Acyl-coenzyme A:cholesterol Acyltransferase Inhibitors Research Articles

YM750, an ACAT Inhibitor, Acts on Adrenocortical Cells to Inhibit Aldosterone Secretion Due to Depolarization.

Primary aldosteronism (PA) is considered the most common form of secondary hypertension, which is associated with excessive aldosterone secretion in the adrenal cortex. The cause of excessive aldosterone secretion is the induction of aldosterone synthase gene (CYP11B2) expression by depolarization of adrenocortical cells. In this study, we found that YM750, an Acyl-coenzyme A: cholesterol acyltransferase (ACAT) inhibitor, acts on adrenocortical cells to suppress CYP11B2 gene expression and aldosterone secretion. YM750 inhibited the induction of CYP11B2 gene expression by KCl stimulation, but not by angiotensin II and forskolin stimulation. Interestingly, YM750 did not inhibit KCl-stimulated depolarization via an increase in intracellular calcium ion concentration. Moreover, ACAT1 expression was relatively abundant in the zona glomerulosa (ZG) including these CYP11B2-positive cells. Thus, YM750 suppresses CYP11B2 gene expression by suppressing intracellular signaling activated by depolarization. In addition, ACAT1 was suggested to play an important role in steroidogenesis in the ZG. YM750 suppresses CYP11B2 gene expression and aldosterone secretion in the adrenal cortex, suggesting that it may be a potential therapeutic agent for PA.

Role of ACAT1 in Atherosclerosis and Alzheimer's Disease

The acyl‐coenzyme A: cholesterol acyltransferase (ACAT) intracellular membrane protein is located in the endoplasmic reticulum and is composed of 427 amino acids. ACAT is a part of the membrane‐bound O‐acyltransferase enzyme family (MBOAT). MBOATs are recognized by their most carried residues, histamine, and arginine, and are used in lipid synthesis, phospholipid remodeling, and peptide acylation. The enzyme itself is structured as a tetramer split into two homodimers. Nine transmembrane helices or domains (TMs) are on each monomer, the most notable being a cavity formed through TMs 4‐9 harboring histidine, an amino acid tied to catalysis and enzymatic activity.The ACAT1 protein‐coupled with its ACAT2 counterpart catalyze the storage of cholesteryl esters. ACAT2 differs from ACAT1 in function and structure. ACAT2 contains 2 domains and is mainly expressed in intestinal cells and controls lipoprotein particle secretion. The enzyme transfers an acyl group to 3β‐hydroxyl of cholesterol, a cholesterol‐binding site, to form cytosolic lipid droplets that are used for reducing membrane fluidity of the cell. Lipo‐protein‐induced ACAT in microphages is key to the creation of atheroma foam cells, cells characterized by the aggregation of these fatty plaque deposits on the walls of arteries, thus appearing like foam. The aggregation of this plaque to eventually narrow and harden arteries to obstruct blood flow is atherosclerosis. ACAT1 also maintains homeostasis during periods of cholesterol overload in the ER. Other functions of the protein include aiding lipoprotein synthesis and dietary LDL absorption.Recent studies have seen a pathogenic connection between cholesterol and Alzheimer’s disease or other neurodegenerative diseases. Cells contain a membrane that contains microdomains rich in cholesterol. The microdomains assist in the maintenance and growth of the cell and cell membrane; however, it is the disruption of these microdomains through individual disease‐causing toxins that cause them to inactivate ACAT1 and block cholesterol storage, disrupting brain activity.ACAT is currently being studied for its targeted drug therapy countering atherosclerosis. Current studies have proved ACAT inhibition to be ineffective, but there still remains potential immunotherapies to neurodegenerative disorders through the ACAT enzyme. Further research remains to be done to analyze the link between neurodegenerative disorders and the ACAT1 protein.

Acyl-Coenzyme A: Cholesterol Acyltransferase (ACAT) in Cholesterol Metabolism: From Its Discovery to Clinical Trials and the Genomics Era.

The purification and cloning of the acyl-coenzyme A: cholesterol acyltransferase (ACAT) enzymes and the sterol O-acyltransferase (SOAT) genes has opened new areas of interest in cholesterol metabolism given their profound effects on foam cell biology and intestinal lipid absorption. The generation of mouse models deficient in Soat1 or Soat2 confirmed the importance of their gene products on cholesterol esterification and lipoprotein physiology. Although these studies supported clinical trials which used non-selective ACAT inhibitors, these trials did not report benefits, and one showed an increased risk. Early genetic studies have implicated common variants in both genes with human traits, including lipoprotein levels, coronary artery disease, and Alzheimer’s disease; however, modern genome-wide association studies have not replicated these associations. In contrast, the common SOAT1 variants are most reproducibly associated with testosterone levels.

Acyl-Coenzyme A: Cholesterol Acyltransferase Inhibition in Cancer Treatment.

Overexpression of acyl-coenzyme A:cholesterol acyltransferase (ACAT) results in increased cholesteryl ester levels and has been involved in a variety of cancer types. As a consequence, cholesterol metabolism has raised interest as a potential target for cancer treatment. Inhibition of ACAT results in suppression of proliferation in a range of cancer cell types both in vitro and in vivo. The exact mechanism of this phenomenon is being investigated, and the most important findings are presented in this review.

Attenuation of Atherosclerosis by 3,4-Dihydroxy-Hydrocinnamic Acid in Rabbits by Partial Inhibition of ACAT

Polyphenols have been reported to have beneficial effects on cardiovascular disease. A polyphenolic compound, 3,4-dihydroxy-hydrocinnamic acid (3,4-DHHCA), has been shown to have antioxidative and antitumorigenic activities. However, the effect of 3,4-DHHCA on atherosclerosis is still unknown. Herein, we investigated the effects of 3,4-DHHCA on atherosclerosis in New Zealand White rabbits. Broad and fused fatty streak lesions were found in rabbits fed with high-cholesterol diet for 8 weeks. Administration of 3,4-DHHCA reduced atherosclerotic lesion formation and lesional accumulation of macrophage in rabbits fed with cholesterol diet without systemic or local toxicity. Hepatic acyl-coenzyme A: cholesterol acyltransferase (ACAT) activity was decreased after treatment with 3,4-DHHCA by 22% in cholesterol diet-fed rabbits compared with the control group. These results indicate that 3,4-DHHCA had antiatherogenic effects in rabbits, possibly by partial inhibition of ACAT.

Role of Acyl‐Coenzyme A:Cholesterol Acyltransferase 1 in Lipogenesis in Adipocytes

Dysregulated lipogenesis in adipose tissue contributes to obesity and its metabolic disorders. However, the molecular mechanisms regulating lipogenesis in adipocytes are not fully understood. Acyl‐coenzyme A: cholesterol acyltransferase (ACAT), an enzyme that catalyzes the formation of cholesteryl ester from free cholesterol (FC), has been identified as a therapeutic target to treat atherosclerosis. However, the role of ACAT in adipogenesis and lipogenesis in adipocytes is unknown. Here, we employed both pharmaceutical inhibitors and a lentivirus‐mediated shRNA‐based gene knockdown technique to investigate the role of ACAT in adipogenesis using murine 3T3‐L1 preadipocytes as an in vitro model of adipogenesis. Avasimibe, an ACAT inhibitor, was added at non‐toxic concentrations and replaced every other day during the course of adipogenesis. Avasimibe treatment inhibited lipid accumulation in a dose‐dependent manner, as judged by Oil red O staining and thin layer chromatography. Consistently, ACAT1 knockdown in 3T3‐L1 cells reduced lipid droplet accumulation by 40%. Real‐time PCR revealed that ACAT inhibition down‐regulated expressions of sterol regulatory element‐binding protein‐1 and genes involved in lipogenesis, with little effect on adipogenic transcription factors. These results suggest that ACAT is required for lipid droplet biogenesis in adipocytes. In addition, ACAT inhibition reduced 37% of intracellular FC and impaired cholesterol uptake in adipocytes, indicating a role of ACAT1 in the biochemical link between lipogenesis and cholesterol homeostasis. Taken together, our results reveal a novel function of ACAT1 in lipogenesis and highlight an important role of ACAT1 in integrating cholesterol metabolism and neutral lipid synthesis in adipocytes.

Cholesterol and lipoprotein metabolism: Early Career Committee contribution.

Cholesterol and lipoprotein metabolism: Early Career Committee contribution.

Palmitoylation of Amyloid Precursor Protein Regulates Amyloidogenic Processing in Lipid Rafts

Brains of patients affected by Alzheimer's disease (AD) contain large deposits of aggregated amyloid β-protein (Aβ). Only a small fraction of the amyloid precursor protein (APP) gives rise to Aβ. Here, we report that ∼10% of APP undergoes a post-translational lipid modification called palmitoylation. We identified the palmitoylation sites in APP at Cys¹⁸⁶ and Cys¹⁸⁷. Surprisingly, point mutations introduced into these cysteines caused nearly complete ER retention of APP. Thus, either APP palmitoylation or disulfide bridges involving these Cys residues appear to be required for ER exit of APP. In later compartments, palmitoylated APP (palAPP) was specifically enriched in lipid rafts. In vitro BACE1 cleavage assays using cell or mouse brain lipid rafts showed that APP palmitoylation enhanced BACE1-mediated processing of APP. Interestingly, we detected an age-dependent increase in endogenous mouse brain palAPP levels. Overexpression of selected DHHC palmitoyl acyltransferases increased palmitoylation of APP and doubled Aβ production, while two palmitoylation inhibitors reduced palAPP levels and APP processing. We have found previously that acyl-coenzyme A:cholesterol acyltransferase (ACAT) inhibition led to impaired APP processing. Here we demonstrate that pharmacological inhibition or genetic inactivation of ACAT decrease lipid raft palAPP levels by up to 76%, likely resulting in impaired APP processing. Together, our results indicate that APP palmitoylation enhances amyloidogenic processing by targeting APP to lipid rafts and enhancing its BACE1-mediated cleavage. Thus, inhibition of palAPP formation by ACAT or specific palmitoylation inhibitors would appear to be a valid strategy for prevention and/or treatment of AD.

Using self-organizing map (SOM) and support vector machine (SVM) for classification of selectivity of ACAT inhibitors

Using a self-organizing map (SOM) and support vector machine, two classification models were built to predict whether a compound is a selective inhibitor toward the two Acyl-coenzyme A: cholesterol acyltransferase (ACAT) isozymes, ACAT-1 and ACAT-2. A dataset of 97 ACAT inhibitors was collected. For each molecule, the global descriptors, 2D and 3D property autocorrelation descriptors and autocorrelation of surface properties were calculated from the program ADRIANA.Code. The prediction accuracies of the models (based on the training/ test set splitting by SOM method) for the test sets are 88.9% for SOM1, 92.6% for SVM1 model. In addition, the extended connectivity fingerprints (ECFP_4) for all the molecules were calculated and the structure-activity relationship of selective ACAT inhibitors was summarized, which may help find important structural features of inhibitors relating to the selectivity of ACAT isozymes.

Lipids-induced Apoptosis Is Aggravated by Acyl-coenzyme A: Cholesterol Acyltransferase Inhibitor

To investigate the role of acyl-coenzyme A: cholesterol acyltransferase inhibitor (ACATI) in apoptosis induced by lipids and whether lipids-induced apoptosis is accompanied by increase of free cholesterol in endoplasmic reticulum (ER), in order to further understand the mechanism of lipids-induced apoptosis in advanced atherosclerosis. Human vascular smooth muscle cells (VSMCs) and phorbol 12-myristate 13-acetate (PMA) differentiated THP-1 macrophages were used. Tritiated thymidine incorporation was applied to detect cell proliferation. Cytotoxicity was assessed by lactate dehydrogenase (LDH) release. 4',6-diamidino-2-phenylindole (DAPI) staining, caspase-3, -7 assay, and Annexin-V/propidium iodide (PI) staining were used to detect apoptosis. High performance liquid chromatography was used in intracellular free cholesterol and cholesterol ester assay. ER free cholesterol was quantified. Different lipids had different effects on proliferation and cytotoxicity of VSMCs. 25-hydroxycholesterol (25OHC) had biphasic effects on the proliferation of VSMCs. At low concentration, it stimulated cell proliferation, but turned to proliferation inhibition as concentration reached 15 mug/mL. 25OHC and acetylated low density lipoprotein (AcLDL) could respectively induce apoptosis in human VSMCs and PMA differentiated THP-1 macrophages, which was aggravated by ACATI, accompanied by increase of intracellular free cholesterol content. There was also an increase of cholesterol content in ER with AcLDL-induced apoptosis in THP-1 macrophages. Lipids could induce apoptosis, accompanied by increase of intracellular free cholesterol content, which could be augmented by ACATI, suggesting that insults resulting in ER free cholesterol rise might be the initiator of apoptosis.

AM-251 and SR144528 are acyl CoA:cholesterol acyltransferase inhibitors

Oxysterol-induced macrophage apoptosis may have a role in atherosclerosis. Macrophages lacking the type 2 cannabinoid receptor (CB2) are partially resistant to apoptosis induced by 7-ketocholesterol (7KC). AM-251 and SR144528 are selective antagonists of CB1 and CB2 receptors, respectively. We observed that both compounds reduce 7KC-induced apoptosis in Raw 264.7 macrophages. As oxysterol-induced macrophage apoptosis requires acyl-coenzymeA:cholesterol acyltransferase (ACAT) activity, we tested their affects on ACAT activity. AM-251 and SR144528 both reduced cholesteryl ester synthesis in unstimulated and acetylated LDL-stimulated Raw 264.7 macrophages, CB2 +/+ and CB2 −/− peritoneal macrophages, as well as in vitro, in mouse liver microsomes. Consistent with inhibition of ACAT, the development of foam cell characteristics in macrophages by treatment with acetylated LDL was reduced by both compounds. This work is the first evidence that AM-251 and SR144528 are inhibitors of ACAT and as a result, might have anti-atherosclerotic activities independent of their affect on cannabinoid signaling.

Bavachin and isobavachalcone, acyl-coenzyme A: Cholesterol acyltransferase inhibitors from Psoralea corylifolia

Acyl-coenzyme A: cholesterol acyltransferase (ACAT) catalyzes cholesterol esterification and plays important roles in intestinal absorption of cholesterol, hepatic production of lipoproteins and accumulation of cholesteryl ester within macrophages and smooth muscle cells. Ethanol extract of Psoralea corylifolia showed a significant inhibition of ACAT enzyme. Via bioactivity-guided fractionation of the ethanol extract of Psoralea corylifolia, two prenylated flavonoids were isolated. Their structures were determined as bavachin (1) and isobavachalcone (2) by spectroscopic analysis ((1)H-, (13)C-NMR, 2DNMR, and ESI-MS). The IC(50) values were 86.0 (1) and 48.0 (2) microM in the ACAT assay system using rat liver microsome. Compound 2 also decreased cholesteryl ester formations in HepG2 cells. In addition, this compound showed a noncompetitive type of inhibition of ACAT.

Do Prion-Induced Changes in Membrane Cholesterol Trigger Neurodegeneration?

The transmissible spongiform encephalopathies (TSEs) are associated with the conversion of a host-encoded cellular protein (PrPC) to alternatively folded, disease-associated isoforms (PrPSc) [1]. These diseases are unique in that the only identified component of the infectious agent is a self-replicating protein (PrPSc); which explains why such diseases are commonly called prion diseases (protein-only infectious agent). The accumulation of PrPSc is closely associated with the main pathological features of prion diseases; loss of synapses, activation of glial cells, neuronal loss and the spongiform degeneration of the brain [2]. However, the cellular and molecular mechanisms leading to neurodegeneration in these diseases are imperfectly understood. In this review, we outline developments implicating prion-induced changes in the levels of membrane cholesterol as a trigger of abnormal cell signaling, which leads to neurodegeneration. Recent studies have demonstrated that prion infection has a significant impact on the biochemistry of neuronal cell membranes; it significantly increased the amounts of free cholesterol, but reduced the amounts of esterified cholesterol [3]. In addition, a highly significant correlation between the amounts of PrPSc and the concentration of free cholesterol in cell membranes was observed. Crucially, the effects of prion infection were not replicated by stimulating cholesterol biosynthesis. Stimulation of cholesterol production in uninfected neurons had the opposite effect to prion infection; it increased cholesterol ester formation without altering the amounts of free cholesterol. Such observations indicate that prion infection increased the capacity of cell membranes to solubilize free cholesterol. Cellular cholesterol is found as either free cholesterol in membranes, or as cholesterol esters that are stored in cytoplasmic droplets. A dynamic equilibrium between the pools of free cholesterol and cholesterol esters is tightly controlled by acyl-coenzyme A:cholesterol acyltransferase (ACAT), an endoplasmic reticulum (ER)-resident enzyme that esterifies free cholesterol with long-chain fatty acids [4]. Therefore, the combination of increased free cholesterol and reduced cholesterol esters observed in prion-infected cells may result from inhibition of ACAT, or from the sequestration of free cholesterol in microenvironments that avoid ACAT.

Cholesterol esterification reduces the neurotoxicity of prions

The transmissible spongiform encephalopathies develop following the conversion of a host-encoded protein (PrP C) into abnormally folded, disease-related isoforms (PrP Sc). Here we report that three acyl-coenzyme A:cholesterol acyltransferase (ACAT) inhibitors, TMP-153, FR179254 or YIC-C8-434, were more toxic to prion-infected neuronal cell lines (ScGT1 and ScN2a cells) than to their uninfected equivalents (GT1 and N2a cells). The toxicity of ACAT inhibitors for ScGT1 cells was not reversed by the addition of cholesterol esters, rather it was increased by the addition of free cholesterol indicating that the toxicity of ACAT inhibitors was related to the increased free cholesterol content of cells rather than reduced amounts of cholesterol esters. This hypothesis was strengthened by the observation that the addition of free cholesterol killed ScGT1, but not GT1 cells. Treatment with ACAT inhibitors increased caspase-3 activity and prostaglandin E 2 production in ScGT1 cells but not in GT1 cells. The addition of the phospholipase A 2 (PLA 2) inhibitors (AACOCF 3 or MAFP) reduced prostaglandin E 2 production and protected ScGT1 cells against the toxicity of ACAT inhibitors. These results indicate that cholesterol esterification is an important cellular response that reduces PrP Sc-induced activation of PLA 2 and protects against cell death in ScGT1 cells.

Inhibition of acyl-coenzyme A:cholesterol acyltransferase stimulates cholesterol efflux from macrophages and stimulates farnesoid X receptor in hepatocytes

We investigated the mechanism of spontaneous cholesterol efflux induced by acyl-coenzyme A:cholesterol acyltransferase (ACAT) inhibition, and how an alteration of cholesterol metabolism in macrophages impacts on that in HepG2 cells. Oleic acid anilide (OAA), a known ACAT inhibitor reduced lipid storage substantially by promotion of cholesterol catabolism and repression of cholesteryl ester accumulation without further increase of cytotoxicity in acetylated low-density lipoprotein-loaded THP-1 macrophages. Analysis of expressed mRNA and protein revealed that cholesterol 7alpha-hydroxylase (CYP7A1), oxysterol 7alpha- hydroxylase (CYP7B1), and cholesterol 27-hydroxylase (CYP27) were highly induced by ACAT inhibition. The presence of a functional cytochrome P450 pathway was confirmed by quantification of the biliary cholesterol mass in cell monolayers and extracelluar medium. Notably, massively secreted biliary cholesterol from macrophages suppressed the expression of CYP7 proteins in a farnesoid X receptor (FXR)-dependent manner in HepG2 cells. The findings reported here provide new insight into mechanisms of spontaneous cholesterol efflux, and suggest that ACAT inhibition may stimulate cholesterol-catabolic (cytochrome P450) pathway in lesion-macrophages, in contrast, suppress it in hepatocyte via FXR induced by biliary cholesterol (BC).

Sitosterol-containing Lipoproteins Trigger Free Sterol-induced Caspase-independent Death in ACAT-competent Macrophages

Sitosterolemia is a disease characterized by very high levels of sitosterol and other plant sterols and premature atherothrombotic vascular disease. One theory holds that plant sterols can directly promote atherosclerosis, but the mechanism is not known. Unesterified, or "free," cholesterol (FC) is a potent inducer of macrophage death, which causes plaque necrosis, a precursor to atherothrombosis. FC-induced macrophage death, however, requires dysfunction of the sterol esterifying enzyme acyl-coenzyme A-cholesterol acyltransferase (ACAT), which likely occurs slowly during lesion progression. In contrast, plant sterols are relatively poorly esterified by ACAT, and so they may cause macrophage death and plaque necrosis in an accelerated manner. In support of this hypothesis, we show here that macrophages incubated with sitosterol-containing lipoproteins accumulate free sterols and undergo death in the absence of an ACAT inhibitor. As with FC loading, sitosterol-induced macrophage death requires sterol trafficking to the endoplasmic reticulum, and sitosterol-enriched endoplasmic reticulum membranes show evidence of membrane protein dysfunction. However, whereas FC induces caspase-dependent apoptosis through activation of the unfolded protein response and JNK, sitosterol-induced death is caspase-independent and involves neither the unfolded protein response nor JNK. Rather, cell death shows signs of necroptosis and autophagy and is suppressed by inhibitors of both processes. These data establish two new concepts. First, a relatively subtle change in sterol structure fundamentally alters the type of death program triggered in macrophages. Understanding the basis of this alteration should provide new insights into the molecular basis of death pathway signaling. Second, sitosterol-induced macrophage death does not require ACAT dysfunction and so may occur in an accelerated fashion. Pending future in vivo studies, this concept may provide at least one mechanism for accelerated plaque necrosis and atherothrombotic disease in patients with sitosterolemia.

Glabrol, an acyl-coenzyme A: Cholesterol acyltransferase inhibitor from licorice roots

Acyl-coenzyme A: cholesterol acyltransferase (ACAT) esterifies free cholesterol in the liver and the intestine. It has relations with production of lipoproteins and accumulation of cholesteryl esters of the atheroma. Therefore, ACAT inhibitors may act as antihypercholesterolemic and antiatherosclerotic agents. One isoprenyl flavonoid was isolated from ethanol extract of licorice roots. On the basis of spectral evidences, the compound was identified as glabrol ( 1). Compound 1 inhibited rat liver microsomal ACAT activity with an IC 50 value of 24.6 μM and decreased cholesteryl ester formation with an IC 50 value of 26.0 μM in HepG2 cells. In addition, 1 showed a non-competitive type of inhibition against ACAT.

Intravascular Ultrasound in Cardiovascular Medicine

Case presentation: A 38-year-old man underwent angiography 2 weeks after a non–ST-segment myocardial infarction. He was overweight and prehypertensive, with a blood pressure of 134/86 mm Hg. His biochemical parameters included low-density lipoprotein cholesterol (LDL-C) 127 mg/dL, high-density lipoprotein cholesterol (HDL-C) 38 mg/dL, and C-reactive protein (CRP) 3.2 mg/L. Angiography revealed mild disease throughout the coronary arteries and a hazy appearance that involved the proximal left anterior descending coronary artery. Intravascular ultrasound (IVUS) imaging revealed diffuse and extensive atherosclerosis with evidence of plaque rupture at multiple sites (Figure 1). In the presence of no significant luminal stenoses, the patient was treated medically with aspirin and low-dose statin therapy. Figure 1. Illustrative example of coronary angiography of the left anterior descending artery that appears to be free of irregularities, apart from a hazy appearance in the proximal segment. A representative cross-sectional tomographic image obtained by IVUS examination (top right) within the proximal segment (white arrow on angiogram) revealed substantial atheroma within the arterial wall. The bottom right segment illustrates a tomographic image of atheroma containing significant ulceration (5 o’clock) at the site of the gold arrow on the angiogram. The application of IVUS in this case highlights a number of important points with regard to the natural history of atherosclerosis and its modification by use of established medical therapies. The ability to image the entire arterial wall represents a significant advantage over coronary angiography. In the presence of minimal luminal stenoses, IVUS imaging in patients with coronary symptoms typically reveals extensive and diffuse atherosclerosis.1 Application of serial IVUS imaging in prospective clinical trials has enabled a greater understanding of the impact of antiatherosclerotic interventions on patients with established coronary artery disease (CAD). Ultrasonic coronary imaging has revealed essential details about the natural history of atherosclerosis. Atheroma formation begins at a …

Genetics and molecular biology: macrophage ACAT depletion – mechanisms of atherogenesis

Genetics and molecular biology: macrophage ACAT depletion – mechanisms of atherogenesis

Inhibitors of Acyl-Coenzyme A: Cholesterol Acyltransferase

Acyl-coenzyme A: cholesterol acyltransferase (ACAT) is an intracellular enzyme that catalyzes the formation of cholesterol esters from cholesterol and fatty acyl-coenzyme A. Animal experiments showed that ACAT inhibitors reduce plasma cholesterol levels by suppressing absorption of dietary cholesterol and by suppressing the assembly and secretion of apolipoprotein B-containing lipoproteins such as very low density lipoprotein in liver and chylomicron in intestine. Moreover, ACAT inhibitors were shown to prevent formation of macrophage-derived foam cells in the arterial walls. However, a recent double-blind, placebo-controlled, randomized trial of a potent ACAT inhibitor, avasimibe, failed to show significant beneficial effects on coronary atherosclerosis assessed by intravascular ultrasound. For clinical application of ACAT inhibitors, development of more potent compounds and improvements of the methods to evaluate their clinical efficacy are strongly needed.