Publications
 
Method for the quantitative assay of fatty acid–bile acid conjugates by tandem mass spectrometry

Fatty acid–bile acid conjugates and especially arachidyl amido cholic acid are synthetic molecules that were shown to be effective in Gallstone disease,in Fatty Liver and Hypercholesterolemia.. To measure these novel compounds a liquid chromatography electrospray tandem mass spectrometry method was developed based on the analysis of 100 μl of plasma with stearyl amido cholic acid (stamchol, 1.5 μmol/l) added as internal standard. Repeatable calibrations between 0 and 50 μmol/l exhibited consistent linearity and reproducibility.Inter-and intraassay C.V.s were 5.3–11.4% and 2.6–6.4%, respectively, at targeted concentrations of 0.1, 2.3 and 50 μmol/l.
 
 
Faecal sterol output is increased by arachidyl amido cholanoic acid (Aramchol) in rats

Fatty acid –bile acid conjugates (FABACs) were shown recently to have important and multiple effects on cholesterol metabolism. In human fibroblasts, they were found to markedly enhance cholesterol efflux by an ATP-binding cassette transporter A1-dependent pathway. In C57L/J mice, they increased CYP7A1 activity and RNA expression, while decreasing moderately 3-hydroxy-3-methylglutaryl-CoA reductase activity. In C57L/J mice and in hamsters, they also decreased serum cholesterol levels, whereas in other animals, this effect was not seen in short-term experiments. In the present study, we investigated potential mechanisms of action of arachidyl amido cholanoic acid (Aramchol), with particular reference to biliary and faecal sterol outputs in rats. Supplementation with Aramchol at a dose of 150 mg ·kg −1 ·day −1 increased neutral sterol output by approx. 50%, while the faecal outputs of bile salts and total sterols increased by almost 2-fold. Biliary lipid outputs were not signi ficantly different between the control and FABAC-supplemented animals. These findings indicate an overall catabolic effect of FABACs on body cholesterol.
 

ABCA1-dependent but apoA-I-independent cholesterol efflux mediated by fatty acid–bile acid conjugates (FABACs)

FABACs (fatty acid–bile acid conjugates) are synthetic molecules that are designed to treat a range of lipid disorders. The compounds prevent cholesterol gallstone formation and diet-induced fatty liver, and increase reverse cholesterol transport in rodents. The aim of the present study was to investigate the effect of FABACs on cholesterol efflux in human cells. Aramchol (3β-arachidylamido-7α, 12α ,5β -cholan-24-oic acid) increased cholesterol efflux from human skin fibroblasts in a dose-dependent manner in the absence of known efflux mediators such as apoA-I (apolipoprotein A-I), but had little effect on phospholipid efflux.An LXR (liver X receptor) agonist strongly increased Aramchol-induced cholesterol efflux; however, in ABCA1 (ATP- binding cassette transporter A1)-deficient cells from Tangier disease patients, the Aramchol effect was absent, indicating that activity of ABCA1 was required.Aramchol did not affect ABCA1 expression, but plasma membrane levels of the transporter increased 2-fold. Aramchol is the first small molecule that induces ABCA1-dependent cholesterol efflux without affecting transcriptional control. These findings may explain the beneficial effect of the compound on atherosclerosis. Key words: ATP-binding cassette transporter A1 (ABCA1), Aramchol, atherosclerosis, cholesterol efflux, fatty acid–bile acid conjugate (FABAC), reverse cholesterol transport.


Fatty Acid Bile Acid Conjugates Inhibit Atherosclerosis in the C57BL6 Mouse Model

Objective: The aim of the current research was to study whether fatty acid bile acid conjugates (FABACs) have a beneficial effect on atherosclerosis progression and blood lipid levels in mice.
Methods: C57BL/6 female mice, fed a high-fat Paigen diet, were administered an oral dose of FABAC or DDH2O daily. Quantification of atherosclerotic fatty-streak lesions at the aortic sinus was performed.
Results: The FABAC-treated mice showed a significant reduction in the atherosclerotic lesion areas as compared to the control group (p = 0.019). A significant elevation in total cholesterol levels was observed in both the FABAC and control groups. Higher FABAC levels were measured in the high-density lipoprotein fraction as compared to the very-low-density and low-density lipoprotein fractions.
Conclusion: Our findings demonstrate that FABACs, given orally, reduce the development of atherosclerosis in mice fed a high-fat high-cholesterol diet, despite a lack of effect on plasma lipid levels.( a marked reduction, ca 50%, in serum cholesterol was subsequently
demonstrated in numerous other experiments--ed.) 


Fatty acid bile acid conjugates (FABACs)—New molecules for the prevention of cholesterol crystallisation in bile

Background—Cholesterol gallstones are a frequent disease for which at present surgery is the usual therapy.Despite the importance of bile acids it has become evident that phospholipids are the main cholesterol solubilisers in bile.Even phospholipid components,such as fatty acids, have anticrystallising activity.
Aim—To synthesise fatty acid bile acid conjugates (FABACs)and study their effects on cholesterol crystallisation in bile in vitro and in vivo.
Methods—FABACs were prepared by conjugation of cholic acid at position 3 with saturated fatty acids of variable chain length using an amide bond.Cholesterol crystallisation and its kinetics (crystal observation time, crystal mass) were studied in model bile,pooled enriched human bile,and fresh human bile using FABACs with saturated fatty acids of varying chain length (C-6 to C-22). Absorption of FABACs into blood and bile was tested in hamsters.Prevention of biliary cholesterol crystallisation in vivo was tested in hamsters and inbred mice.
Results—FABACs strongly inhibited cholesterol crystallisation in model as well as native bile.The FABACs with longer acyl chains (C-16 to C-22) were more effective. At a concentration of 5 mM,FABACs almost completely inhibited cholesterol crystallisation in fresh human bile for 21 days. FABACs were absorbed and found in both portal and heart blood of hamsters. Levels in bile were 2–3 times higher than in blood, indicating active secretion. Appreciable levels were found in the systemic circulation 24–48 hours after a single administration.Ingested FABACs completely prevented the formation of cholesterol crystals in the gall bladders of hamsters and mice fed a lithogenic diet.
Conclusions—FABACs are potent inhibitors of cholesterol crystallisation in bile. They are absorbed and secreted into bile and prevent the earliest step of cholesterol gallstone formation in animals. These compounds may be of potential use in cholesterol gall stone disease in humans.
 
 
 
 
 
Gallstones, mostly cholesterol stones, affect some 15% of the population.Oral bile salts dissolve human cholesterol gallstones,but with low efficacy, and surgery remains the main therapeutic option. Fatty acid bile acid conjugates (FABACs) were shown to prevent formation of cholesterol gallstones in experimental animals.The aim of this study was to test whether these compounds could dissolve preexisting cholesterol gallstones via oral administration. Inbred, gallstone-susceptible C57J/L mice were given a lithogenic diet for 2 months, and the presence of gallstones was ascertained. The mice were then switched to a regular diet while part of them were given in addition C20-FABAC, by gavage, at a dose of 0.5 or 3 mg per animal per day. All mice tested had cholesterol gallstones after 2 months on the lithogenic diet. In study I, after 2 months on the regular diet, 3 of 4 (75%) of the controls had gallstones, whereas none of the 6 FABAC-fed animals (3 mg/d) had stones (P = .033). In study II, evaluating 2 FABAC doses, after 2 months on the regular diet, 8 of 8 (100%) of the controls had gallstones, which were found in 2 of 7 (28%) and 1 of 8 (12%) of the mice supplemented with 0.5 mg/d (P = .007) or 3 mg/d (P = .001) FABAC, respectively. On a molar basis, the dose of 0.5 mg FABAC is equivalent to 14 mg/kg/d of a bile acid. In conclusion, FABACs given orally can dissolve preexisting cholesterol gallstones in mice. This was accomplished with a dose of FABAC equivalent to the dose of bile acids used in human gallstone dissolution.
 
 
  
Fatty acid bile acid conjugates (FABACs) are a new family of synthetic molecules designed to solubilize biliary cholesterol. They were shown to prevent and dissolve cholesterol gallstones in inbred C57L/J mice fed a lithogenic, high-fat diet (HFD). In these mice, fatty liver was observed in the controls but not in the FABAC-treated ones. The present study was designed to study the effect of FABAC (arachidyl-amido-cholanoic acid) on diet-induced fatty liver in rats, hamsters, and mice. The fatty liver score (on a scale of 0-4 by light microscopy) was 4.0 in control hamsters and 0.3 in the FABAC-fed hamsters (P <.001). In mice it was 1.5 and 0.4, respectively (P <.01). The lipid/protein ratio in the liver was 1.3 ± 0.44 (mg lipid/mg protein) in control rats and 0.66 ± 0.04 in the FABAC group (P = .001) after 14 days. In hamsters it was 1.41 ± 0.27 and 1.11 ± 0.20, respectively (P = .03), after 21 days. In Imperial Charles River (ICR) mice the ratio was 0.34 ± 0.10 and 0.17 ± 0.07 (P = .03), respectively, after 24 days. Liver fat concentration, measured as mg lipid/g liver tissue, decreased similarly by FABAC feeding. The decrease in liver fat affected mainly the triglyceride levels. FABAC-fed animals gained weight similarly to the controls. Triglyceride absorption was unaffected by FABAC supplementation. In conclusion, oral FABAC therapy prevents/reduces the development of fatty liver in animals consuming a HFD. 
                                                               
 
 
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