Knock-down of osteopontin expression through lentiviral-mediated RNA interference suppresses the proliferation and invasiveness of hepatocellular carcinoma

AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA 1961 In our previous study, osteopontin (OPN) was identified as one of the leading genes that promoting the metastasis of hepatocellular carcinoma (HCC). However, the mechanism by which OPN is involved in metastasis of HCC is not understood yet. In this study, RNA interference (RNAi) mediated by viral vectors, which could induce down-regulation in gene expression for a longer time, was applied to analyze the roles of OPN in the proliferation, invasion and metastasis of HCC. Three lentiviral vectors encoding microRNA (miRNA) against OPN, Len.OPNi-1, -2 and -3 were constructed and found to down-regulate the OPN expression by 62%, 78% and 95% respectively in HCCLM3 cells ell line which had a higher expression level of OPN and a higher metastatic potential. Consequently, of the expression levels of MMP-2 and uPA were significantly down-regulated which led to a markedly inhibition of invasion and metastasis of HCCLM3 cells ( P< 0.001),. Moreover, besides the inhibitory effect on the invasion and metastasis, Len.OPNi-3 could also suppress the in vitro proliferation and in vivo tumor growth ( P< 0.001), however, it had no significant effect on the proliferation of HepG2 and CCL13 cells which had no overexpression of OPN). Len.OPNi-3, rather than Len.OPNi-2, significantly suppressed the mitogen-activated protein kinase kinase/ extracellular signal-regulated kinase (MEK/ERK1/2) signaling pathway in HCCLM3 cells. The recombinant OPN was found to induce the translocation of NF-κB (p65) into the nucleus of HCC cells and triggered the activation of pro-MMP-2 and MEK/ERK/1/2. These suggest that OPN plays an important role in both the growth and metastasis of HCC, which have a different minimum threshold for the basal expression level of OPN. Lentiviral-mediated RNAi targeting OPN may be a useful strategy to control the HCCs with a high OPN expression.