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Int J Med Sci 2014; 11(8):779-787. doi:10.7150/ijms.7405

Research Paper

Prognostic CpG Methylation Biomarkers Identified by Methylation Array in Esophageal Squamous Cell Carcinoma Patients

I-Ying Kuo1, Jia-Ming Chang2,3, Shih-Sheng Jiang4, Chung-Hsin Chen4, I-Shou Chang4, Bor-Shyang Sheu5, Pei-Jung Lu2, Wei-Lun Chang5, Wu-Wei Lai6, Yi-Ching Wang1,2,7✉

1. Institute of Basic Medical Sciences, National Cheng Kung University, Tainan, Taiwan;
2. Institute of Clinical Medicine, National Cheng Kung University, Tainan, Taiwan;
3. Department of Surgery, Chia-Yi Christian Hospital, Chiayi, Taiwan;
4. National Institute of Cancer Research, National Health Research Institutes, Miaoli, Taiwan;
5. Department of Internal Medicine, National Cheng Kung University Hospital, Tainan, Taiwan;
6. Department of Surgery, National Cheng Kung University Hospital, Tainan, Taiwan;
7. Department of Pharmacology, National Cheng Kung University, Tainan, Taiwan.


Background: Esophageal squamous cell carcinoma (ESCC) is an aggressive cancer with poor prognosis. We aimed to identify a panel of CpG methylation biomarkers for prognosis prediction of ESCC patients.

Methods: Illumina's GoldenGate methylation array, supervised principal components, Kaplan-Meier survival analyses and Cox regression model were conducted on dissected tumor tissues from a training cohort of 40 ESCC patients to identify potential CpG methylation biomarkers. Pyrosequencing quantitative methylation assay were performed to validate prognostic CpG methylation biomarkers in 61 ESCC patients. The correlation between DNA methylation and RNA expression of a validated marker, SOX17, was examined in a validation cohort of 61 ESCC patients.

Results: We identified a panel of nine CpG methylation probes located at promoter or exon1 region of eight genes including DDIT3, FES, FLT3, NTRK3, SEPT5, SEPT9, SOX1, and SOX17, for prognosis prediction in ESCC patients. Risk score calculated using the eight-gene panel statistically predicted poor outcome for patients with high risk score. These eight-gene also showed a significantly higher methylation level in tumor tissues than their corresponding normal samples in all patients analyzed. In addition, we also detected an inverse correlation between CpG hypermethylation and the mRNA expression level of SOX17 gene in ESCC patients, indicating that DNA hypermethylation was responsible for decreased expression of SOX17.

Conclusions: This study established a proof-of-concept CpG methylation biomarker panel for ESCC prognosis that can be further validated by multiple cohort studies. Functional characterization of the eight prognostic methylation genes in our biomarker panel could help to dissect the mechanism of ESCC tumorigenesis.

Keywords: esophageal squamous cell carcinoma, CpG methylation, DNA methylation array, pyrosequencing, prognosis.

This is an open access article distributed under the terms of the Creative Commons Attribution (CC BY-NC) License. See for full terms and conditions.
How to cite this article:
Kuo IY, Chang JM, Jiang SS, Chen CH, Chang IS, Sheu BS, Lu PJ, Chang WL, Lai WW, Wang YC. Prognostic CpG Methylation Biomarkers Identified by Methylation Array in Esophageal Squamous Cell Carcinoma Patients. Int J Med Sci 2014; 11(8):779-787. doi:10.7150/ijms.7405. Available from