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RUNX3 gene promoter demethylation by 5-Aza-CdR induces apoptosis in breast cancer MCF-7 cell line

Authors Kang H, Dai Z , Bai H, Lu W, Ma X, Bao X, Lin S, Wang X

Received 6 February 2013

Accepted for publication 18 March 2013

Published 17 April 2013 Volume 2013:6 Pages 411—417


Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 3

Hua-Feng Kang,* Zhi-Jun Dai,* He-Ping Bai,* Wang-Feng Lu, Xiao-Bin Ma, Xing Bao, Shuai Lin, Xi-Jing Wang

Department of Oncology, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, People's Republic of China

*These authors contributed equally to this work

Abstract: Runt-related transcription factor 3 (RUNX3) is a tumor suppressor gene, its inactivation due to hypermethylation related to carcinogenesis. The aim of this study was to investigate the effects of 5-aza-2'-deoxycytidine (5-Aza-CdR) on cell proliferation and apoptosis by demethylation of the promoter region and restoring the expression of RUNX3 in the breast cancer MCF-7 cell line. MCF-7 cells were cultured with different concentrations (0.4–102.4 µmol/L) of 5-Aza-CdR in vitro. MTT assay was used to determine the proliferation of MCF-7 cells. Flow cytometry and Hoechst staining were used for analyzing cell apoptosis. The methylation status and expression of RUNX3 in mRNA and protein levels were measured by methylation-specific polymerase chain reaction (PCR [MSP]), reverse transcription (RT)-PCR, and Western blot. It was shown that the RUNX3 gene downregulated and hypermethylated in MCF-7 cells. 5-Aza-CdR induced demethylation, upregulated the expression of RUNX3 on both mRNA and protein levels in cancer cells, and induced growth suppression and apoptosis in vitro in a dose- and time-dependent manner. The results demonstrate that RUNX3 downregulation in breast cancer is frequently due to hypermethylation, and that 5-Aza-CdR can inhibit cell proliferation and induce apoptosis by eliminating the methylation status of RUNX3 promoter and restoring its expression.

Keywords: breast cancer, RUNX3, methylation, apoptosis

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