來自於pri-mir-218-1的miR-218可藉由調控N-Cadherin來抑制肺腺癌細胞的移動

Abstract

Lung cancer is the leading cause of cancer-related mortality in the world. Brain is a major migratory site of lung cancer[1]. So far, the major therapy of brain metastasis is irradiation, but it has a serious side effect. Hence, there is a great need to find efficiency methods for curing brain metastasis. One possible therapeutic strategy is using miRNA to inhibit brain metastatic lung cancer. However, the knowledge of its regulatory mechanism is still very limited. Therefore, as an initial step, the purpose of this study is trying to identify miRNAs that can inhibit the migration of lung adenocarcinoma cells. Here, a brain metastatic lung adenocarcinoma cell line, BM#7, was used as an experimental model. The expression of CDH2 was used as a biomarker for representing metastatic ability. Illumina miRNA microarrays were used to screen the differentially expressed miRNAs in BM#7 cells and parental F4 cells. Bioinformatic tools were used to predict which miRNA can target to CDH2. We identified nine miRNAs that differentially expressed in BM#7 and predicted to target to CDH2. Among these nine miRNAs, since CDH2 was up-regulated in BM#7, we focused on miR-218 that was down-regulated and predicted to target CDH2 by several algorithms. The endogenous expression levels of CDH2 and miR-218 were validated by real-time PCR. Next, to understand the regulatory mechanism of miR-218, we examined the expression levels of miR-218 precursors. Since miR-218 had two precursors, pri-mir-218-1 and pri-mir-218-2, located in SLIT2 and SLIT3 respectively, real-time PCR was used to measure their expression levels. The results showed that decline of miR-218 in BM#7 were due to down-regulation of pri-mir-218-1. Third, to explore whether miR-218 could bind to CDH2, luciferase assays were conducted. The results showed that miR-218 could directly bind to CDH2-3’UTR at two binding sites. Lastly, to investigate the function role of miR-218 on CDH2, transwell migration assay was conducted in BM#7 with miR-218 over-expression. The ability of cell migration was suppressed when miR-218 was up-regulated in BM#7 cells. Taken together, this study showed that the down-regulation of miR-218 was attributed to the decreased expression of pri-mir-218-1 in BM#7, and that miR-218 could inhibit cell migration by targeting to CDH2-3’UTR. MiR-218 might be a novel drug for developing clinical therapies for lung cancer with brain metastasis.