Hypermethylation of BEND5 contributes to cell proliferation and is a prognostic marker of colorectal cancer

// Ruo-Kai Lin 1, 2, 9 , Wan-Yu Hung 2 , Yu-Fang Huang 1 , Yu-Jia Chang 3 , Chien-Hsing Lin 4 , Wei-Yu Chen 5, 6 , Shih-Feng Chiu 7 , Shih-Ching Chang 8 and Shih-Feng Tsai 4 1 Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan, R.O.C. 2 Master Program for Clinical Pharmacogenomics and Pharmacoproteomics, Taipei Medical University, Taipei, Taiwan, R.O.C. 3 Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan, R.O.C. 4 Institute of Molecular and Genomic Medicine, National Health Research Institutes, Zhunan, Miaoli, Taiwan 5 Department of Pathology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan, R.O.C. 6 Department of Pathology, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan, R.O.C. 7 Professional Master Program in Pharmaceutics and Biotechnology, Taipei Medical University, Taipei, Taiwan, R.O.C. 8 Division of Colon and Rectal Surgery, Department of Surgery, Taipei Veterans General Hospital, Taipei, Taiwan, R.O.C. 9 PH.D Program for Clinical Drug Development of Chinese Herbal Medicine, Ph.D Program in Biotechnology Research and Development, College of Pharmacy, Taipei Medical University, Taipei, Taiwan, R.O.C. Correspondence to: Shih-Feng Tsai, email: petsai@nhri.org.tw Shih-Ching Chang, email: changsc@vghtpe.gov.tw Keywords: BEND5, colorectal cancer, DNA methylation, prognostic marker, tumor suppressor genes Received: September 09, 2016      Accepted: May 10, 2017      Published: November 01, 2017 ABSTRACT Aberrant hypermethylation of CpG islands in tumor suppressor genes (TSGs) contributes to colorectal tumorigenesis. To identify new colorectal cancer (CRC) screening marker, we investigated DNA methylation alterations in novel TSGs. Using HumanMethylation450 BeadChip arrays, CpG regions in BEND5 were the most highly methylated among all genomic regions in 26 colorectal tumors compared to paired non-neoplastic tissues from a Taiwan cohort. Therefore, BEND5 was selected for further analysis. Quantitative methylation-specific real-time PCR revealed that 86.7% (117/135) of CRC patients exhibited hypermethylated BEND5 . Real-time reverse transcription PCR identified that BEND5 mRNA expression was downregulated in 68% (32/47) of the analyzed samples. BEND5 hypermethylation was associated with poor overall survival (OS) in Taiwan patients with early-stage CRC ( P = 0.037). In a CRC tissue set from South Korea, OS was higher in patients with high BEND5 protein expression than in those with low BEND5 protein expression ( P = 0.037) by using immunohistochemistry assays. Consistently, BEND5 hypermethylation was associated with poor OS in patients with early-stage CRC in The Cancer Genome Atlas (TCGA) data set ( P = 0.003). Multivariate Cox proportional hazards regression analysis further supported that hypermethylation of BEND5 genes was significantly associated with OS in Taiwan and TCGA CRC patients ( P = 0.023 and 0.033, respectively). Finally, the cell model assay with transient transfection of BEND5 or si-BEND5 knockdown indicated that BEND5 inhibited cancer cell proliferation. In conclusion, epigenetic alteration in the candidate TSG BEND5 contributes to colorectal cancer development and is a prognostic marker of CRC.