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Oncotarget. 2017 Sep 28;8(48):84434-84448. doi: 10.18632/oncotarget.21375. eCollection 2017 Oct 13.

DNA hypermethyation and silencing of PITX1 correlated with advanced stage and poor postoperative prognosis of esophageal squamous cell carcinoma.

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Department of Gastroenterology, The Research Center for Hepatitis and Immunology, Research Institute, National Center for Global Health and Medicine, Chiba, Japan.
Department of Surgery, National Center for Global Health and Medicine, Tokyo, Japan.
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Science, The University of Tokyo, Chiba, Japan.
Medical Research Support Center, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
Integrated Database Unit, Advanced Center for Computing and Communication (ACCC), RIKEN, Saitama, Japan.
Pathology Division of Clinical Laboratory, National Center for Global Health and Medicine, Tokyo, Japan.
Cooperative Major in Advanced Health Science, Graduate School of Advanced Science and Engineering, Waseda University, Tokyo, Japan.


Esophageal squamous cell carcinoma (ESCC) is associated with the accumulation of genetic and epigenetic changes in the background mucosa. Dysregulated DNA methylation is known to lead to the inactivation of tumor suppressor genes and the activation of oncogenes. To identify the genes whose expression is perturbed by abnormal DNA methylation in ESCC, integrative transcriptomics by serial analysis of gene expression (SAGE) and methylome sequencing by methyl-DNA immunoprecipitation (MeDIP) analysis were performed. We found 159 genes with significantly decreased expression in ESCC compared to that in noncancerous esophageal mucosa. MeDIP-seq analysis identified hypermethylation in the promoter region of 56 of these genes. Using surgically resected tissues of 40 cases, we confirmed that the paired-like homeodomain 1 (PITX1) gene was hypermethylated in ESCC compared to that in normal tissues (P < 0.0001) by pyrosequencing. PITX1 overexpression in ESCC cell lines inhibited cell growth and colony formation, whereas PITX1 knockdown accelerated cell growth. A PITX1-transfected ESCC cell line, KYSE30, formed smaller tumors in nude mice than in mock-transfected cells. Hypermethylation of PITX1 was associated with tumor depth (P = 0.0011) and advanced tumor stage (P = 0.0052) and predicted poor survival in ESCC (hazard ratio, 0.1538; 95% confidence interval, 0.03159-0.7488; P = 0.0169). In this study, we found a novel tumor suppressor gene of ESCC, PITX1, which is silenced by DNA hypermethylation. Downregulation of PITX1 contributes to the growth and progression of ESCC. Hypermethylation of the PITX1 in ESCC correlated with tumor progression and advanced stage cancer, and may predict a poor prognosis.


homeobox gene; methylome; prognosis marker; transcriptome; tumor suppressor gene

Conflict of interest statement

CONFLICTS OF INTEREST The authors declare no potential conflicts of interest.

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