Identification of genes involved in hepatocellular carcinoma : evaluation of hCdc4 and B-Raf genes

Abstract

Hepatocellular carcinoma (HCC), the major type of primary liver cancer, is one of the most common cancers worldwide. Development and progression of HCC occur as a multistep process, requiring the activation of oncogenes and the inactivation of several tumor suppressor genes. Although, inactivation of tumor suppressor genes, including p53, Axin and activation of oncogene β-catenin, have been shown to be involved in HCC development, the molecular mechanism of hepatocarcinogenesis is still unclear. The identification of additional genes that are involved in hepatocarcinogenesis is, not only, an important task to understand this process, but also for development of novel strategies for prevention or therapy. The aim of this study was to elucidate a possible function of hCdc4 and B-Raf genes in HCCs. A gene or genes on chromosome 4q have been implicated in hepatocarcinogenesis by the observation of frequent deletions of this region in HCCs. More recently, hCdc4, a gene in this region has been proposed as a candidate tumor suppressor gene. hCdc4 is an F-box protein which is shown to be involved in ubiquitination of cyclin E, thus targeting it for destruction. Cyclin E overexpression, is reported to be a frequent event in different cancers including HCC suggesting a problem in its destruction. 15 HCC cell lines were analysed for expression and mutation of hCdc4 gene by RT-PCR and direct sequencing of PCR products. No abnormal transcript and mutation observed in HCC cell lines tested. Our findings suggest that alteration of this gene is not a frequent event in hepatocarcinogenesis. iv B-Raf, which is one of the human isoforms of RAF, is activated by oncogenic Ras (leading to cooperative effects in cells responding to growth factor signals). B-Raf mutations are found in a wide range of cancers. Eventhough, mutational activation of Ras is not a frequent event in human hepatocarcinogenesis, few Ras mutations were reported in HCC cases. Thus activation of oncogenic MAP kinase pathway by another component of this pathway such as BRAF is worth to analyze. HCC cell lines and tumours were searched for B-Raf mutations. Activating BRAF missense mutations were identified in 2/72 HCCs (3%). Our results suggest that B-Raf may be occasionally involved in hepatocarcinogenesis. Thus MAP kinase pathway might be involved in hepatocarcinogenesis but neither B-Raf nor Ras are the major players of this pathway in this event. For this reason, other members of this pathway should be evaluated for mutations in HCC.