Discovery of novel phosphonate derivatives as hepatitis C virus NS3 protease inhibitors

A novel class of phosphonate derivatives was designed to mimic the interaction of product-like carboxylate based inhibitors of HCV NS3 protease. A phosphonic acid (compound 2) was demonstrated to be a potent HCV NS3 protease inhibitor, and a potential candidate for treating HCV infection. The syntheses and preliminary biological evaluation of this phosphonate class of inhibitor are described.     

Novel, potent and orally bioavailable indolizidinone-derived inhibitors of the hepatitis C virus NS3 protease

A novel, potent, and orally bioavailable class of product-like inhibitors of the HCV NS3 protease was discovered by constraining the P2-P3 amide bond and the P3 hydrocarbon substituent to the protease-bound conformation. This preorganization was accomplished by incorporation of the P2-P3 amide into a six-membered ring attached to the P2-proline 5-position. Isothermal calorimetric characterization of the role of hydrocarbon substitution of this six-membered ring, upon binding the HCV NS3 protease, was found to be exclusively entropic in nature. The synthesis, preliminary SAR and pharmacokinetic profiling of this compact, indolizidinone-derived scaffold are described.     

Design, synthesis and evaluation of a series of novel fumagillin analogues

A series of fumagillin analogues targeted at understanding tolerability of MetAP2 toward substitution at C4 and C6 were synthesized. Initially, the C6 side chain was maintained as cinnamoyl ester and C4 was modified. It was concluded that replacing the natural C4 of fumagillin with a benzyl oxime at C4 resulted in moderate loss of activity toward binding to MetAP2. Placement of a primary or secondary carbamate at C6 did not improve the potency of compounds toward inhibition of MetAP2. However, the inhibitory activity against MetAP2 was gained back by placing polar groups such as piperazinyl carbamate at C6. Small alkyl substituents on the amine of piperazinyl carbamate were well tolerated.     

Traditional Chinese Medicine improves dysfunction of peroxisome proliferator-activated receptor alpha and microsomal triglyceride transfer protein on abnormalities in lipid metabolism in ethanol-fed rats

We report the effects of Traditional Chinese Medicine (TCM) on alcohol-induced fatty liver in rats. TCM consists of Astragalus membranaceus, Morus alba, Crataegus pinnatifida, Alisma oriental, Salvia miltiorrhiza and Pueraria lobata. The rats were separated randomly into five groups; the CD group (n=10), which was fed a control diet for 10 weeks, the ED group (n=10), which was fed an isocaloric liquid diet containing ethanol for 10 weeks and given daily oral doses of TCM (0.222 g/kg/day; TCM222, 0.667 g/kg/day; TCM667, and 2.000 g/kg/day; TCM2000, n=10, respectively) over the last four weeks of the study. The ED group developed fatty livers, as determined by their lipid profiles and liver histological findings. Compared with the control group, liver/body weight, plasma triglyceride (TG) and total cholesterol (TC), liver TG and TC, plasma alanine aminotransferase (ALT) and aspartic aminotransferase (AST) significantly increased in the ED group. Also, free fatty acids (FFA) levels increased in both plasma and liver during the administration of ethanol. On the other hand, when rats were administrated with TCM, their liver/body weight, plasma TG, TC and FFA, liver TG, TC and FFA, plasma ALT and AST decreased significantly and the degree of hepatic lipid droplets was markedly improved compared with those in the ED group. Proper function of the peroxisome proliferator-activated receptor alpha (PPARalpha) is essential for the regulation of hepatic fatty acid metabolism. Microsomal triglyceride transfer protein (MTP) is essential for the secretion of triglycerides from the liver. mRNAs for PPARalpha and MTP were reduced in the livers of ethanol-fed rats. TCM restored the mRNA levels of PPARalpha and MTP, and prevented development of fatty livers in ethanol-fed rats. Impairment of PPARalpha and MTP function during ethanol consumption contributes to the development of alcohol-induced fatty liver, which can be overcome by TCM.     

Transduction efficacy of Tat-Cu,Zn-superoxide dismutase is enhanced by copper ion recovery of the fusion protein

We previously reported that Tat-Cu,Zn-superoxide dismutase (Tat-SOD) can be directly transduced into mammalian cells across the lipid membrane barrier. To enhance the therapeutic potential of Tat-SOD for the treatment of various disorders that are related to this antioxidant enzyme, the transduction efficacy of Tat-SOD should be heightened. Therefore, we investigated whether copper ion recovery of the fusion protein could enhance the transduction potential of Tat-SOD in cultured HeLa cells. The results showed that the transduction potential of Tat-SOD was markedly enhanced by copper ions, and moderately increased by zinc ions. Compared with Tat-SOD, the Tat-SOD that recovered the copper ion (CR-Tat-SOD) achieved a significant increase in intracellular concentration and enzymatic activity. Therefore, CR-Tat-SOD was transduced into HeLa cells in a rapid saturation manner, but Tat-SOD was shown in a time-dependent manner. With the higher transduction efficacy of CR-Tat-SOD than that of Tat-SOD, the transduced CR-Tat-SOD significantly increased the viability of HeLa cells that were pretreated with paraquat, an intracellular superoxide anion generator. Although the mechanism of the enhanced transduction of Tat-SOD by copper ions is still unanswered, these results indicate that copper ions facilitate the transduction of SOD. These then significantly increase the biological effectiveness of this antioxidant enzyme.     

Hemolytic activity of a cyclic peptide Ro09-0198 isolated from Streptoverticillium

Ro09-0198, a cyclic peptide isolated from culture filtrates of Streptoverticillium griseoverticillatum, induced lysis of erythrocytes. Preincubation of the peptide with phosphatidylethanolamine reduced the hemolytic activity, whereas other phospholipids present in erythrocytes in nature had no effect. A study of the structural requirements on phosphatidylethanolamine necessary for interaction with the peptide indicates that Ro09-0198 recognizes strictly a particular chemical structure of phosphatidylethanolamine: dialkylphosphoethanolamine as well as 1-acylglycerophosphoethanolamine showed the same inhibitory effect on hemolysis induced by Ro09-0198 as diacylphospatidylethanolamine, whereas phosphoethanolamine gave no inhibitory effect. Neither phosphatidyl-N-monomethylethanolamine nor alkylphosphopropanolamine had an inhibitory effect. Consequently, the hydrophobic chain is necessary for the interaction and the phosphoethanolamine moiety is exactly recognized by the peptide. Ro-09-0198-induced hemolysis was temperature-dependent and the sensitivity of hemolysis differed greatly among animal species.     

Discovery of the bifunctional modulator of angiotensin II type 1 receptor (AT1R) and PPARγ derived from the AT1R antagonist,Fimasartan

Inspired by the well-known PPARγ partial agonism of angiotensin II type 1 receptor (AT1R) antagonists exemplified by an antihypertensive drug, Telmisartan, efforts to identify compounds with the dual activities have been pursued in order to control the two major metabolic disorders, hypertension and hyperglycemia simultaneously. Lead compound 18 derived from the AT1R antagonist, Fimasartan, has successfully presented the possibility to control the medical conditions by a single molecule.     

Effects of intra-abomasal infusion of beta-casomorphins on circulating concentrations of hyperglycaemic insulin and glucose in dairy cows

The effects of intra-abomasal infusion of a mixture of -casomorphins on circulating concentrations of insulin and glucose prestimulated by either abomasal (experiment 1) or intravenous (experiment 2) glucose were studied using non-lactating dairy cows. In both experiments, bolus infusion of 240 mg of a mixture of three beta-casomorphins (beta-casomorphin-4-amide, -5 and -7) was given via an abomasal infusion line. The beta-casomorphins significantly lowered the responses of serum insulin to both abomasal and intravenous glucose infusions (P<0.05). However, the beta-casomorphins did not significantly affect circulating glucose concentrations. The insulinopenic action of the beta-casomorphins is consistent with the action of somatostatin-28 (SS-28) as judged from the effects of SS-28 on the insulin secretion when administered intravenously in experiment 1.     

Characterization of a biosurfactant, mannosylerythritol lipid produced from Candida sp. SY16

One yeast strain, SY16, was selected as a potential producer of a biosurfactant, and identified as a Candida species. A biosurfactant produced from Candida sp. SY16 was purified and confirmed to be a glycolipid. This glycolipid-type biosurfactant lowered the surface tension of water to 29 dyne/cm at critical micelle concentration of 10 mg/l (1.5 × 10⁻⁵ M), and the minimum interfacial tension was 0.1 dyne/cm against kerosene. Thin-layer and high-pressure liquid chromatography studies demonstrated that the glycolipid contained mannosylerythritol as a hydrophilic moiety. The hydrophilic sugar moiety of the biosurfactant was determined to be β-d-mannopyranosyl-(1 → 4)-O-meso-erythritol by nuclear magnetic resonance (NMR) and fast atom bombardment mass–spectroscopy analyses. The hydrophobic moiety, fatty acids, of the biosurfactant was determined to be hexanoic, dodecanoic, tetradecanoic, and tetradecenoic acid by gas chromatography–mass spectroscopy. The structure of the native biosurfactant was determined to be 6-O-acetyl-2,3- di-O-alkanoyl-β-d-mannopyranosyl-(1 → 4)-O-meso-erythritol by NMR analyses. We newly determined that an acetyl group was linked to the C-6 position of the d-mannose unit in the hydrophilic sugar moiety. Received: 18 December 1999 / Received last revision: 2 June 1999 / Accepted: 4 June 1999