Marc Issandou

Inhibition of lipid synthesis through activation of AMP kinase: an additional mechanism for the hypolipidemic effects of berberine

The alkaloid drug berberine (BBR) was recently described to decrease plasma cholesterol and triglycerides (TGs) in hypercholesterolemic patients by increasing expression of the hepatic low density lipoprotein receptor (LDLR). Using HepG2 human hepatoma cells, we found that BBR inhibits cholesterol and TG synthesis in a similar manner to the AMP-activated protein kinase (AMPK) activator 5-aminoimidazole-4-carboxamide 1-β-ribofuranoside (AICAR). Significant increases in AMPK phosphorylation and AMPK activity were observed when the cells were incubated with BBR. Activation of AMPK was also demonstrated by measuring the phosphorylation of acetyl-CoA carboxylase, a substrate of AMPK, correlated with a subsequent increase in fatty acid oxidation. All of these effects were abolished by the mitogen-activated protein kinase kinase inhibitor PD98059. Treatment of hyperlipidemic hamsters with BBR decreased plasma LDL cholesterol and strongly reduced fat storage in the liver. These findings indicate that BBR, in addition to upregulating the LDLR, inhibits lipid synthesis in human hepatocytes through the activation of AMPK. These effects could account for the strong reduction of plasma TGs observed with this drug in clinical trials.

SCAP ligands are potent new lipid-lowering drugs

Upregulation of low-density lipoprotein receptor (LDLr) is a key mechanism to control elevated plasma LDL-cholesterol levels. Here we identify a new class of compounds that directly binds to the sterol regulatory element-binding protein (SREBP) cleavage-activating protein (SCAP). We show that a 14C-labeled, photo-activatable analog specifically labeled both SCAP and a truncated form of SCAP containing the sterol-sensing domain. When administered to hyperlipidemic hamsters, SCAP ligands reduced both LDL cholesterol and triglycerides levels by up to 80% with a three-fold increase in LDLr mRNA in the livers. Using human hepatoma cells, we show that these compounds act through the sterol-responsive element of the LDLr promoter and activate the SCAP/SREBP pathway, leading to increased LDLr expression and activity, even in presence of excess of sterols. These findings have led to the identification of a class of compounds that represent a promising new class of hypolipidemic drugs.

Pharmacological inhibition of stearoyl-CoA desaturase 1 improves insulin sensitivity in insulin-resistant rat models.

Stearoyl-CoA Desaturase 1 (SCD1) is a central enzyme that catalyzes the biosynthesis of monounsaturated fatty acids from saturated fatty acids. SCD1 is an emerging target in obesity and insulin resistance due to the improved metabolic profile obtained when the enzyme is genetically inactivated. Here, we have investigated if the pharmacological inhibition of SCD1 could elicit the same profile. We have identified a small molecule, GSK993 and characterized it as a potent and orally available SCD1 inhibitor. In Zucker(fa/fa) rats, GSK993 exerted a marked reduction in hepatic lipids as well as a significant improvement of glucose tolerance. Furthermore, in a diet-induced insulin resistant rat model, GSK993 induced a very strong reduction in Triton-induced hepatic Very Low Density Lipoprotein-Triglyceride production. In addition, following a hyperinsulinemic-euglycemic clamp in GSK993-treated animals, we observed an improvement in the whole body insulin sensitivity as reflected by an increase in the glucose infusion rate. Taken together, these findings demonstrate that the pharmacological inhibition of SCD1 translates into improved lipid and glucose metabolic profiles and raises the interest of SCD1 inhibitors as potential new drugs for the treatment of insulin resistance.

Liver X receptor activation promotes macrophage-to-feces reverse cholesterol transport in a dyslipidemic hamster model

Liver X receptor (LXR) activation promotes reverse cholesterol transport (RCT) in rodents but has major side effects (increased triglycerides and LDL-cholesterol levels) in species expressing cholesteryl ester transfer protein (CETP). In the face of dyslipidemia, it remains unclear whether LXR activation stimulates RCT in CETP species. We therefore used a hamster model made dyslipidemic with a 0.3% cholesterol diet and treated with vehicle or LXR agonist GW3965 (30 mg/kg bid) over 10 days. To investigate RCT, radiolabeled 3H-cholesterol macrophages or 3H-cholesteryl oleate-HDL were then injected to measure plasma and feces radioactivity over 72 or 48 h, respectively. The cholesterol-enriched diet increased VLDL-triglycerides and total cholesterol levels in all lipoprotein fractions and strongly increased liver lipids. Overall, GW3965 failed to improve both dyslipidemia and liver steatosis. However, after 3H-cholesterol labeled macrophage injection, GW3965 treatment significantly increased the 3H-tracer appearance by 30% in plasma over 72 h, while fecal 3H-cholesterol excretion increased by 156% (P < 0.001). After 3H-cholesteryl oleate-HDL injection, GW3965 increased HDL-derived cholesterol fecal excretion by 64% (P < 0.01 vs. vehicle), while plasma fractional catabolic rate remained unchanged. Despite no beneficial effect on dyslipidemia, LXR activation promotes macrophage-to-feces RCT in dyslipidemic hamsters. These results emphasize the use of species with a more human-like lipoprotein metabolism for drug profiling.

Design and evaluation of a novel series of 2,3-oxidosqualene cyclase inhibitors with low systemic exposure, relationship between pharmacokinetic properties and ocular toxicity.

We describe the discovery of novel potent inhibitors of 2,3-oxidosqualene:lanosterol cyclase inhibitors (OSCi) from a focused pharmacophore-based screen. Optimization of the most tractable hits gave a series of compounds showing inhibition of cholesterol biosynthesis at 2mg/kg in the rat with distinct pharmacokinetic profiles. Two compounds were selected for toxicological study in the rat for 21 days in order to test the hypothesis that low systemic exposure could be used as a strategy to avoid the ocular side effects previously described with OSCi. We demonstrate that for this series of inhibitors, a reduction of systemic exposure is not sufficient to circumvent cataract liabilities.

Selective CETP Inhibition and PPARα Agonism Increase HDL Cholesterol and Reduce LDL Cholesterol in Human ApoB100/Human CETP Transgenic Mice

Cholesteryl ester transfer protein (CETP) plays a key role in high-density lipoprotein (HDL) cholesterol metabolism, but normal mice are deficient in CETP. In this study, transgenic mice expressing both human apolipoprotein B 100 (ApoB-100) and human CETP (hApoB100/hCETP) were used to characterize the effects of CETP inhibition and peroxisome proliferator—activated receptor α (PPARα) agonism on lipid profiles. Torcetrapib (3, 10, and 30 mg/kg), a CETP inhibitor, fenofibrate (30 mg/kg), a weak PPARα agonist, and GW590735 (3 and 10 mg/kg), a potent and selective PPARα agonist were given orally for 14 days to hApoB100/hCETP mice and lipid profiles were assessed. The average percentages of HDL, low-density lipoprotein (LDL), and very-low-density lipoprotein (VLDL) cholesterol fractions in hApoB100/hCETP mice were 34.8%, 61.6%, and 3.6%, respectively, which is similar to those of normolipidemic humans. Both torcetrapib and fenofibrate significantly increased HDL cholesterol and reduced LDL cholesterol, and there was a tendency for torcetrapib to reduce VLDL cholesterol and triglycerides. GW590735 significantly increased HDL cholesterol, decreased LDL and VLDL cholesterol, and significantly reduced triglycerides. Maximal increases in HDL cholesterol were 37%, 53%, and 84% with fenofibrate, torcetrapib, and GW590735, respectively. These results, in mice that exhibit a more human-like lipid profile, demonstrate an improved lipid profile with torcetrapib, fenofibrate, and GW590735, and support the use of selective PPARα agonism for the treatment of lipid disorders. In addition, these data demonstrate the use of hApoB100/hCETP transgenic mice to identify, characterize, and screen compounds that increase HDL cholesterol.