Tumorsuppressive MicroRNA-188-5p Reveals Novel Oncogenes for Hepatocellular Carcinoma

 

Background & Aims: MicroRNAs are known to be dysregulated in Hepatocellular Carcinoma (HCC) and to play crucial roles in tumor development and progression. In a preliminary study, our group found that miR-188-5p is downregulated in activated synovial fibroblasts in rheumatoid arthritis (RASF). RASF reveal tumor-like features such as enhanced migration and proliferation. Re-Expression of miR-188-5p strongly inhibited migration in RASF (Int J Clin Exp Pathol, 2015). Chloride voltage-gated channel 5 (CLCN5), which is the host gene for miR-188-5p, was shown to be induced by Hepatocyte nuclear factor (HNF) 1 (Am J Physiol Renal Physiol., 2010). Interestingly, our group demonstrated in a previous study that HNF-1 is a lost tumorsuppressor in HCC (Gut, 2008). Therefore, in this project, our aim was to explore a possible role of miR-188-5p in cancer development and progression with a focus on HCC.

Methods & Results: For functional analysis, transient transfection of pre-miR-188-5p was performed in HCC cell lines (PLC, Hep3B, and HepG2). MiR-188-5p markedly inhibited migration and proliferation and reduced clonogenicity in HCC cell lines. Quantitative RT-PCR analysis showed that miR-188-5p expression was significantly downregulated in HCC tissue samples from patients as compared to corresponding non-tumorous liver tissues. Moreover, miR-188-5p expression was strongly reduced in HCC cell lines as compared to primary human hepatocytes (PHH). Furthermore, first experiments showed that HNF-1 overexpression enhances miR-188-5p expression in HCC cell lines, and HNF-1 induced inhibition of clonogenicity was ‘rescued’ by anti-miR-188-5p. For determination of potential novel target genes of the miR-188-5p in HCC, cDNA array analysis comparing HCC cells with and without re-expressed miR-188-5p was performed, and revealed a list of several potential novel target genes for miR-188-5p, including Ephrin B2 (EFNB2), Discs large MAGUK scaffold protein 5 (DLG5), and Tumor protein p63 regulated 1 like (TPRG1L). We will further investigate these potential novel oncogenes in HCC development and progression and their regulation by miR-188-5p.

Conclusions: We found that a mostly unknown microRNA in cancer, miR-188-5p, is downregulated in HCC cells and re-expression experiments revealed potent tumorsuppressive functions. Moreover, we found that miR-188-5p expression is regulated by Hepatocyte Nuclear Factor 1, a downregulated tumorsuppressor in HCC. cDNA array analysis comparing HCC cells with and without re-expressed miR-188-5p revealed a list of several potential novel target genes for miR-188-5p. We will further investigate these potential novel oncogenes and evaluate possible therapeutic approaches for HCC.

References:

[1] Hellerbrand C, Amann T, Schlegel J, et al. The novel gene MIA2 acts as a tumour suppressor in hepatocellular carcinoma. Gut (2008)

[2] Ruedel A*, Dietrich P*, Schubert T, et al. Expression and function of microRNA-188 – 5 p in activated rheumatoid arthritis synovial fibroblasts. Int J Clin Exp Pathol (2015) (*contributed equally)

[3] Tanaka K, Terryn S, Geffers L, et al. The transcription factor HNF1α regulates expression of chloride-proton exchanger ClC-5 in the renal proximal tubule. Am J Physiol Renal Physiol. (2010)