel 2-(adamantan-1-ylamino)thiazol-4(5H)-one derivatives containing different substituents at C-5 of thiazole ring and tested their activity towards inhibition of two 11β-HSD isoforms. For most of them, over 50% inhibition of 11β-HSD1 and less than 45% inhibition of 11β-HSD2 activity at the concentration of 10 µM was observed. The binding energies found during docking simulations for 11β-HSD1 correctly reproduced the experimental IC50 values for analyzed compounds. The most active compound 2-(adamantan-1-ylamino)-1-thia-3-azaspiro[4.5]dec-2-en-4-one (3i) inhibits the activity of isoform 1 by 82.82%. This value is comparable to the known inhibitor-carbenoxolone. The IC50 value is twice the value determined by us for carbenoxolone, however inhibition of the enzyme isoform 2 to a lesser extent makes it an excellent material for further tests.

A novel highly potent and selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor, INU-101

11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) is a cortisol regenerating enzyme that amplifies tissue glucocorticoid levels, especially in the liver and adipose tissue. Knockout mice or a selective inhibitor of 11β-HSD1 improves metabolic syndrome parameters in preclinical models and human clinical trials. Here, we evaluated the therapeutic potential of INU-101, a potent and selective oral inhibitor of 11β-HSD1. The in vitro activity of 11β-HSD1 was measured using the homogeneous time-resolved fluorescence (HTRF) assay. Differentiated adipocytes were used to evaluate the cellular 11β-HSD1 activity. To determine the inhibitory effects on 11β-HSD1 in tissues, we performed ex vivo studies using liver and adipose tissue isolated from C57BL/6 J mice and Cynomolgus monkeys. KKAy mice, ob/ob mice and ZDF rats were administered INU-101 to evaluate whether this compound ameliorated metabolic abnormalities in obese and diabetic animals. INU-101 had highly potent inhibitory activity in mouse, monkey and human 11β-HSD1, derived from liver microsomes. The oral administration of INU-101 significantly inhibited 11β-HSD1 activity in the liver and adipose tissue of mice and monkeys. In KKAy mice, ob/ob mice and ZDF rats, the oral administration of INU-101 enhanced insulin sensitivity and lowered the fasting blood glucose level. Furthermore, INU-101 treatment decreased the body weight and ameliorated an improved lipid profile in the diabetic mouse model. These results suggest that the 11β-HSD1 inhibitor, INU-101 may serve as a novel drug candidate for the treatment of type 2 diabetes and metabolic syndrome.

Discovery of BI 135585, an in vivo efficacious oxazinanone-based 11β hydroxysteroid dehydrogenase type 1 inhibitor

A potent, in vivo efficacious 11β hydroxysteroid dehydrogenase type 1 (11β HSD1) inhibitor (11j) has been identified. Compound 11j inhibited 11β HSD1 activity in human adipocytes with an IC50 of 4.3nM and in primary human adipose tissue with an IC80 of 53nM. Oral administration of 11j to cynomolgus monkey inhibited 11β HSD1 activity in adipose tissue. Compound 11j exhibited >1000× selectivity over other hydroxysteroid dehydrogenases, displays desirable pharmacodynamic properties and entered human clinical trials in 2011.

Novel 11β-hydroxysteroid dehydrogenase 1 inhibitors reduce cortisol levels in keratinocytes and improve dermal collagen content in human ex vivo skin after exposure to cortisone and UV.

Activity and selectivity assessment of new bi-aryl amide 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) inhibitors, prepared in a modular manner via Suzuki cross-coupling, are described. Several compounds inhibiting 11β-HSD1 at nanomolar concentrations were identified. Compounds 2b, 3e, 7b and 12e were shown to selectively inhibit 11β-HSD1 over 11β-HSD2, 17β-HSD1 and 17β-HSD2. These inhibitors also potently inhibited 11β-HSD1 activity in intact HEK-293 cells expressing the recombinant enzyme and in intact primary human keratinocytes expressing endogenous 11β-HSD1. Moreover, compounds 2b, 3e and 12e were tested for their activity in human skin biopsies. They were able to prevent, at least in part, both the cortisone- and the UV-mediated decreases in collagen content. Thus, inhibition of 11β-HSD1 by these compounds can be further investigated to delay or prevent UV-mediated skin damage and skin aging.

11β-Hydroxysteroid Dehydrogenase Type 1 Inhibition Attenuates the Adverse Effects of Glucocorticoids on Dermal Papilla Cells.

PurposeGlucocorticoids, stress-related hormones, inhibit hair growth. Intracellular glucocorticoid availability is regulated by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). 11β-HSD1 was recently detected in keratinocytes and fibroblasts. However, the expression of 11β-HSD1 in human hair follicles remains unknown. We aimed to examine 11β-HSD1 expression in human dermal papilla cells (DPCs) and to investigate whether modulation of 11β-HSD1 activity can regulate the negative effects of glucocorticoids on DPCs.Materials and Methods11β-HSD1 expression in normal human scalp skin was examined by immunohistochemistry. 11β-HSD1 protein was detected in Western blots of human DPCs. Cultured human DPCs were treated with cortisol with or without a selective 11β-HSD1 inhibitor and subsequently stained for Ki-67 antibody. Expression levels of 11β-HSD1, Wnt5a, alkaline phosphatase (ALP), and vascular endothelial growth factor (VEGF) were analyzed by Western blotting.Results11β-HSD1 was detected in dermal papilla in human scalp skin by immunohistochemistry. Human DPCs expressed 11β-HSD1 protein in vitro. Furthermore, cortisol stimulated the expression of 11β-HSD1 in DPCs. Glucocorticoids decreased cellular proliferation and the expression of Wnt5a, ALP, and VEGF in DPCs. A specific 11β-HSD1 inhibitor significantly attenuated the anti-proliferative effects of cortisol and reversed the cortisol-induced suppression of Wnt5a, ALP, and VEGF expression in DPCs.ConclusionOur data demonstrated the expression of 11β-HSD1 in human DPCs and revealed that inhibition of 11β-HSD1 activity can partially prevent the negative effect of glucocorticoids on DPCs, suggesting the possible application of 11β-HSD1 inhibitors for stress-related hair loss.

Anti-inflammatory effect of a selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor via the stimulation of heme oxygenase-1 in LPS-activated mice and J774.1 murine macrophages

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts inactive cortisone to the active cortisol. 11β-HSD1 may be involved in the resolution of inflammation. In the present study, we investigate the anti-inflammatory effects of 2-(3-benzoyl)-4-hydroxy-1,1-dioxo-2H-1,2-benzothiazine-2-yl-1-phenylethanone (KR-66344), a selective 11β-HSD1 inhibitor, in lipopolysaccharide (LPS)-activated C57BL/6J mice and macrophages. LPS increased 11β-HSD1 activity and expression in macrophages, which was inhibited by KR-66344. In addition, KR-66344 increased survival rate in LPS treated C57BL/6J mice. HO-1 mRNA expression level was increased by KR-66344, and this effect was reversed by the HO competitive inhibitor, ZnPP, in macrophages. Moreover, ZnPP reversed the suppression of ROS formation and cell death induced by KR-66344. ZnPP also suppressed animal survival rate in LPS plus KR-66344 treated C57BL/6J mice. In the spleen of LPS-treated mice, KR-66344 prevented cell death via suppression of inflammation, followed by inhibition of ROS, iNOS and COX-2 expression. Furthermore, LPS increased NFκB-p65 and MAPK phosphorylation, and these effects were abolished by pretreatment with KR-66344. Taken together, KR-66344 protects against LPS-induced animal death and spleen injury by inhibition of inflammation via induction of HO-1 and inhibition of 11β-HSD1 activity. Thus, we concluded that the selective 11β-HSD1 inhibitor may provide a novel strategy in the prevention/treatment of inflammatory disorders in patients.

Yeast-based assays for screening 11β-HSD1 inhibitors.

BackgroundIntracellular metabolism of glucocorticoid hormones plays an important role in the pathogenesis of metabolic syndrome and regulates, among many physiological processes, collagen metabolism in skin. At the peripheral level the concentration of active glucocorticoids is mainly regulated by the 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) enzyme, involved in the conversion of cortisone into the biologically active hormone cortisol. Cortisol interacts with the glucocorticoid receptor and regulates the expression of different classes of genes within the nucleus. Due to its implication in glucocorticoid metabolism, the inhibition of 11β-HSD1 activity has become a dominant strategy for the treatment of metabolic syndrome. Moreover, inhibitors of this target enzyme can be used for development of formulations to counteract skin ageing. Here we present the construction of two yeast cell based assays that can be used for the screening of novel 11β-HSD1 inhibitors.ResultsThe yeast Saccharomyces cerevisiae is used as a host organism for the expression of human 11β-HSD1 as well as a genetically encoded assay system that allows intracellular screening of molecules with 11β-HSD1 inhibitory activity. As proof of concept the correlation between 11β-HSD1 inhibition and fluorescent output signals was successfully tested with increasing concentrations of carbenoxolone and tanshinone IIA, two known 11β-HSD1 inhibitors. The first assay detects a decrease in fluorescence upon 11β-HSD1 inhibition, whereas the second assay relies on stabilization of yEGFP upon inhibition of 11β-HSD1, resulting in a positive read-out and thus minimizing the rate of false positives sometimes associated with read-outs based on loss of signals. Specific inhibition of the ABC transporter Pdr5p improves the sensitivity of the assay strains to cortisone concentrations by up to 60 times.ConclusionsOur yeast assay strains provide a cost-efficient and easy to handle alternative to other currently available assays for the screening of 11β-HSD1 inhibitors. These assays are designed for an initial fast screening of large numbers of compounds and enable the selection of cell permeable molecules with target inhibitory activity, before proceeding to more advanced selection processes. Moreover, they can be employed in yeast synthetic biology platforms to reconstitute heterologous biosynthetic pathways of drug-relevant scaffolds for simultaneous synthesis and screening of 11β-HSD1 inhibitors at intracellular level.

Identification of a novel 11β-HSD1 inhibitor from a high-throughput screen of natural product extracts

Selective inhibitors of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) have considerable potential as a treatment for metabolic syndrome including type 2 diabetes mellitus and obesity. To identify 11β-HSD1 inhibitors, we conducted high-throughput screening (HTS) of active natural product extracts from the Korea Chemical Bank, including Tanshinone I, Tanshinone IIA, and flavanone derivatives, and 2- and 3-phenyl-4H-chromen-4-one. Then Tanshinone IIA and its derivatives were targeted for the development of a lead compound according to the HTS results. However, the mechanism for anti-adipogenic effect through 11β-HSD1 enzyme inhibition by Tanshinone IIA is not clear. Tanshinone IIA (2a) concentration-dependently inhibited 11β-HSD1 activity in human and mouse 11β-HSD1 overexpressed cells and 3T3-L1 adipocytes. Tanshinone IIA (2a) also inhibited 11β-HSD1 enzyme activities in murine liver and fats. Furthermore, Tanshinone IIA (2a)-suppressed adipocyte differentiation of cortisone-induced adipogenesis in 3T3-L1 cells was associated with the suppression of the cortisone-induced adipogenesis-specific markers mRNA and protein expression. In 3T3-L1 preadipocytes, Tanshinone IIA (2a)-inhibited cortisone induced reactive oxygen species formation in a concentration-dependent manner. Thus, these results support the therapeutic potential of Tanshinone IIA (2a) as a 11β-HSD1 inhibitor in metabolic syndrome patients.

A novel highly potent and selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor, UI-1499

AimsElevated levels of glucocorticoid hormones cause glucose intolerance, visceral obesity, insulin resistance, hypertension, and dyslipidemia. 11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) represents as an attractive therapeutic target for treatment of metabolic syndrome and type 2 diabetes. This study investigated whether 11β-HSD1 inhibition by a novel selective inhibitor, (1S,3R,4S,5S,7S)-4-(1-((2-fluoro-N-methylphenylsulfonamido)-methyl) cyclopropanecarboxamido) adamantane-1-carboxamide (UI-1499) ameliorated metabolic abnormalities in diabetic mice. Main methodsThe in vitro activity of 11β-HSD1 was measured using the homogeneous time resolved fluorescence (HTRF) assay. Differentiated adipocytes were used to evaluate cellular 11β-HSD1 activity. To determine the inhibitory effects on 11β-HSD1 in tissues, we performed ex vivo studies using liver and epididymal fat isolated from C57BL/6J mice. KKAy mice were administered with UI-1499 to evaluate whether this compound ameliorated metabolic abnormalities in vivo in diabetic animals. Key findingsUI-1499 had highly potent inhibitory activity in mouse, monkey and human 11β-HSD1, derived from liver microsomes. Oral administration of 45 mg/kg UI-1499 significantly inhibited 11β-HSD1 activity in C57BL/6J mouse liver and epididymal fat tissues. In KKAy mice, oral administration of UI-1499 (10 and 30 mg/kg for 3 weeks) lowered fasting blood glucose and HbA1c levels; these effects were comparable to those of pioglitazone. Further, UI-1499 treatment lowered plasma low-density lipoprotein (LDL) level in KKAy mice. SignificanceThese results suggest that the 11β-HSD1 inhibitor, UI-1499, may serve as a novel drug candidate for the treatment of type 2 diabetes and metabolic syndrome.

Anti-diabetic and anti-inflammatory effect of a novel selective 11β-HSD1 inhibitor in the diet-induced obese mice

It has been reported that the selective inhibitors of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) have considerable potential for treating type 2 diabetes mellitus, metabolic syndrome and inflammation. In the present study, we investigated the anti-diabetic and anti-inflammatory effects of N-(5-carbamoyladamantan-2-yl)-3-((2-fluorophenyl) sulfonyl)thiazolidine-2-carboxamide (KR-67105), a novel 11β-HSD1 inhibitor, in diabetic mice model and preadipocyte model. KR-67105 concentration dependently inhibited 11β-HSD1 activity in human and mouse 11β-HSD1 overexpressing cells and mouse 3T3-L1 adipocytes. Furthermore, KR-67105 concentration-dependently inhibited 11β-HSD1 activity in the ex vivo assay of C57BL/6 mice. In the study with diet-induced obese (DIO) mice, the administration of KR-67105 (100mg/kg/day, orally for 28 days) improved the glucose tolerance and insulin sensitivity as determined by the oral glucose tolerance test and the insulin tolerance test. Anti-diabetic effect by KR-67105 was associated with the suppression of diabetic related genes expression in liver and fat. Furthermore, KR-67105 suppressed 11β-HSD1 activity in liver and fat of diabetic mice, but showed no effect on adrenal grand weight/body weight ratio and plasma corticosterone concentration in diabetic mice. In 3T3-L1 preadipocytes, cortisone induced the mRNA of inflammatory cytokines and 11β-HSD1 and reactive oxygen species formation. This effect was abolished by co-incubation with KR-67105 in a concentration-dependent manner. Moreover, KR-67105 attenuated cortisone induced iNOS expression and phosphorylation of NF-κB p65, p38 MAPK, and ERK1/2 in preadipocytes. Taken together, it is concluded that a selective 11β-HSD1 inhibitor, KR-67105, may provide a new therapeutic window in the prevention and treatment of type 2 diabetes with chronic inflammation without toxicity.

Inhibition of microsomal cortisol production by (–)‐epigallocatechin‐3‐gallate through a redox shift in the endoplasmic reticulum—A potential new target for treating obesity‐related diseases

Conversion of cortisone to cortisol by 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1) in the endoplasmic reticulum (ER) of the target cells is a major determinant of glucocorticoid action, and plays an important role in the development of obesity-related diseases. Inhibition of 11βHSD1 activity is, therefore, considered as a promising novel strategy for the treatment of metabolic syndrome and diabetes. Tea flavanols and their major representative, epigallocatechin gallate are known as antiobesity and antidiabetic agents. Their impacts on blood glucose level, hepatic glucose production, and insulin responsiveness resemble those observed on inhibition or depletion of 11βHSD1. We aimed to study the effect of epigallocatechin gallate on 11βHSD1 activity in ER-derived rat liver microsomes by measuring cortisone and cortisol with HPLC. Cortisol production was efficiently suppressed in a concentration dependent manner in intact microsomal vesicles. However, this effect was abolished by membrane permeabilization; and the three proteins involved in the overall process (11βHSD1, hexose 6-phosphate dehydrogenase, and glucose 6-phosphate transporter) were not or only mildly affected. Further investigation revealed the oxidation of luminal NADPH to NADP⁺, which attenuates cortisone reduction and favors cortisol oxidation in this compartment. Such a redox shift in the ER lumen might contribute to the beneficial health effects of tea flavanols and should be regarded as a promising strategy for the development of novel selective 11βHSD1 inhibitors to treat obesity-related diseases.

Carbenoxolone treatment ameliorated metabolic syndrome in WNIN/Ob obese rats, but induced severe fat loss and glucose intolerance in lean rats.

Background11beta-hydroxysteroid dehydrogenase type 1 (11β-HSD1) regulates local glucocorticoid action in tissues by catalysing conversion of inactive glucocorticoids to active glucocorticoids. 11β-HSD1 inhibition ameliorates obesity and associated co-morbidities. Here, we tested the effect of 11β-HSD inhibitor, carbenoxolone (CBX) on obesity and associated comorbidities in obese rats of WNIN/Ob strain, a new animal model for genetic obesity.Methodology/Principal FindingsSubcutaneous injection of CBX (50 mg/kg body weight) or volume-matched vehicle was given once daily for four weeks to three month-old WNIN/Ob lean and obese rats (n = 6 for each phenotype and for each treatment). Body composition, plasma lipids and hormones were assayed. Hepatic steatosis, adipose tissue morphology, inflammation and fibrosis were also studied. Insulin resistance and glucose intolerance were determined along with tissue glycogen content. Gene expressions were determined in liver and adipose tissue. CBX significantly inhibited 11β-HSD1 activity in liver and adipose tissue of WNIN/Ob lean and obese rats. CBX significantly decreased body fat percentage, hypertriglyceridemia, hypercholesterolemia, insulin resistance in obese rats. CBX ameliorated hepatic steatosis, adipocyte hypertrophy, adipose tissue inflammation and fibrosis in obese rats. Tissue glycogen content was significantly decreased by CBX in liver and adipose tissue of obese rats. Severe fat loss and glucose- intolerance were observed in lean rats after CBX treatment.Conclusions/SignificanceWe conclude that 11β-HSD1 inhibition by CBX decreases obesity and associated co-morbidities in WNIN/Ob obese rats. Our study supports the hypothesis that inhibition of 11β-HSD1 is a key strategy to treat metabolic syndrome. Severe fat loss and glucose -intolerance by CBX treatment in lean rats suggest that chronic 11β-HSD1 inhibition may lead to insulin resistance in normal conditions.

Anti-diabetic and anti-adipogenic effects of a novel selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor in the diet-induced obese mice

Glucocorticoid excess (Cushing's syndrome) causes metabolic syndrome such as visceral obesity, insulin resistance, diabetes mellitus, dyslipidaemia and hypertension. The selective inhibitors of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) have considerable potential for treating type 2 diabetes mellitus and metabolic syndrome. In the present study, we investigated the anti-diabetic and anti-adipogenic effects of 4-(2-(1,1-dioxido-6-(2,4,6-trichlorophenyl)-1,2,6-thiadiazinan-2-yl)acetamido)adamantane-1-carboxamide (KR-67183), a novel selective 11β-HSD1 inhibitor; we also investigated the underlying molecular mechanisms in the cortisone-induced 3T3-L1 adipogenesis model system and diet-induced obese (DIO) mice. KR-67183 concentration-dependently inhibited 11β-HSD1 activity in human and mouse 11β-HSD1 over-expressed cells and in the ex vivo assay of C57BL/6 mice. In the study with DIO mice, the administration of KR-67183 (20 and 50mg/kg/day, orally for 28 days) improved the glucose tolerance and insulin sensitivity with suppressed 11β-HSD1 activity in the liver and fat. However, KR-67183 showed no change in the adrenal gland weight/body weight ratio and plasma corticosterone concentration in DIO mice. Further, KR-67183 suppressed adipocyte differentiation on cortisone-induced adipogenesis in 3T3-L1 cells is associated with the suppression of the cortisone-induced mRNA levels of FABP4, PPARγ2 and GLUT4, and 11β-HSD1 activity. Taken together, it is suggested that a selective 11β-HSD1 inhibitor, KR-67183, may provide a new therapeutic window in the prevention and treatment without toxicity in type 2 diabetes with obesity.

Small Molecule 11β- Hydroxysteroid Dehydrogenase Type 1 Inhibitors

11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) catalyzes the interconversion of inactive cortisone to active cortisol in a NADPH dependent manner. Excess cortisol or 11β-HSD1 leads to insulin resistance and metabolic syndrome. Inhibition of 11β-HSD1 activity has been pursued vigorously by the pharmaceutical industry as a potential therapeutic strategy for the treatment of type 2 diabetes. As a result, a large number of chemical classes have been identified as potent and selective small molecule inhibitors for 11β-HSD1. Here we review the recent progress in the discovery and development of small molecule inhibitors of 11β-HSD1 by highlighting the medicinal chemistry, SAR, in vivo pharmacodynamic effects and efficacy of a few representative classes of inhibitors in models of diabetes. Furthermore, we also review the structural characteristics of each class of inhibitors by analyzing the inhibitor co-crystal structures of 11β-HSD1.

Discovery of Adamantyl Heterocyclic Ketones as Potent 11β‐Hydroxysteroid Dehydrogenase Type 1 Inhibitors

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) plays a key role in converting intracellular cortisone to physiologically active cortisol, which is implicated in the development of several phenotypes of metabolic syndrome. Inhibition of 11β-HSD1 activity with selective inhibitors has beneficial effects on various conditions, including diabetes, dyslipidemia and obesity, and therefore constitutes a promising strategy to discover novel therapies for metabolic and cardiovascular diseases. A series of novel adamantyl heterocyclic ketones provides potent and selective inhibitors of human 11β-HSD1. Lead compounds display low nanomolar inhibition against human and mouse 11β-HSD1 and are selective with no activity against 11β-HSD2 and 17β-HSD1. Selected potent 11β-HSD1 inhibitors show moderate metabolic stability upon incubation with human liver microsomes and weak inhibition of human CYP450 enzymes.

Anti-diabetic and anti-adipogenic effects of a novel selective 11β-hydroxysteroid dehydrogenase type 1 inhibitor, 2-(3-benzoyl)-4-hydroxy-1,1-dioxo-2H-1,2-benzothiazine-2-yl-1-phenylethanone (KR-66344)

The selective inhibitors of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) have considerable potential for treating type 2 diabetes mellitus and metabolic syndrome. In the present study, we investigated the anti-diabetic and anti-adipogenic effects of 2-(3-benzoyl)-4-hydroxy-1,1-dioxo-2H-1,2-benzothiazine-2-yl-1-phenylethanone (KR-66344), as a 11β-HSD1 inhibitor; we also investigated the underlying molecular mechanisms in the cortisone-induced 3T3-L1 adipogenesis model system and C57BL/6-Lep ob/ob mice. KR-66344 concentration-dependently inhibited 11β-HSD1 activity in human liver microsome, mouse C2C12 myotube and human SW982 cells. In the C57BL/6-Lep ob/ob mice study, the administration of KR-66344 (200 mg/kg/d, orally for 5 days) improved the glucose intolerance as determined by the oral glucose tolerance test, in which the area under the curve (AUC) of the plasma glucose concentration was significantly reduced by 27% compared with the vehicle treated group. Further, KR-66344 suppressed adipocyte differentiation on cortisone-induced adipogenesis in 3T3-L1 cells is associated with the suppression of the cortisone-induced mRNA levels of FABP4, G3PD, PPARγ2 and Glut4, and 11β-HSD1 expression and activity. Our results additionally demonstrate evidence showing that KR-66344 improved glycemic control and inhibited adipogenesis via 11β-HSD1 enzyme activity. Taken together, these results may provide evidence of the therapeutic potential of KR-66344, as a 11β-HSD1 inhibitor, in obesity and type 2 diabetes patients with metabolic syndrome.

Synthesis and 11β hydroxysteroid dehydrogenase 1 inhibition of thiazolidine derivatives with an adamantyl group

A new series of thiazolidine derivatives with an adamantyl group was synthesized and evaluated for their ability to inhibit 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1). Our initial compound 5a showed a weak inhibitory activity. Significant improvements in potency were achieved by substituent modification. The potent compound 8g (E) showed good in vitro inhibitory activity toward human 11β-HSD1, selectivity toward 11β-HSD2, metabolic stability, pharmacokinetic, and safety profile. Furthermore, this compound significantly inhibited 11β-HSD1 activity in rat and monkey models, and showed improved glycemic control in KKAy mice.

17β-Estradiol inhibits 11β-hydroxysteroid dehydrogenase type 1 activity in rodent adipocytes

17Beta-estradiol (E(2)) serves as an anti-obesity steroid; however, the mechanism underlying this effect has not been fully clarified. The effect of E(2) on adipocytes opposes that of glucocorticoids, which potentiate adipogenesis and anabolic lipid metabolism. The key to the intracellular activation of glucocorticoid in adipocytes is 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), which catalyses the production of active glucocorticoids (cortisol in humans and corticosterone in rodents) from inactive 11-keto steroids (cortisone in humans and 11-dehydrocorticosterone in rodents). Using differentiated 3T3-L1 adipocytes, we showed that E(2) inhibited 11beta-HSD1 activity. Estrogen receptor (ER) antagonists, ICI-182 780 and tamoxifen, failed to reverse this inhibition. A significant inhibitory effect of E(2) on 11beta-HSD1 activity was observed within 5-10 min. Furthermore, acetylation or alpha-epimerization of 17-hydroxy group of E(2) attenuated the inhibitory effect on 11beta-HSD1. These results indicate that the inhibition of 11beta-HSD1 by E(2) depends on neither an ER-dependent route, transcriptional pathway nor non-specific fashion. Hexose-6-phosphate dehydrogenase, which provides the cofactor NADPH for full activation of 11beta-HSD1, was unaffected by E(2). A kinetic study revealed that E(2) acted as a non-competitive inhibitor of 11beta-HSD1. The inhibitory effect of E(2) on 11beta-HSD1 was reproduced in adipocytes isolated from rat mesenteric fat depots. This is the first demonstration that E(2) inhibits 11beta-HSD1, thereby providing a novel insight into the anti-obesity mechanism of estrogen.

Insulin and Dexamethasone Dynamically Regulate Adipocyte 11β-Hydroxysteroid Dehydrogenase Type 1

The adipocyte enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) amplifies local glucocorticoid action by generating active glucocorticoids from inactive metabolites and has emerged as a key player in the pathogenesis of central obesity and metabolic syndrome. However, the regulation of adipocyte 11beta-HSD1 is incompletely understood. Therefore, the present study was designed to investigate the effects of insulin and glucocorticoid as well as their underlying molecular mechanisms on 11beta-HSD1 activity and expression in 3T3-L1 adipocytes and determine whether the in vitro findings could be confirmed in vivo. Our main in vitro findings are 1) insulin stimulated whereas dexamethasone inhibited 11beta-HSD1 activity and expression in a time- and concentration-dependent manner; 2) the effect of dexamethasone was mimicked by both cortisol and corticosterone but blocked by the glucocorticoid receptor antagonist RU486; 3) the p38 MAPK inhibitor SB220025, but not the ERK inhibitor U0126 or the phosphatidylinositol 3-kinase inhibitor LY294002, prevented insulin stimulation of 11beta-HSD1 activity; and 4) although dexamethasone did not alter the half-life of 11beta-HSD1 mRNA, insulin doubled it. Taken together, these in vitro results demonstrate that insulin stimulates adipocyte 11beta-HSD1 through a posttranscriptional mechanism that involves activation of the p38 MAPK signaling pathway, whereas dexamethasone exerts an opposite effect by a glucocorticoid receptor-mediated transcriptional mechanism. In contrast, both insulin and dexamethasone augmented 11beta-HSD1 activity and expression in rat white adipose tissue in vivo, thus confirming the role of insulin but revealing a fundamental difference regarding the role of dexamethasone in regulating adipocyte 11beta-HSD1 between the two model systems.

Implication of cortisol and 11β-hydroxysteroid dehydrogenase enzymes in the development of porcine (Sus scrofa domestica) ovarian follicles and cysts

This study investigated cortisol inactivation by 11beta-hydroxysteroid dehydrogenase (11beta HSD) enzymes in porcine granulosa cells from antral follicles at different developmental stages and in ovarian cysts. In granulosa cells, cortisol oxidation increased threefold with antral follicle diameter (P < 0.001). This trend was paralleled by a threefold increase in NADP(+)-dependent 11beta-dehydrogenase activity in granulosa cell homogenates with follicle diameter. Intact granulosa cells from ovarian cysts exhibited significantly lower enzyme activities than cells from large antral follicles. Neither intact cells norcell homogenates displayed net 11-ketosteroid reductase activities. Since porcine follicular fluid (FF) from large antral follicles and ovarian cysts contains hydrophobic inhibitors of glucocorticoid metabolism by type 1 11beta HSD, this studyalso investigated whether levels of 11beta HSD inhibitors changed during follicle growth and could affect cortisol metabolism in granulosa cells. The extent of inhibition of 11beta HSD1 activity in rat kidney homogenates decreased progressively from 50 +/- 8% inhibition by FF from small antral follicles (P < 0.001) to 23 +/- 6% by large antral FF (P < 0.05). Cyst fluid inhibited 11beta HSD1 activity by 59 +/- 4% (P < 0.001). Likewise, net cortisol oxidation in granulosa cells was significantly decreased by large antral FF (35-48% inhibition, P < 0.05) and cyst fluid (45-75% inhibition, P < 0.01). We conclude that inactivation of cortisol by 11beta HSD enzymes in porcine granulosa cells increases with follicle development but is significantly decreased in ovarian cysts. Moreover, changes in ovarian cortisol metabolism are accompanied by corresponding changes in the levels of paracrine inhibitors of 11beta HSD1 within growing ovarian follicles and cysts, implicating cortisol in follicle growth and cyst development.