Inhibition of acetyl-CoA carboxylase (ACC), with its resultant inhibition of fatty acid synthesis and stimulation of fatty acid oxidation, has the potential to favorably affect the multitude of cardiovascular risk factors associated with the metabolic syndrome. To achieve maximal effectiveness, an ACC inhibitor should inhibit both the lipogenic tissue isozyme (ACC1) and the oxidative tissue isozyme (ACC2). Herein, we describe the biochemical and acute physiological properties of CP-610431, an isozyme-nonselective ACC inhibitor identified through high throughput inhibition screening, and CP-640186, an analog with improved metabolic stability. CP-610431 inhibited ACC1 and ACC2 with IC 50 s of ϳ50 nM. Inhibition was reversible, uncompetitive with respect to ATP, and non-competitive with respect to bicarbonate, acetyl-CoA, and citrate, indicating interaction with the enzymatic carboxyl transfer reaction. CP-610431 also inhibited fatty acid synthesis, triglyceride (TG) synthesis, TG secretion, and apolipoprotein B secretion in HepG2 cells (ACC1) with EC 50 s of 1.6, 1.8, 3.0, and 5.7 M, without affecting either cholesterol synthesis or apolipoprotein CIII secretion. CP-640186, also inhibited both isozymes with IC 50 s of ϳ55 nM but was 2-3 times more potent than CP-610431 in inhibiting HepG2 cell fatty acid and TG synthesis. CP-640186 also stimulated fatty acid oxidation in C2C12 cells (ACC2) and in rat epitrochlearis muscle strips with EC 50 s of 57 nM and 1.3 M. In rats, CP-640186 lowered hepatic, soleus muscle, quadriceps muscle, and cardiac muscle malonyl-CoA with ED 50 s of 55, 6, 15, and 8 mg/kg. Consequently, CP-640186 inhibited fatty acid synthesis in rats, CD1 mice, and ob/ob mice with ED 50 s of 13, 11, and 4 mg/kg, and stimulated rat whole body fatty acid oxidation with an ED 50 of ϳ30 mg/kg. Taken together, These observations indicate that isozyme-nonselective ACC inhibition has the potential to favorably affect risk factors associated with the metabolic syndrome.

Sterol regulatory element-binding proteins (SREBPs) are major transcriptional regulators of cholesterol, fatty acid, and glucose metabolism. Genetic disruption of SREBP activity reduces plasma and liver levels of cholesterol and triglycerides and insulin-stimulated lipogenesis, suggesting that SREBP is a viable target for pharmacological intervention. The proprotein convertase SREBP site 1 protease (S1P) is an important posttranscriptional regulator of SREBP activation. This report demonstrates that 10 M PF-429242 (Bioorg Med Chem Lett 17:4411-4414, 2007), a recently described reversible, competitive aminopyrrolidineamide inhibitor of S1P, inhibits endogenous SREBP processing in Chinese hamster ovary cells. The same compound also down-regulates the signal from an SREluciferase reporter gene in human embryonic kidney 293 cells and the expression of endogenous SREBP target genes in cultured HepG2 cells. In HepG2 cells, PF-429242 inhibited cholesterol synthesis, with an IC 50 of 0.5 M. In mice treated with PF-429242 for 24 h, the expression of hepatic SREBP target genes was suppressed, and the hepatic rates of cholesterol and fatty acid synthesis were reduced. Taken together, these data establish that small-molecule S1P inhibitors are capable of reducing cholesterol and fatty acid synthesis in vivo and, therefore, represent a potential new class of therapeutic agents for dyslipidemia and for a variety of cardiometabolic risk factors associated with diabetes, obesity, and the metabolic syndrome.

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Phospholipid transfer protein (PLTP) facilitates the transfer of phospholipids from triglyceride-rich lipoproteins into HDL. PLTP has been shown to be an important factor in lipoprotein metabolism and atherogenesis. Here, we report that chronic high-fat, high-cholesterol diet feeding markedly increased plasma cholesterol levels in C57BL/6 mice. PLTP deficiency attenuated diet-induced hypercholesterolemia by dramatically reducing apolipoprotein E-rich lipoproteins (288%) and, to a lesser extent, LDL (240%) and HDL (235%). Increased biliary cholesterol secretion, indicated by increased hepatic ABCG5/ABCG8 gene expression, and decreased intestinal cholesterol absorption may contribute to the lower plasma cholesterol in PLTP-deficient mice. The expression of proinflammatory genes (intercellular adhesion molecule-1 and vascular cell adhesion molecule-1) is reduced in aorta of PLTP knockout mice compared with wild-type mice fed either a chow or a high-cholesterol diet. Furthermore, plasma interleukin-6 levels are significantly lower in PLTP-deficient mice, indicating reduced systemic inflammation. These data suggest that PLTP appears to play a proatherogenic role in dietinduced hyperlipidemic mice. Plasma phospholipid transfer protein (PLTP) plays an important role in the metabolism of lipoproteins (1). PLTP belongs to the family of lipid transfer/lipopolysaccharide binding proteins, including cholesteryl ester transfer protein, lipopolysaccharide binding protein, and bactericidal permeability-increasing protein (2, 3). It has been shown that PLTP facilitates the transfer and exchange of phospholipids between VLDL and HDL (4). It also transfers phospholipids between HDL particles that result in the conversion of HDL 3 into larger and smaller HDL particles (5, 6). PLTP can also bind several other amphipathic molecules, including a-tocopherol, diacylglycerides, cerebrosides, and lipopolysaccharides (7). Several clinical studies suggest that high plasma PLTP activity is a risk factor for coronary artery disease and a determinant of carotid intimamedia thickness in type 2 diabetes mellitus (8, 9).Studies using genetically modified mice strongly suggest that PLTP functions as a proatherogenic factor (10-12). PLTP deficiency causes a marked decrease in HDL lipids and apolipoprotein A-I (apoA-I), as a result of higher catabolism, in both chow-fed and 2 week Western diet-fed mice (13,14). The absence of PLTP in hyperlipidemic apoE-deficient and human apoB transgenic mouse strains results in reduced production in plasma levels of apoBcontaining lipoproteins, mostly LDL (10). Atherosclerotic lesion areas are also decreased in PLTP knockout mice in the LDL receptor-or apoE-deficient or apoB transgenic background, despite decreased HDL. Furthermore, reduced lipoprotein oxidation and improved anti-inflammatory properties of HDL in PLTP knockout mice may also contribute to the antiatherogenic potential in these mouse models (15,16). The proatherogenic role of PLTP is further supported by results that demonstrate increased ...

Phospholipid transfer protein (PLTP) plays an important role in atherogenesis, and its function goes well beyond that of transferring phospholipids between lipoprotein particles. Previous studies showed that genetic deficiency of PLTP in mice causes a substantially impaired hepatic secretion of apolipoprotein-B (apoB), the major protein of atherogenic lipoproteins. To understand whether the impaired apoB secretion is a direct result from lack of PLTP activity, in this study, we further investigated the function of PLTP in apoB secretion by using PLTP inhibitors. We identified a series of compounds containing a 3-benzazepine core structure that inhibit PLTP activity. Compound A, the most potent inhibitor, was characterized further and had little cross-reactivity with microsomal triglyceride transfer protein. Compound A reduced apoB secretion in human hepatoma cell lines and mouse primary hepatocytes. Furthermore, we confirmed that the reduction of apoB secretion mediated by compound A is PLTP-dependent, because the PLTP inhibitor had no effect on apoB secretion from PLTP-deficient hepatocytes. These studies provided evidence that PLTP activity regulates apoB secretion and pharmacologic inhibition of PLTP may be a new therapy for dyslipidemia by reducing apoB secretion.