Oncogenes and Tumor Suppressors miRNA-145 Targets vets Erythroblastosis Virus E 26 Oncogene Homolog 1 to Suppress the Invasion , Metastasis , and Angiogenesis of Gastric Cancer Cells

Recent evidence shows that v-ets erythroblastosis virus E26 oncogene homolog 1 (Ets1) is implicated in tumor development and progression.However, the clinical potentials and underlyingmechanisms of Ets1 in gastric cancer progression and metastasis remain largely unknown. In this study, Ets1 immunostaining was identified in 56 of 84 (66.7%) gastric cancer tissues, whichwas correlatedwith tumor invasion andmetastasis. In gastric cancer specimens and cell lines, miRNA-145 (miR-145) was downregulated and inversely correlated with Ets1 expression. Bioinformatics analysis and luciferase reporter assay revealed that miR-145 directly targeted the 30-untranslated region (30-UTR) of Ets1mRNA. Overexpression or knockdown of miR-145 responsively altered both the mRNA and protein levels of Ets1 and its downstream genes, matrix metalloproteinase (MMP-1)-1 and -9, in gastric cancer cell lines SGC-7901 and MKN-45. Ectopic expression of miR-145 suppressed the invasion, metastasis, and angiogenesis of SGC-7901 and MKN-45 cells in vitro and in vivo. In addition, the effects of miR-145 on Ets1 expression, migration, invasion, and angiogenesis were rescued by restoration of Ets1 expression in these cells. Furthermore, anti-miR-145 inhibitor promoted the migration, invasion, and angiogenesis, whereas siRNAmediated Ets1 knockdown phenocopied the effects of miR-145 overexpression in gastric cancer cells. These results show that miR-145 suppresses Ets1 expression via the binding site in the 30-UTR, thus inhibiting the invasion, metastasis, and angiogenesis of gastric cancer cells. Mol Cancer Res; 11(2); 182–93. 2012 AACR. Introduction Gastric cancer is the fourth most common cancer in humans, and its prognosis still remains poor due to the invasion andmetastasis (1). The metastatic process of cancer cells is complicated and consists of multiple steps, including detachment, local invasion, motility, angiogenesis, extravasation, and growth in distant organs (2). Better elucidating the mechanisms of metastasis is important for improving the therapeutic efficiencies of gastric cancer (1, 2). v-ets erythroblastosis virus E26 oncogene homolog 1 (Ets1) is a member of the Ets transcription factor family that participates in the migration, invasion, and angiogenesis of cancer cells (3). To date, emerging clinical studies have shown that Ets1 plays a critical role in the development and progression of various cancers, including breast cancer, prostate cancer, colon cancer, and liver cancer (4). However, themechanisms underlying Ets1 expression in cancers still remain largely unknown. miRNAs, a novel class of small noncoding RNAs, participate in the posttranscriptional regulation of gene expression through partial complementary binding with the 30 untranslated regions (30-UTR) of target mRNA (5). Because of this unique feature, a singlemiRNAhasmultiple target genes (5). It has been established that miRNAs can function as tumor suppressors or oncogenes, and altered expression of miRNA is involved in the development of cancer (5). For example, miR-10b is highly expressed in metastatic breast cancer cells (6), and positively regulates cell migration and invasion through targeting E-cadherin (7) and syndecan-1 (8). Conversely, the miR-200 family members are downregulated in breast cancer cell lines and reduce epithelial to mesenchymal transition during carcinogenesis by targeting zinc finger E-box binding homeobox 1 and 2 (9). The let-7 miRNAs are underexpressed in lung cancer and negatively regulate the expression of oncogenes Ras (10) and high mobility group AT-hook 2 (11). Thus, it is currently urgent to investigate the expression and function of miRNAs in tumor biology by experimental models. miRNA-145 (miR-145) was first discovered in mouse heart muscle (12) and later verified in humans (13). Recently, downregulatedmiR-145 expression has been identified in Authors' Affiliations: Departments of Pathology, Surgery, and Cardiology; and Clinical Center ofHumanGenomicResearch, UnionHospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province, PR China Note: Supplementary data for this article are available at Molecular Cancer Research Online (http://mcr.aacrjournals.org/). Corresponding Author: Qiangsong Tong, Department of Surgery, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, PR China. Phone: 86-2785726005; Fax: 86-27-85726821; E-mail: qs_tong@hotmail.com or qstong@mail.hust. edu.cn doi: 10.1158/1541-7786.MCR-12-0534 2012 American Association for Cancer Research. Molecular Cancer Research Mol Cancer Res; 11(2) February 2013 182 on June 19, 2017. © 2013 American Association for Cancer Research. mcr.aacrjournals. org Downloaded from Published OnlineFirst December 11, 2012; DOI: 10.1158/1541-7786.MCR-12-0534