Unit: Oncogenesis and Molecular Virology - INSERM U.579
Director: Marie-Annick BUENDIA
Our Unit is investigating the molecular mechanisms of liver carcinogenesis in humans and in murine models. We are particularly interested in better understanding the role of hepatitis B virus, notably the regulatory protein HBx, and in defining the factors implicated in liver cell transformation upon abnormal reactivation of the WNT/ß-catenin signaling pathway in hepatocellular carcinoma and hepatoblastoma.
Hepatocellular carcinoma (HCC) is the predominant type of primary liver cancer, affecting epithelial cells of the hepatic parenchyma, the hepatocytes. This tumor ranks among of the commonest human cancers, and because it is usually fatal within a few months of diagnosis, it represents a leading cause of cancer death worldwide. HCC is also one of the rare human neoplasms epidemiologically related to viral infections : over 70% of HCC cases are associated with chronic infection with hepatitis B (HBV) or hepatitis C virus (HCV). In particular, HBV infection remains a major risk factor, despite the availability of an efficient and safe vaccine. However, the molecular basis of liver carcinogenesis remains incompletely understood. Several oncogenic pathways have been implicated, including abnormal reactivation of WNT/ß-catenin signaling. This pathway plays a predominant role in the development of hepatoblastoma, a rare embryonic liver tumor.
Our research Unit was created in April 2003. During this year, we have analyzed the mechanisms regulating the functional activity of two proteins involved in liver oncogenesis : the transcriptional transactivator HBx of hepatitis B virus, and the oncoprotein ß-catenin, which is frequently mutated in human cancers. A major objective is to identify the cellular genes that are activated by these proteins, and to better understand their role in human and murine hepatocarcinogenesis.
1- Analysis of the viral regulatory protein HBx
(Christine Neuveut, Yuanfei WU, Julie Caramel, Delphine Cougot)
The HBx protein of hepatitis B virus is a transcriptional transactivator endowed with weak oncogenic properties, and it has been implicated in cell cycle control and apoptosis, and as a co-factor in liver carcinogenesis. Our project aims at better understanding the regulation of HBx transcriptional activity. We are currently analyzing HBx interactions with histone acetyltransferases and deacetylases and subsequent post-translational modifications of HBx. Our recent data indicate that HBx is acetylated in vivo, and we are investigating the effects of this modification on known HBx functions, including interaction with cellular partners, subcellular localization, and role in viral replication and oncogenesis. A second theme is the search for cellular genes that are regulated by HBx in primary human hepatocytes and in mouse liver. By its activities in the viral life cycle and in liver tumorigenesis, HBx represents a major therapeutic target for combating HBV-induced carcinogenesis.
2- Oncogenic activation of ß-catenin
(Yu Wei, Charlotte Labalette, Christine Neuveut, Claire-Angélique Renard)
We have previously demonstrated that the Wnt pathway is frequently activated by stabilizing mutations of ß-catenin in HCC and in hepatoblastoma. Our objectives are to characterize the network of factors involved in liver carcinogenesis induced by oncogenic ß-catenin, including interacting partners and downstream target genes of ß-catenin in the liver context. In a recent work, we have identified the LIM protein FHL2 as a new ß-catenin binding protein. LIM domains are made of double zinc fingers rich in cysteines and histidines, and LIM proteins are viewed as adaptors capable of mediating protein interactions. We have shown that FHL2 behaves as a coactivator of the nuclear ß-catenin/Tcf complex, and that it interacts and synergizes with CBP/p300 in transcriptional activation of Wnt-responsive genes. In parallel studies, we have demonstrated that interaction of ß-catenin with p300 leads to acetylation of ß-catenin at lysine 345, thus increasing ß-catenin affinity for Tcf4 and transcriptional activity of the ß-catenin-Tcf complex. This process is stimulated by FHL2. The second aspect of our project is to identify downstream targets of activated ß-catenin using microarray analysis in primary human hepatocytes and murine hepatocellular carcinomas. Our recent work has allowed us to characterize a number of ß-catenin target genes in the liver context, and we are studying the role of these genes in liver carcinogenesis. Our group is the main investigator for the study of hepatoblastoma in a national program of tumor expression profiling (CIT project of the Ligue Nationale contre le Cancer), in collaboration with different French hospitals. The aim of our project is to generate new molecular markers for tumor diagnosis and prognosis, and novel approaches for liver cancer therapy.
3- Murine models of liver tumorigenesis
(Claire-Angélique Renard, Charlotte Labalette, Yu Wei)
In WHV/myc transgenic mice, liver-specific expression of the c-myc or N-myc oncogene driven by hepatitis virus regulatory elements leads to the development of primary liver tumors in all animals within 12 months in average. These models are used to analyze the oncogenic steps in liver oncogenesis. A major project is to analyze gene expression profiles and genetic alterations of liver tumors induced by Myc in the context of the CIT program of the Ligue Nationale contre le Cancer. In a collaborative project recently initiated within Pasteur Institute (PTR 136), the WHV/myc transgenic mice will be used to monitor potential anti-tumor effects of proapoptotic biopeptides.
The importance of the coactivator FHL2 in tumorigenesis will be evaluated in murine models. Both FHL2-null mice (collaboration: Ju Chen, UCSD) after crossing with different oncomice that develop liver or colon tumors, and transgenic mice expressing FHL2 in liver and intestine under control of the ApoA-IV promoter-enhancer.
Photo : ß-catenin expression in human liver (left) and in hepatocellular carcinoma carrying a stabilizing mutation of ß-catenin (right). The normal ß-catenin protein is localized at the lateral plasma membrane in hepatocytes, and the mutated form accumulates in cytoplasmic and nuclear compartments of tumor cells.
Keywords: cancer, virology, hepatitis, transcription regulation, acetylation, transgenic mice