The combination of anti-cancer drugs with nutritional factors is a potential strategy for improving the efficacy of chemotherapy, particularly for hepatocellular carcinoma because its conventional therapies are mostly ineffective. Using a highly invasive hepatoma SK-Hep-1 cell line, we investigated the possible synergistic anti-metastatic efficacy of a combination of sorafenib (SF), a multi-kinase inhibitor, and β-ionone (BI), a precursor of carotenoids. We found that SF (1 μM) in combination with BI (1 μM) synergistically inhibited cell invasion and additively inhibited cell migration, especially at 48 h of incubation. Mechanistically, the combination of SF and BI was found to decrease the protein expression of focal adhesion kinase (FAK) and Rho, and to enhance the protein expression of tissue inhibitor matrix metalloproteinase (TIMP)-1 and TIMP-2. In addition, the combination of SF and BI inhibited the activity of matrix metalloproteinase (MMP)-2 and MMP-9 and decreased the phosphorylation of FAK and of Rac1 proteins. Importantly, SF enhanced the suppressing effect of BI (1-50 μM) on the viability of SK-Hep-1 cells, but not on murine hepatic BNL CL.2 cells, indicating the selective cytotoxicity of this combination on tumor cells. The combination of SF and BI could be a potential therapeutic strategy against human hepatoma cells.

b-Ionone (BI), a precursor for carotenoids, is widely distributed in fruit and vegetables. Recent in vitro studies have demonstrated the potential anti-metastatic effects of BI, but the mechanisms underlying such actions are not clear. Because liver cancer is the most endemic cancer in Taiwan and in a large region of the world, we hereby investigate the anti-metastatic effects of BI and its mechanisms of actions in a highly metastatic human hepatocarcinoma SK-Hep-1 cells. We show that incubation of cells with BI (1-50 mM) for 24 and 48 h significantly inhibited cell invasion, migration and adhesion. Mechanistically, incubation of cells with BI (1-50 mM) for 24 h resulted in the following: (1) significant inhibition of matrix metalloproteinase (MMP)-2, MMP-9 and urokinase-type plasminogen activator activities, (2) up-regulation of protein expression of the tissue inhibitor of matrix metalloproteinase (TIMP)-1, TIMP-2 and plasminogen activator inhibitor-1, (3) down-regulation of the expression of migration-related proteins, including focal adhesion kinase (FAK), phosphorylated form of FAK, Rho, Rac1 and Cdc42 and (4) up-regulation of the expression of nm23-H1 protein (P, 0·05). Overall, the results show that BI effectively inhibits the metastasis of SK-Hep-1 cells, and this effect involves the regulation of gene expression and signal pathways related to invasion and migration.Key words: b-Ionone: Metastasis: Nm23-H1: Focal adhesion kinase: Rho GTPase Hepatocellular carcinoma is the major cause of cancer death in Taiwan and the most endemic cancer in a large region of the world. Tumour metastasis, both intrahepatic and extrahepatic, is a major factor of mortality in hepatocellular carcinoma patients. Tumour cell metastasis is characteristic of tumour progression involving complex processes including the ability to dissolve the basement membrane and the extracellular matrix (ECM) and to migrate through the ECM. The degradative process is mediated largely by matrix metalloproteinases (MMP), cathepsins and plasminogen activator systems (1) .MMP-2, MMP-9 and urokinase-type plasminogen activator (uPA) are the most vital proteases for degradation of base membrane and, therefore, are deeply involved in cancer metastasis (1 -3) . MMP-2 (72 kDa) and MMP-9 (92 kDa) activities are regulated extracellularly, and their regulations are primarily affected by the balance of pro-enzyme activation and inhibition by tissue inhibitors of matrix metalloproteinase (TIMP), TIMP-2 and TIMP-1, respectively (3 -5) . In addition, serine protease uPA is a protease that cleaves the ECM and activates the conversion of plasminogen to plasma (6) . The conversion of plasminogen to active plasmin is regulated by two specific and fast-acting plasminogen activator inhibitors (PAI), PAI-1 and PAI-2, with PAI-1 being more important (7) . The inhibition of MMP and uPA activity has been adopted as an anti-metastasis therapeutic strategy. Focal adhesion kinase (FAK) is the most extensively studied focal adhesion protein in hepatocellular carc...