COMPLETE INHIBITION OF REGENERATION IN MICE FOLLOWING SIMULTANEOUS ELIMINATION OF MET‐EGFR SIGNALING

Abstract

The Liver has an amazing ability to regenerate following loss of mass. Regeneration is a complex process that involves integration of multiple growth signaling pathways. Liver regeneration has been experimentally studied by surgical removal of 2/3 rd of liver in rodents. Following a partial hepatectomy (PHx), more than 100 genes exhibit differential expression and multiple signaling pathways are activated. Among the growth factors that are associated with hepatocyte proliferation, only Hepatocyte growth factor (HGF) and Epidermal growth factor (EGF) are complete mitogens that stimulate DNA synthesis in hepatocyte cultures maintained in chemically defined media. They are also the only ones that stimulate DNA synthesis in the liver of normal mice and rats. EGFR and Met phosphorylation increases over basal levels by 60 minutes after a PHx. Cross talk between these two pathways has been documented and integration of these two pathways may be required for regeneration to proceed efficiently. We decided to study the effect on regeneration in mice following complete elimination of EGFR‐MET signaling pathway. To this end, a post natal elimination of c‐met by using ER‐Cre‐Tamoxiphen‐inducible system was utilized. Mice were injected with Tamoxiphen (1mg/ml) and injected i.p everyday for 5 days. Controls were injected with corn oil alone. This treatment deleted exon 16 that contains a critical ATP‐binding site in the intracellular tyrosine kinase (TK) domain, essential for the activation of c‐met. PhosphoEGFR was inhibited by using Canertinib, an irreversible pan‐pEGFR inhibitor that was administered i.p at 80 mg/kg. Mice were injected a day before PHx and then everyday for 7 days. Treatment of Met‐KO mice with Canertinib inhibited EGFR signaling significantly. Treatment resulted in global changes in signaling pathways that seemed to promote block in proliferation and liver damage at time 0 Hrs. post PHx. A significant reduction in Ki67 labeling was also evident, as was a reduction in liver to body weight ratio. The size of the lobules and the hepatocytes was decreased in Met KO mice treated with Canertinib. Met KO mice treated with Canertinib had increased morbidity and had elevated AST, ALT values, and lower albumin levels. Combined elimination of both MET and EGFR signaling resulted in sustained inhibition of liver regeneration up to 14 days suggesting a liver regeneration deficit. The AKT‐mTOR and the ERK pathways associated with cellular proliferation were down regulated in MET‐EGFR inhibited mice. Suppression of MET‐EGFR signaling caused profound alterations in expression of many cell cycle associated genes and genes associated with fatty acid biosynthesis pathway. There appears to be is no redundancy and compensatory scheme for liver regeneration following complete elimination of EGFR‐MET signaling network.Support or Funding InformationNIH Grant No. CA 35373, CA 103958 & Rangos Fund for Enhancement of Research in Pathology.