Amplification and/or activation of the c-Myc protooncogene is one of the leading genetic events along hepatocarcinogenesis. The oncogenic potential of c-Myc has been proven experimentally by the finding that its overexpression in the mouse liver triggers tumor formation. However, the molecular mechanism whereby c-Myc exerts its oncogenic activity in the liver remains poorly understood. Here, we demonstrate that the mammalian target of rapamycin complex 1 (mTORC1) cascade is activated and necessary for c-Myc dependent hepatocarcinogenesis. Specifically, we found that ablation of Raptor, the unique member of the mTORC1 complex, strongly inhibits c-Myc liver tumor formation. Also, p70S6K/ribosomal protein S6 (RPS6) and eukaryotic translation initiation factor 4E-binding protein 1/eukaryotic translation initiation factor 4E (4EBP1/eIF4E) signaling cascades downstream of mTORC1 are required for c-Myc-driven tumorigenesis. Intriguingly, microarray expression analysis revealed the upregulation of multiple amino acid transporters, including SLC1A5 and SLC7A6, leading to robust uptake of amino acids, including glutamine, into c-Myc tumor cells. Subsequent functional studies showed that amino acids are critical for activation of mTORC1, as their inhibition suppressed mTORC1 in c-Myc tumor cells. In human HCC specimens, levels of c-Myc directly correlate with those of mTORC1 activation as well as of SLC1A5 and SLC7A6. Conclusion Our current study indicates that an intact mTORC1 axis is required for c-Myc-driven hepatocarcinogenesis. Thus, targeting mTOR pathway or amino acid transporters may be an effective and novel therapeutic option for the treatment of HCC with activated c-Myc signaling.

Activation of the AKT/mTOR cascade and overexpression of c-Met have been implicated in the development of human hepatocellular carcinoma (HCC). To elucidate the functional crosstalk between the two pathways, we generated a model characterized by the combined expression of activated AKT and c-Met in the mouse liver. Co-expression of AKT and c-Met triggered rapid liver tumor development and mice required to be euthanized within 8 weeks after hydrodynamic injection. At the molecular level, liver tumors induced by AKT/c-Met display activation of AKT/mTOR and Ras/MAPK cascades as well as increased lipogenesis and glycolysis. Since a remarkable lipogenic phenotype characterizes liver lesions from AKT/c-Met mice, we determined the requirement of lipogenesis in AKT/c-Met driven hepatocarcinogenesis using conditional Fatty Acid Synthase (FASN) knockout mice. Of note, hepatocarcinogenesis induced by AKT/c-Met was fully inhibited by FASN ablation. In human HCC samples, coordinated expression of FASN, activated AKT, and c-Met proteins was detected in a subgroup of biologically aggressive tumors. Altogether, our study demonstrates that co-activation of AKT and c-Met induces HCC development that depends on the mTORC1/FASN pathway. Suppression of mTORC1 and/or FASN might be highly detrimental for the growth of human HCC subsets characterized by concomitant induction of the AKT and c-Met cascades.

Background & Aims Cumulating evidence underlines the crucial role of aberrant lipogenesis in human hepatocellular carcinoma (HCC). Here, we investigated the oncogenic potential of fatty acid synthase (FASN), the master regulator of de novo lipogenesis, in the mouse liver. Methods FASN was overexpressed in the mouse liver, either alone or in combination with activated N-Ras, c-Met, or SCD1, via hydrodynamic injection. Activated AKT was overexpressed via hydrodynamic injection in livers of conditional FASN or Rictor knockout mice. FASN was suppressed in human hepatoma cell lines via specific small interfering RNA. Results Overexpression of FASN, either alone or in combination with other genes associated with hepatocarcinogenesis, did not induce histological liver alterations. In contrast, genetic ablation of FASN resulted in the complete inhibition of hepatocarcinogenesis in AKT-overexpressing mice. In human HCC cell lines, FASN inactivation led to a decline in cell proliferation and a rise in apoptosis, which were paralleled by a decrease in the levels of phosphorylated/activated AKT, an event controlled by the mammalian target of rapamycin complex 2 (mTORC2). Downregulation of AKT phosphorylation/activation following FASN inactivation was associated with strong inhibition of rapamycin-insensitive companion of mTOR (Rictor), the major component of mTORC2, at post-transcriptional level. Finally, genetic ablation of Rictor impaired AKT-driven hepatocarcinogenesis in mice. Conclusions FASN is not oncogenic per se in the mouse liver, but is necessary for AKT-driven hepatocarcinogenesis. Pharmacological blockade of FASN might be highly useful in the treatment of human HCC characterized by activation of the AKT pathway.

ObjectiveIncreased de novo fatty acid (FA) synthesis and cholesterol biosynthesis have been independently described in many tumour types, including hepatocellular carcinoma (HCC).DesignWe investigated the functional contribution of fatty acid synthase (Fasn)-mediated de novo FA synthesis in a murine HCC model induced by loss of Pten and overexpression of c-Met (sgPten/c-Met) using liver-specific Fasn knockout mice. Expression arrays and lipidomic analysis were performed to characterise the global gene expression and lipid profiles, respectively, of sgPten/c-Met HCC from wild-type and Fasn knockout mice. Human HCC cell lines were used for in vitro studies.ResultsAblation of Fasn significantly delayed sgPten/c-Met-driven hepatocarcinogenesis in mice. However, eventually, HCC emerged in Fasn knockout mice. Comparative genomic and lipidomic analyses revealed the upregulation of genes involved in cholesterol biosynthesis, as well as decreased triglyceride levels and increased cholesterol esters, in HCC from these mice. Mechanistically, loss of Fasn promoted nuclear localisation and activation of sterol regulatory element binding protein 2 (Srebp2), which triggered cholesterogenesis. Blocking cholesterol synthesis via the dominant negative form of Srebp2 (dnSrebp2) completely prevented sgPten/c-Met-driven hepatocarcinogenesis in Fasn knockout mice. Similarly, silencing of FASN resulted in increased SREBP2 activation and hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase (HMGCR) expression in human HCC cell lines. Concomitant inhibition of FASN-mediated FA synthesis and HMGCR-driven cholesterol production was highly detrimental for HCC cell growth in culture.ConclusionOur study uncovers a novel functional crosstalk between aberrant lipogenesis and cholesterol biosynthesis pathways in hepatocarcinogenesis, whose concomitant inhibition might represent a therapeutic option for HCC.