Abstract 4944: The impact of adenoviral gene transfer of microRNA-34b/c on malignant pleural mesotheliomas

Abstract

Abstract Background: Altered expression of microRNA (miRNA) is strongly implicated in human malignancies. miRNAs are small non-coding RNAs that regulate translations of messenger RNAs of various genes. Similar to encoding genes, tumor suppressive miRNAs are silenced with DNA methylation. The miR-34b/c is direct transcriptional targets of p53 and has been reported to be down-regulated in human malignancies. Unlike other malignant tumors, mutation of TP53 gene is rare in malignant pleural mesotheliomas (MPMs). In our previous study, aberrant methylations of miR-34b/c were frequent in MPMs (−100%), resulting in the slicing of their expression. Ectopic expression of miR-34b/c by transit or stable plasmid transfection, that promoted physiological expression level of miR-34b/c, could inhibit the proliferation and invasion of MPM cells. In this study, we investigate the anti proliferative effect of adenoviral gene transfer of miR-34b/c on MPM cells to aim development of miR-34b/c gene therapy. Methods: We examined three MPM cell lines (H28, H290, and H2052) and a lung cancer cell line (H125). Three adenovirus vectors expressing miR-34b/c (Ad-miR-34b/c), p53 (Ad-p53) and luciferase (Ad-Luc) driven by CMV promoter were generated by homologous recombination and were infected to cell lines above. Expression of miR-34b/c was determined by quantitative PCR. Protein expression, cell proliferation, and cell cycle were analyzed by western blotting, MTS assay, and flow cytometry, respectively. Results: TP53 was wild-type and miR-34b/c was methylated in all three MPM cell lines, while TP53 was mutated and miR-34b/c was unmethylated in H125. After Ad-p53 transfer, p53 and p21 were up-regulated in all four cell lines. Of note, Ad-p53 transfer increased expression of miR-34b/c in H125 but not in all three MPM cell lines, resulting in a decrease of the cell viability in H125 but not in MPM cell lines. The expression of miR-34b and 34c were highly un-regulated after Ad-miR-34b/c transfer in all three MPM cell lines (100 to 2000 fold), that were much higher than physiological level. In addition, the cell viability was also decreased after Ad-miR-34b/c transfer compared with that after Ad-Luc transfer (P < 0.01) in MPM cell lines. Ad-miR-34b/c transfer promoted the strong down-regulation of Bcl-2 protein expression in MPM cell lines and caused an increase in the sub-G0-G1 DNA content, compared with that in cell lines infected with Ad-Luc, indicating the induction of apoptosis. In contrast, Ad-p53 transfer did not induce a drastic change in the sub-G0-G1 DNA content in MPM cell lines. Conclusion: Current study suggests that the adenoviral gene therapy of miR-34b/c, but not TP53, inhibits the cell proliferation of MPM cells by induction of apoptosis, indicating the possibility of miR-34b/c gene therapy in MPMs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4944. doi:10.1158/1538-7445.AM2011-4944