During the past decade, research on primary liver cancers has particularly highlighted the uncommon plasticity of differentiated parenchymal liver cells (that is, hepatocytes and cholangiocytes (also known as biliary epithelial cells)), the role of liver progenitor cells in malignant transformation, the importance of the tumour microenvironment and the molecular complexity of liver tumours. Whereas other reviews have focused on the landscape of genetic alterations that promote development and progression of primary liver cancers and the role of the tumour microenvironment, the crucial importance of the cellular origin of liver cancer has been much less explored. Therefore, in this Review, we emphasize the importance and complexity of the cellular origin in tumour initiation and progression, and attempt to integrate this aspect with recent discoveries in tumour genomics and the contribution of the disrupted hepatic microenvironment to liver carcinogenesis.

HGF/c-Met supports a pleiotrophic signal transduction pathway that controls stem cell homeostasis. Here, we directly addressed the role of c-Met in stem cell-mediated liver regeneration by utilizing mice harboring c-met floxed alleles and Alb-Cre or Mx1-Cre transgenes. To activate oval cells, the hepatic stem cell (HSC) progeny, we used a model of liver injury induced by diet containing the porphyrinogenic agent, 3, 5-diethocarbonyl-1,4-dihydrocollidine (DDC). Deletion of c-met in oval cells was confirmed in both models by PCR analysis of FACS- sorted EpCam-positive cells. Loss of c-Met receptor decreased sphere-forming capacity of oval cells in vitro as well as reduced oval cell pool, impaired migration and decreased hepatocytic differentiation in vivo as demonstrated by double immunofluorescence using oval- (A6 and EpCam) and hepatocyte-specific (HNF-4α) antibodies. Furthermore, lack of c-Met had a profound effect on tissue remodeling and overall composition of HSC niche which was associated with greatly reduced MMP9 activity and decreased expression of SDF1. Using a combination of double immunofluorescence of cell type-specific markers with MMP9 and gelatin zymography on the isolated cell populations, we identified macrophages as a major source of MMP9 in DDC-treated livers. The Mx1-Cre-driven c-met deletion caused the greatest phenotypic impact on HSCs response as compared to the selective inactivation in the epithelial cell lineages achieved in c-Metfl/fl; Alb-Cre+/- mice. However, in both models, genetic loss of c-met triggered a similar cascade of events leading to failure of HSCs mobilization and death of the mice. Conclusion: These results establish a direct contribution of c-Met in regulation of HSC response, and support a unique role for HGF/c-Met as an essential growth factor signaling pathway for regeneration of diseased liver.

Background & Aims The cancer stem cells (CSCs) have important therapeutic implications for multi-resistant cancers including hepatocellular carcinoma (HCC). Among the key pathways frequently activated in liver CSCs is NF-kB signaling. Methods We evaluated the CSCs-depleting potential of NF-kB inhibition in liver cancer achieved by the IKK inhibitor curcumin, RNAi and specific peptide SN50. The effects on CSCs were assessed by analysis of Side Population (SP), sphere formation and tumorigenicity. Molecular changes were determined by RT-qPCR, global gene expression microarray, EMSA, and Western blotting. Results HCC cell lines exposed to curcumin exhibited differential responses to curcumin and were classified as sensitive and resistant. In sensitive lines, curcumin-mediated induction of cell death was directly related to the extent of NF-kB inhibition. The treatment also led to a selective CSC-depletion as evidenced by a reduced SP size, decreased sphere formation, down-regulation of CSC markers and suppressed tumorigenicity. Similarly, NF-kB inhibition by SN50 and siRNA against p65 suppressed tumor cell growth. In contrast, curcumin-resistant cells displayed a paradoxical increase in proliferation and expression of CSC markers. Mechanistically, an important component of the CSC-depleting activity of curcumin could be attributed to a NF-kB-mediated HDAC inhibition. Co-administration of the class I/II HDAC inhibitor trichostatine sensitized resistant cells to curcumin. Further, integration of a predictive signature of curcumin sensitivity with human HCC database indicated that HCCs with poor prognosis and progenitor features are most likely to benefit from NF-kB inhibition. Conclusions These results demonstrate that blocking NF-kB can specifically target CSC populations and suggest a potential for combined inhibition of NF-kB and HDAC signaling for treatment of liver cancer patients with poor prognosis.

Despite great progress in diagnosis and management of hepatocellular carcinoma (HCC), the exact biology of the tumor remains poorly understood overall limiting the patients' outcome. Detailed analysis and characterization of the molecular mechanisms and subsequently individual prediction of corresponding prognostic traits would revolutionize both diagnosis and treatment of HCC and is the key goal of modern personalized medicine. Over the recent years systematic approaches for the analysis of whole tumor genomes and transcriptomes as well as epigenomes became affordable tools in translational research. This includes simultaneous analyses of thousands of molecular targets using microarray-based technologies as well as next-generation sequencing. Although currently diagnosis and classification of hepatocellular cancers still rely on histological examination of tumor sections, these technologies show great promise to advance the current knowledge of hepatocarcinogenesis, complement diagnostic classification in a setting of microarray-aided pathology, and rationalize the individual drug selection. This review aims to summarize recent progress of system biological approaches in hepatocarcinogenesis and outline potential areas for translational application in a clinical setting. Further, we give an update about known signaling pathways active in HCC, summarize the historical application of whole genomic approaches in liver cancer and indicate ongoing experimental research utilizing novel technologies in diagnosis and treatment of this deadly disease. This will also include the discussion and characterization of new molecular and cellular targets such as Cancer Stem Cells (CSCs).

http://medicalgenomics.org/rna_seq_atlas.

BACKGROUND & AIMS Human primary liver cancer (PLC) is classified into biologically distinct subgroups, based on cellular origin. Liver cancer stem cells (CSCs) have been recently described. We investigated the ability of distinct lineages of hepatic cells to become liver CSCs and the phenotypic and genetic heterogeneity of PLC. METHODS We transduced mouse primary hepatic progenitor cells (HPC), lineage-committed hepatoblasts, and differentiated adult hepatocytes with transgenes encoding oncogenic H-Ras and simian virus 40 large-T antigen. The CSC properties of transduced cells and their ability to form tumors were tested by standard in vitro and in vivo assays and transcriptome profiling. RESULTS Irrespective of origin, all transduced cells acquired markers of CSC/progenitor cells, side populations, and self-renewal capacity in vitro. They also formed a broad spectrum of liver tumors, ranging from cholangiocarcinoma to hepatocellular carcinoma, which resembled human liver tumors, based on genomic and histologic analyses. The tumor cells co-expressed hepatocyte (HNF4A), biliary progenitor cell (keratin 19, EpCAM, A6), and mesenchyme (vimentin) markers and showed disregulation of genes that control the epithelial–mesenchymal transition. Gene expression analyses could distinguish tumors of different cellular origin, indicating the contribution of lineage-stage dependent genetic changes to malignant transformation. Activation of c-Myc and its target genes was required to reprogram adult hepatocytes into CSC and for tumors to develop. Stable knockdown of c-Myc in transformed adult hepatocytes reduced their CSC properties in vitro and suppressed growth of tumors in immunodeficient mice. CONCLUSIONS Any cell type in the mouse hepatic lineage can undergo oncogenic reprogramming into a CSC, by activating different cell type-specific pathways. Identification of common and cell-of-origin specific phenotypic and genetic changes could provide new therapeutic targets for liver cancer.

No abstract

Sirtuin 6 (SIRT6) is a member of the sirtuin family of NAD-dependent deacetylases. Genetic deletion of Sirt6 in mice results in a severe degenerative phenotype with impaired liver function and premature death. So far the role of SIRT6 in development and progression of hepatocellular carcinoma is unknown. We first investigated SIRT6 expression in 153 primary human liver cancers, normal and cirrhotic livers using microarray analysis. SIRT6 was significantly downregulated in both cirrhotic livers and cancer. A Sirt6 knock out (KO) gene expression signature was generated from primary hepatoctyes isolated from three week old Sirt6-deficient animals. Sirt6-deficient hepatocytes showed upregulation of established HCC-biomarkers Alpha-fetoprotein (Afp), Insulin-like growth factor 2 (Igf2), H19 and Glypican-3 (Gpc3). Furthermore decreased SIRT6 expression was observed in hepatoma cell lines that are known to be apoptosis insensitive. Re-expression of SIRT6 in HepG2 cells increased apoptosis sensitivity to CD95-stimulation or chemotherapy treatment. Loss of Sirt6 was characterized by oncogenic changes including global hypomethylation as well as metabolic changes including hypoglycemia and increased fat deposition. The hepatocyte-specific Sirt6-KO signature had prognostic impact and was enriched in patients with poorly differentiated tumors with high AFP levels as well as recurrent disease. Finally, we could demonstrate that the Sirt6-KO signature possessed a predictive value for tumors other than HCC, i.e. breast and lung cancer. Conclusion Loss of SIRT6 induces epigenetic changes which may be relevant to chronic liver diseases and HCC development. Downregulation of SIRT6 and genes dysregulated by loss of SIRT6 possess oncogenic effects in hepatocarcinogenesis. Our data demonstrate that deficiency in one epigenetic regulator predisposes a tumorigenic phenotype which ultimately has relevance for outcome of HCC and other cancer patients.