Hepatitis B virus-X protein upregulates TSC1/m-TOR pathway and enhance angiogenesis in hepatitis B associated hepatocellular carcinoma

[[abstract]]Within the hepatic system, chronic hepatitis caused by hepatitis B and C virus predisposes into hepatocellular carcinoma. Hepatocellular carcinoma (HCC) is one of the major causes of cancer death in South-East Asian countries and frequently caused by Hepatitis B virus (HBV) infection. Critical genetic factors involved in the development of HCC by HBV infection are yet to be identified. Recent reports suggest that the Hepatitis B virus-X protein may induce a signaling pathway involving Akt, IKKb, Erk, GSK-3b and b-catenin, which is critical for hepatocytes transformation by HBV infection. TSC1, a well-known tumor suppressor responsible for the development of tuberous sclerosis complex (TSC), negatively regulates the mammalian target of rapamycin (mTOR) pathway. Dysregulation of the TSC/mTOR signaling pathway has been implicated in the development of cancers. To elucidate the molecular pathways of Hepatitis B virus-X protein induced liver tumorigenesis, the signaling pathway involving I B kinases β (IKKβ), tuberous sclerosis complex 1(TSC1), and mammalian target of rapamycin (mTOR) downstream effector S6 kinase(S6K1) was confirmed to be activated in HBV X gene stable expression hepatoma cancer Hep 3B and Hep G2 cells. Phosphorylation of TSC1 and S6K1 was induced in Hepatitis B virus-X protein expression cells. Moreover, overexpression of HBV X protein stimulates activation of IKKb and mTOR and increases secretion of VEGF in the culture supernatants of Hep3B and HepG2 cells. Treatment of these cells with the mTOR inhibitor rapamycin or the IKKβ inhibitor Bay 11-7082 suppressed X protein induced cell proliferation and VEGF production. We next evaluated immunostaining of clinical hepatitis B associated hepatoma tissue specimens which from 95 patients who received curative surgery. Of interest, we found that pIKKβ expression was strongly correlated with pTSC1 (P<0.01) and pS6K1 (P<0.01) level. Furthermore, the over expression of pIKKβ (P<0.001), pTSC1 (P=0.04) and pS6K1 (P