The role of microRNA-9 in hepatocellular oncogenesis.

Abstract

168 Background: Hepatocellular carcinoma (HCC) is the main type of liver cancer. MicroRNAs are non-coding RNAs that have been involved in the pathogenesis of different cancer types. Our aim was to identify microRNAs that have functional and clinical significance in liver oncogenesis and understand how manipulation of their levels could have a therapeutic potential. Methods: MicroRNA expression analysis was performed in 38 HCC tissues and 25 normal liver tissues. A microRNA library (318 microRNAs) was transfected in SNU-449 liver cancer cells and invasiveness was measured 24 h later by a matrigel invasion assay. Cell growth and matrigel invasion assays were performed in SNU-449 cells that were transfected with miR-9 or antisense-miR-9 (20nM). TargetScan algorithm was used to identify miR-9 direct downstream target genes. The mRNA levels of PPARalpha, and E-cadherin were assessed by real-time PCR 48h after miR-9 overexpression and down-regulation in SNU-449 cells. Results: MicroRNA expression analysis in 38 HCCs and 25 normal liver tissues identified a 25-microRNA signature of HCC. Integration of the library screen and patient data revealed that miR-9 has clinical and functional relevance for liver cancer. Specifically, we found that miR-9 increases 2.7-fold the invasiveness and the growth of SNU-449 cells, 48h post transfection. Bioinformatic analysis revealed that miR-9 has a binding site in the 3’UTR of PPARalpha gene. MiR-9 overexpression inhibited 35% PPARalpha 3’UTR luciferase activity and 60% mRNA levels, while miR-9 down-regulation led to 75% increased PPARalpha mRNA levels. These data suggest that miR-9 targets directly the 3'UTR of PPARalpha and regulates its expression levels in liver cancer cells. In addition, miR-9 overexpression led to 95% suppression of E-cadherin mRNA levels in SNU-449 cells, thus increasing their metastatic potential. Conclusions: We found that miR-9 is highly overexpressed in liver cancer patients and its up-regulation increases the growth and metastatic potential of liver cancer cells. Overall these data suggest that miR-9 is a novel oncogene involved in liver oncogenesis and its suppression could have therapeutic potential in liver cancer patients.