Random integration of hepatitis B virus (HBV) DNA into the host genome is frequent in human hepatocellular carcinoma (HCC) and this leads to truncation of the HBV DNA, particularly at the C-terminal end of the HBV X protein (HBx). In this study, we investigated the frequency of this natural C-terminal truncation of HBx in human HCCs and its functional significance. In 50 HBV-positive patients with HCC, full-length HBx was detected in all nontumorous livers. However, full-length HBx was found in only 27 (54%) of the HCC tumors, whereas natural carboxylic acid (COOH)-truncated HBx was found in the remaining 23 (46%) tumors. Upon clinicopathological analysis, the presence of natural COOH-truncated HBx significantly correlated with the presence of venous invasion, a hallmark of metastasis (P 5 0.005). Inducible stable expression of the COOH-truncated HBx protein (with 24 amino acids truncated at the C-terminal end) enhanced the cellinvasive ability of HepG2 cells, as compared to full-length HBx, using the Matrigel cellinvasion assay. It also resulted in increased C-Jun transcriptional activity and enhanced transcription of matrix metalloproteinase 10 (MMP10), whereas activation of the MMP10 promoter by COOH-truncated HBx was abolished when the activator protein 1-binding sites on the MMP10 promoter were mutated. Furthermore, silencing of MMP10 by short interfering RNA in HBxDC1-expressing HepG2 cells resulted in significant reduction of cell invasiveness. Conclusions: Our data suggest that COOH truncation of HBx, particularly with 24 amino acids truncated at the C-terminal end, plays a role in enhancing cell invasiveness and metastasis in HCC by activating MMP10 through C-Jun. (HEPATOLOGY 2012;57:131-139) H epatocellular carcinoma (HCC) is one of the major malignancies worldwide and the second-most common fatal cancer in Southeast Asia, China, and Hong Kong, as a result of the high prevalence of hepatitis B virus (HBV) infection. HBV is a partial double-stranded DNA virus with a 3.2-kb genome containing four open reading frames, including the viral DNA polymerase (P), viral envelope (surface antigens) proteins (PreS1, PreS2, or S), core proteins (PreC or C), and HBV X protein (HBx). Integration of the HBV DNA into the host genome is common in HCC and this may lead to alterations of the host cells by disrupting the expression of cellular genes that are important for cell growth, survival, and cellular differentiation. These cellular genes include cyclin A2, 1 retinoic acid receptor, 2 and human telomerase reverse transcriptase (hTERT).3,4 Moreover, full-length HBx can alter the expression of cellular

HBV is involved in the mammalian pre-mRNA splicing machinery in the generation of potential tumorigenic HBV-human chimeras. This study also provided insight on the impact of intronic HBV integration with the gain of splice sites in the development of HBV-associated HCC.