Epigenetic mechanisms and metabolic reprogramming in fibrogenesis: dual targeting of G9a and DNMT1 for the inhibition of liver fibrosis

Gut. 2021 Feb;70(2):388-400. doi: 10.1136/gutjnl-2019-320205. Epub 2020 Apr 23.

Abstract

Objective: Hepatic stellate cells (HSC) transdifferentiation into myofibroblasts is central to fibrogenesis. Epigenetic mechanisms, including histone and DNA methylation, play a key role in this process. Concerted action between histone and DNA-mehyltransferases like G9a and DNMT1 is a common theme in gene expression regulation. We aimed to study the efficacy of CM272, a first-in-class dual and reversible G9a/DNMT1 inhibitor, in halting fibrogenesis.

Design: G9a and DNMT1 were analysed in cirrhotic human livers, mouse models of liver fibrosis and cultured mouse HSC. G9a and DNMT1 expression was knocked down or inhibited with CM272 in human HSC (hHSC), and transcriptomic responses to transforming growth factor-β1 (TGFβ1) were examined. Glycolytic metabolism and mitochondrial function were analysed with Seahorse-XF technology. Gene expression regulation was analysed by chromatin immunoprecipitation and methylation-specific PCR. Antifibrogenic activity and safety of CM272 were studied in mouse chronic CCl4 administration and bile duct ligation (BDL), and in human precision-cut liver slices (PCLSs) in a new bioreactor technology.

Results: G9a and DNMT1 were detected in stromal cells in areas of active fibrosis in human and mouse livers. G9a and DNMT1 expression was induced during mouse HSC activation, and TGFβ1 triggered their chromatin recruitment in hHSC. G9a/DNMT1 knockdown and CM272 inhibited TGFβ1 fibrogenic responses in hHSC. TGFβ1-mediated profibrogenic metabolic reprogramming was abrogated by CM272, which restored gluconeogenic gene expression and mitochondrial function through on-target epigenetic effects. CM272 inhibited fibrogenesis in mice and PCLSs without toxicity.

Conclusions: Dual G9a/DNMT1 inhibition by compounds like CM272 may be a novel therapeutic strategy for treating liver fibrosis.

Keywords: fibrogenesis; gene regulation; glucose metabolism.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chromatin Immunoprecipitation
  • DNA (Cytosine-5-)-Methyltransferase 1 / genetics
  • DNA (Cytosine-5-)-Methyltransferase 1 / metabolism*
  • Epigenesis, Genetic
  • Gene Expression Regulation
  • Gene Knockdown Techniques
  • Hepatic Stellate Cells / metabolism*
  • Histocompatibility Antigens / genetics
  • Histocompatibility Antigens / metabolism*
  • Histone-Lysine N-Methyltransferase / genetics
  • Histone-Lysine N-Methyltransferase / metabolism*
  • Humans
  • Liver Cirrhosis / etiology*
  • Liver Cirrhosis / genetics
  • Liver Cirrhosis / metabolism
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Polymerase Chain Reaction
  • Transforming Growth Factor beta1 / metabolism

Substances

  • Histocompatibility Antigens
  • Transforming Growth Factor beta1
  • DNA (Cytosine-5-)-Methyltransferase 1
  • DNMT1 protein, human
  • EHMT2 protein, human
  • Histone-Lysine N-Methyltransferase