The Mechanism Of HBV Integration Into TERT And Integration-associated Upregulation Of TERT

Hepatocellular carcinoma(HCC) is the sixth most prevalent malignancy and ranks as the second leading events of cancer deaths worldwide. Integration of hepatitis B virus(HBV) DNA in the host’s genome has been shown to be one cause of hepatocellular carcinoma(HCC) development, with infection appearing to occur in the liver during the early stages of the disease. In the early days, some studies, mainly limited by small-scale or sequencing technique, have suggested that the integration of HBV was a random issue;Nowadays, benefited from improvement of sequencing technique, especially the usage of whole genome sequencing, recent research indicates that the position of hepatitis B virus(HBV) DNA integration in the host genome is not random, with telomerase reverse transcriptase(TERT) being the most recurrent integration gene, and HBV integration was significantly associated with poor prognosis; However, as only small-scale studies have been performed, the molecular mechanism and clinical impact/oncogenicity of this preferential integration into TERT remain poorly understood. So we taken advantage of a high-throughput viral integration detection(HIVID) method with high accuracy could be accurate to a single base-pair(bp) in a large scale cohort(445 patients),focusing on HBV-TERT integration this most recurrent events and get the well understanding of this issue may will be a powerful tool to increase our understanding of molecular mechanism of HBV integration and of its tumorigenesis.Part one: The clinical and pathological significance of HBV-TERT integration in HCC and the related mechanism of integration associated upregulation of TEXT expressionPurposes: To analysis the Clinical and pathological significance of HBV-TERT integration in HBV associated HCC and the related mechanism of integration associated upregulation of TERT expression.Methods : We utilized a high-throughput viral integration detection(HIVID)method with high single base-pair(bp) accuracy to analyze a large patient cohort(445 patients) and putted our attention to the HBV-TERT integration events in order to better understand the clinical significance and mechanisms underlying HBV-TERT integration. After detection of the aberrant expression of TERT caused by HBV integration, the underlying mechanism was used by the Dual-luciferase assays. In addition to the HBV integration events, we further analyzed the two hot mutation, methylation condition and chromatic accessibility of TERT.Results:In the present study, we demonstrate that TERT is not only a recurrent HBV-integrated gene, but also appears to have a region of preferential integration sites. In patients, HBV-TERT integration correlated with gender and microvascular invasion, and patients with integration had a significantly poorer prognosis compared with those without(P=0.046). Notably, HBV integration significantly upregulated the expression of TERT in an integration location and HBV insertion sequence dependent manner Moreover, we analyzed the association between HBV integration and other genomic events that could occur in the TERT promoter,including somatic mutations, aberrant DNA methylation, and changes in chromatic accessibility. While no correlation was observed between HBV integration and methylation or chromatin accessibility, it appears that somatic mutations in the TERT promoter are mutually exclusive with HBV integration.Conclusion: HBV integration into TERT plays an important role in HCC pathogenesis and is the strongest regulator of TERT expression.Part one: the promoting metastasis function of TERT in SMMC-7721 cell linesPurposes: In order to analysis the telomere-independent function of TERT in HCCMethods: We analysis the Clinical and pathological significance of TERT expression in HCC. Furthermore, we constructed a stable TERT-overexpression SMMC-7721 cell lines, and analysis the function of promoting metastasis in vitro and in vivo.Results : TERT upregulation in HCC patients is significantly associated with microvascular invasion(χ 2 =4.061, P=0.044). Further more, the overexpression SMMC-7721 cell showed a more ablity of migration in vitro and could promote the lung metastasis in vivo.Conclusion: In addition to the function of maintaining the telomere, TERT can promote the metastasis in HCC.