Study On The Regulation Of MicroRNA - 7 In The Growth And Metastasis Of Hepatocellular Carcinoma

MicroRNAs (miRNAs) are a family of small, non-coding RNAs which play important roles in various biological progressions. Its dysfunction is found in several diseases including cancers. By previous researches, miRNAs were known to be involved in carcinogenesis and tumor progression in hepatocellular carcinoma (HCC). Recently, miRNA-7(miR-7) has been proven to play a substantial role in glioblastoma and breast cancer, but its functions in the context of HCC remain unknown. In our study, we investigated the predictive signaling pathway(s) which miR-7might be involved in and we gave a hypothesis on its regulatory mechanism of HCC progression. All these may give a help for its future application on HCC targeting therapy.We first screened and identified a novel miR-7target, phosphoinositide3-kinase catalytic subunit delta (PIK3CD), by TargetScan and Dual-Luciferase Reporter Aassays. This gene is a key component of phosphoinositide3-kinase (PI3K)/Akt pathway which is associated with cell proliferation and migration. There are4target sites of miR-7harbored in the3’UTR (3’untranslated regions) of PIK3CD mRNA. Overexpression of miR-7would specifically and markedly down-regulate its expression at post-transcriptional level by binding with these target sites. miR-7-overexpressing could arrest cell cycle in G0-G1phase and showed significant cell growth inhibition and significant impairment of cell migration in vitro. To identify the mechanisms, we investigated the PI3K/Akt pathway and found that miR-7-overexpressing could down-regulate the expression of Akt; mTOR and p70S6K but up-regulate the4EBP1expression. We again identified other two novel, putative miR-7target genes, mTOR and p70S6K, which further suggests that miR-7may be a key regulator of the PI3K/Akt pathway. In xenograft animal experiments, we found that overexpressed miR-7effectively repressed tumor growth (3.5-fold decrease in mean tumor volume) in vivo. It also abolished extrahepatic migration from liver to lung in a nude mouse model of metastasis. The number of visible nodules on the lung surface was reduced by32-fold. A negative correlation between miR-7and the expression of PIK3CD and PI3K/Akt pathway was confirmed in xenograft tumor tissues; lung-metastatic nodules and human clinical specimens of HCC respectively. In clinical HCC specimens it also showed a relationship between miR-7expressional level and HCC progression in patients.These findings indicate that miR-7functions as a tumor suppressor and plays a substantial role in inhibiting the tumorigenesis and reversing the metastasis of HCC through the PI3K/Akt/mTOR-signaling pathway in vitro and in vivo. By targeting PIK3CD, mTOR, and p70S6K, miR-7efficiently targets the PI3K/Akt pathway. We herein give a novel hypothesis on the mechanism of microRNA in regulating HCC progression. Given these results, miR-7may be a prospective therapeutic or diagnostic/prognostic tool for HCC.