Guirui Yan

Sildenafil promotes adipogenesis through a PKG pathway

Sildenafil is the first oral PDE5 inhibitor for the treatment of erectile dysfunction and pulmonary arterial hypertension. In the present study, we investigated the effect of sildenafil on adipogenesis in 3T3L1 preadipocytes. Treatment with sildenafil for 8 days significantly promoted adipogenesis characterized by increased lipid droplet and triglyceride content in 3T3L1 cells. Meanwhile, sildenafil induced a pronounced up-regulation of the expression of adipocyte-specific genes, such as aP2 and GLUT4. The results by RT-PCR and Western blotting further showed that sildenafil increased the sequential expression of C/EBP beta, PPAR gamma and C/EBP alpha. Additionally, we found that the other two PDE5 inhibitors (vardenafil and tadalafil) and the cGMP analog 8-pCPT-cGMP also increased adipogenesis. Likewise, 8-pCPT-cGMP could up-regulate the expression of adipogenic and adipocyte-specific genes. Importantly, the PKG inhibitor Rp-8-pCPT-cGMP was able to inhibit both sildenafil and 8-pCPT-cGMP-induced adipogenesis. Furthermore, sildenafil promoted basal and insulin-mediated glucose uptake in 3T3L1 cells, which was counteracted by Rp-8-pCPT-cGMP. These results indicate that sildenafil could promote adipogenesis accompanied by increased glucose uptake through a PKG pathway at least partly.

PDE5 inhibitor promotes melanin synthesis through the PKG pathway in B16 melanoma cells

PDE inhibitors could increase cellular cGMP levels and are used to treat erectile dysfunction as well as pulmonary arterial hypertension. cGMP production was reported to be necessary for UVB‐induced melanin synthesis, however, the effect of PDE5 inhibitor on melanin synthesis has not been examined. We found that PDE5 inhibitor (sildenafil or vardenafil) and the cGMP analog 8‐CPT‐cGMP stimulated CREB phosphorylation, leading to increased tyrosinase expression and melanin synthesis, which was counteracted by KT5823, a selective cGMP‐dependent protein kinase (PKG) inhibitor. However, KT5823 did not affect cAMP‐elevating agent‐mediated melanin synthesis, indicating that KT5823 selectively inhibited cGMP‐induced melanin synthesis. This is the first study to find that PDE5 inhibitor can promote melanin synthesis and reveal that PKG‐dependent CREB phosphorylation and tyrosinase expression is involved in cGMP‐induced melanin synthesis. Our results suggest that PDE5 inhibitor may be beneficial for the treatment of hypopigmentation diseases. J. Cell. Biochem. 113: 2738–2743, 2012.

Discovery of highly selective inhibitors of human fatty acid binding protein 4 (FABP4) by virtual screening

In this study, a series of small molecule inhibitors of human FABP4 were identified through virtual screening. Compound 1 is the most potent hit against FABP4 with a selectivity of more than 144-fold preferences over human FABP3. In addition, MD simulation and mutation studies revealed key residues for inhibitory potency and selectivity, which provides a guideline for further drug design against obesity, diabetes and atherosclerosis.

Benzbromarone, an old uricosuric drug, inhibits human fatty acid binding protein 4 in vitro and lowers the blood glucose level in db/db mice.

Aim:Fatty acid-binding protein 4 (FABP4) plays an important role in maintaining glucose and lipid homeostasis. The aim of this study was to find new inhibitors of FABP4 for the treatment of type 2 diabetes.Methods:Human FABP4 protein was expressed, and its inhibitors were detected in 1,8-ANS displacement assay. The effect of the inhibitor on lipolysis activity was examined in mouse 3T3-L1 preadipocytes. The db/db mice were used to evaluate the anti-diabetic activity of the inhibitor. Molecular docking and site-directed mutagenesis studies were carried out to explore the binding mode between the inhibitor and FABP4.Results:From 232 compounds tested, benzbromarone (BBR), an old uricosuric drug, was discovered to be the best inhibitor of FABP4 with an IC50 value of 14.8 μmol/L. Furthermore, BBR (25 μmol/L) significantly inhibited forskolin-stimulated lipolysis in 3T3-L1 cells. Oral administration of BBR (25 or 50 mg/kg, for 4 weeks) dose-dependently reduced the blood glucose level and improved glucose tolerance and insulin resistance in db/db mice. Molecular docking revealed that the residues Ser55, Asp76, and Arg126 of FABP4 formed important interactions with BBR, which was confirmed by site-directed mutagenesis studies.Conclusion:BBR is an inhibitor of FABP4 and a potential drug candidate for the treatment of type 2 diabetes and atherosclerosis.

New Isoprenylated 2-Arylbenzofurans and Pancreatic Lipase Inhibitory Constituents from Artocarpus nitidus

Two new isoprenylated 2‐arylbenzofurans, artonitidin A (=(2′R)‐2′,3′‐dihydro‐2′‐(1‐hydroxy‐1‐methylethyl)‐5′,7‐bis(3‐methylbut‐2‐en‐1‐yl)‐2,4′‐bi‐1‐benzofuran‐6,6′‐diol; 1) and artonitidin B (=5‐[6‐hydroxy‐7‐(3‐methylbut‐2‐en‐1‐yl)‐1‐benzofuran‐2‐yl]‐4‐(3‐methylbut‐2‐en‐1‐yl)benzene‐1,3‐diol; 2), together with 14 known compounds, 3–16, were isolated from the stems of Artocarpus nitidus Trec. The structures were elucidated by spectroscopic methods. Norartocarpin (3), cudraflavone C (5), brosimone I (8), artotonkin (11), albanin A (13), and artopetelin M (14) showed inhibitory effects on pancreatic lipase with IC50 values ranging from 1.8±0.1 to 63.8±3.6 μM.

New isoprenylated flavones and stilbene derivative from Artocarpus hypargyreus.

Three new isoprenylated flavones, hypargyflavones A–C (1–3, resp.), and one novel stilbene derivative, hypargystilbene A (4), together with seven known compounds, 5–11, were isolated from the stems of Artocarpus hypargyreus Hance. The structures were elucidated by spectroscopic methods. Hypargyflavone A (1), cudraflavone C (8), brosimone I (10), and norartocarpin (11) showed inhibitory effects on pancreatic lipase.

2-Arylbenzofuran and tyrosinase inhibitory constituents of Morus notabilis

Phytochemical investigation of the stem of Morus notabilis led to the isolation and characterization of 10 compounds of 2-arylbenzofurans (1–10), including two new compounds, (2′R)-2′,3′-dihydro-2′-(1-hydroxy-1-methylethyl)-2,6′-bibenzofuran-6,4′-diol (1) and 5,6-dimethoxy-2-(3-hydroxy-5-methoxyphenyl)benzofuran (2). Moracins O (6) and P (10) showed inhibitory effects on mushroom tyrosinase with IC50 values being lower than that of kojic acid.

Novel fatty acid binding protein 4 (FABP4) inhibitors: virtual screening, synthesis and crystal structure determination.

Fatty acid binding protein 4 (FABP4) is a potential drug target for diabetes and atherosclerosis. For discovering new chemical entities as FABP4 inhibitors, structure-based virtual screening (VS) was performed, bioassay demonstrated that 16 of 251 tested compounds are FABP4 inhibitors, among which compound m1 are more active than endogenous ligand linoleic acid (LA). Based on the structure of m1, new derivatives were designed and prepared, leading to the discovery of two more potent inhibitors, compounds 9 and 10. To further explore the binding mechanisms of these new inhibitors, we determined the X-ray structures of the complexes of FABP4-9 and FABP4-10, which revealed similar binding conformations of the two compounds. Residue Ser53 and Arg126 formed direct hydrogen bonding with the ligands. We also found that 10 could significantly reduce the levels of lipolysis on mouse 3T3-L1 adipocytes. Taken together, in silico, in vitro and crystallographic data provide useful hints for future development of novel inhibitors against FABP4.

Combining pharmacophore, docking and substructure search approaches to identify and optimize novel B-RafV600E inhibitors

In this study for searching novel B-Raf(V600E) inhibitors, pharmacophore-based virtual screening identified 1 as a hit bearing 5-benzylidene-2-thioxodihydropyrimidine-4,6(1H,5H)-dione. Based on 1, scaffold hopping inspired by molecular docking discovered 5-(furan-2-ylmethylene)-2-thioxodihydropyrimidine-4,6(1H,5H)-dione as a new and better scaffold. Substructure search with the new scaffold identified 28 active compounds, among which 12 compounds (42.9%) showed IC(50) less than 1 μM. Especially, compound 3o, which is 10-fold more potent than the hit 1, is a potent inhibitor comparable to that of the marketed drug vemurafenib.

Pharmacological inhibition of diacylglycerol acyltransferase 1 reduces body weight gain, hyperlipidemia, and hepatic steatosis in db/db mice

AbstractAim:To test whether pharmacological inhibition of Diacylglycerol acyltransferase 1 (DGAT1) by a small-molecule inhibitor H128 can improve metabolism disorders in leptin receptor-deficient db/db mice.Methods:To investigate the effect of H128 on intestinal fat absorption,db/db mice were acutely given a bolus of corn oil by gavage. The mice were further orally administered H128 (3 and 10 mg/kg) for 5 weeks. Blood glucose, lipids, insulin, ALT, and AST as well as hepatic triglycerides were measured. The insulin tolerance test was performed to evaluate insulin sensitivity. The expression of genes involved in fatty acid oxidation was detected by RT-PCR.Results:Oral administration of H128 (10 mg/kg) acutely inhibited intestinal fat absorption following a lipid challenge in db/db mice. Chronic treatment with H128 significantly inhibited body weight gain, decreased food intake, and induced a pronounced reduction of serum triglycerides. In addition, H128 treatment markedly ameliorated hepatic steatosis, characterized by decreased liver weight, lipid droplets, and triglyceride content as well as serum ALT and AST levels. Furthermore, H128 treatment increased the expression of the CPT1 and PPARα genes in liver, suggesting that H128 enhanced fatty acid oxidation in db/db mice. However, neither blood glucose nor insulin tolerance was affected by H128 treatment throughout the 5-week experimental period.Conclusion:DGAT1 may be an effective therapeutic target for the treatment of obesity, hyperlipidemia and hepatic steatosis.