MiR-139 suppresses metastasis and progression of hepatocellular carcinoma through regulation of Rho-Kinase 2 (ROCK2)

Abstract

Background: Hepatocellular carcinoma (HCC) is an aggressive cancer associated with poor prognosis. Knowledge of the biological mechanisms conferring metastatic capability to cancer cells helps better understanding this lethal disease and may potentially enhance new therapies. Objectives: MicroRNA (miRNA) play important roles in modulating gene expression and has been implicated in human carcinogenesis. However, knowledge about the functions of miRNAs in mediating HCC metastasis is still scarce. In this study, we characterized the miR-139 deregulation in human HCC and its pathological implications in HCC metastasis. Methods: We examined the expression profiles of miRNAs in four paired primary advanced HCCs with q-RT-PCR array. miR-139, the most significantly underexpressed miRNA was selected for further investigation. Expression and clinicopathological significance of miR-139 was determined in an expanded HCC cohort of 67 paired primary HCCs. The roles of miR-139 downregulation and HCC metastasis was further confirmed in 20 microdissected HCC venous metastases. miR-139 stably expressing HCC cells were established with lenti-viral transduction and the effect of miR-139 on HCC migration was tested using transwell cell migration assay. Orthotopic liver tumor implantation model was used to test the effects of miR-139 on HCC metastasis in vivo. The potential miR-139 target, ROCK2, was identified by in silico analysis and verified with luciferase reporter assay and Western Blot analysis. Results: We identified an anti-metastatic microRNA, miR-139, that was significantly downregulated in human HCCs. Underexpression of miR-139 in HCC was closely associated with aggressive and metastatic HCC features. Expression of miR-139 was progressively reduced along hepatocarcinogenesis and was found to be further decreased in metastatic HCC tissues. Overexpression of miR-139 in HCC cells significantly suppressed cell migration and invasion in vitro and remarkably reduced the incidence and severity of lung metastasis in the orthotopic liver tumor implantation model in vivo. Interestingly, we found that miR-139 interacted with the 3’ untranslated region of Rhokinase 2 (ROCK2) and reduced ROCK2 protein expression in HCC cells. Significantly, miR-139 level correlated inversely with ROCK2 protein expression in human HCC specimens. Also, overexpression of miR-139 failed to impede HCC cell motility in ROCK2 knock down cells. Conclusion: Our findings have unraveled a novel mechanistic link between miR-139, ROCK2, and HCC metastasis. We found that underexpression of miR-139 in HCC endowed cancer cells with increased metastatic potential through modulating ROCK2 expression.