Arachidyl amido cholanoic acid improves liver glucose and lipid homeostasis in nonalcoholic steatohepatitis via AMPK and mTOR regulation

Abstract

BACKGROUND

Arachidyl amido cholanoic acid (Aramchol) is a potent downregulator of hepatic stearoyl-CoA desaturase 1 (SCD1) protein expression that reduces liver triglycerides and fibrosis in animal models of steatohepatitis. In a phase IIb clinical trial in patients with nonalcoholic steatohepatitis (NASH), 52 wk of treatment with Aramchol reduced blood levels of glycated hemoglobin A1c, an indicator of glycemic control.

AIM

To assess lipid and glucose metabolism in mouse hepatocytes and in a NASH mouse model [induced with a 0.1% methionine and choline deficient diet (0.1MCD)] after treatment with Aramchol.

METHODS

Isolated primary mouse hepatocytes were incubated with 20 μmol/L Aramchol or vehicle for 48 h. Subsequently, analyses were performed including Western blot, proteomics by mass spectrometry, and fluxomic analysis with ¹³C-uniformly labeled glucose. For the in vivo part of the study, male C57BL/6J mice were randomly fed a control or 0.1MCD for 4 wk and received 1 or 5 mg/kg/d Aramchol or vehicle by intragastric gavage for the last 2 wk. Liver metabolomics were assessed using ultra-high-performance liquid chromatography-time of flight-MS for the determination of glucose metabolism-related metabolites.

RESULTS

Combination of proteomics and Western blot analyses showed increased AMPK activity while the activity of nutrient sensor mTORC1 was decreased by Aramchol in hepatocytes. This translated into changes in the content of their downstream targets including proteins involved in fatty acid (FA) synthesis and oxidation [P-ACCα/β(S79), SCD1, CPT1A/B, HADHA, and HADHB], oxidative phosphorylation (NDUFA9, NDUFB11, NDUFS1, NDUFV1, ETFDH, and UQCRC2), tricarboxylic acid (TCA) cycle (MDH2, SUCLA2, and SUCLG2), and ribosome (P-p70S6K[T389] and P-S6[S235/S236]). Flux experiments with ¹³C-uniformely labeled glucose showed that TCA cycle cataplerosis was reduced by Aramchol in hepatocytes, as indicated by the increase in the number of rounds that malate remained in the TCA cycle. Finally, liver metabolomic analysis showed that glucose homeostasis was improved by Aramchol in 0.1MCD fed mice in a dose-dependent manner, showing normalization of glucose, G6P, F6P, UDP-glucose, and Rbl5P/Xyl5P.

CONCLUSION

Aramchol exerts its effect on glucose and lipid metabolism in NASH through activation of AMPK and inhibition of mTORC1, which in turn activate FA β-oxidation and oxidative phosphorylation.

Keywords: Nonalcoholic fatty liver disease, Steatohepatitis, Methionine and choline deficient diet, Tricarboxylic acid cycle, Hemoglobin A1c, Stearoyl-CoA desaturase 1

Core Tip: Arachidyl amido cholanoic acid (Aramchol), a phase III investigational drug for the treatment of nonalcoholic steatohepatitis (NASH), has been demonstrated to be able to reduce liver steatosis, inflammation, and fibrosis in various animal models of steatohepatitis. In a phase IIb clinical trial, it reduced blood levels of glycated hemoglobin A1c, an indicator of glycemic control. In this study, we showed that Aramchol-treated hepatocytes had activated AMPK and inhibited mTORC1, which in turn activated fatty acid β-oxidation and oxidative phosphorylation, inhibiting de novo lipogenesis, gluconeogenesis, and cataplerosis. These results explain the mechanism by which Aramchol exerts its effect on glucose and lipid metabolism in NASH.