Expression of ECRG4, a Novel Esophageal Cancer-Related Gene, Downregulated by CpG Island Hypermethylation in Human Esophageal Squamous Cell Carcinoma

World J Gastroenterol. 2003 Jun;9(6):1174-8. doi: 10.3748/wjg.v9.i6.1174.


Aim: To study the mechanisms responsible for inactivation of a novel esophageal cancer related gene 4 (ECRG4) in esophageal squamous cell carcinoma (ESCC).

Methods: A pair of primers was designed to amplify a 220 bp fragment, which contains 16 CpG sites in the core promoter region of the ECRG 4 gene. PCR products of bisulfite-modified CpG islands were analyzed by denaturing high-performance liquid chromatography (DHPLC), which were confirmed by DNA sequencing. The methylation status of ECRG 4 promoter in 20 cases of esophageal cancer and the adjacent normal tissues, 5 human tumor cell lines (esophageal cancer cell line-NEC, EC109, EC9706; gastric cancer cell line- GLC; human embryo kidney cell line-Hek293) and 2 normal esophagus tissues were detected. The expression level of the ECRG 4 gene in these samples was examined by RT-PCR.

Results: The expression level of ECRG 4 gene was varied. Of 20 esophageal cancer tissues, nine were unexpressed, six were lowly expressed and five were highly expressed compared with the adjacent tissues and the 2 normal esophageal epithelia. In addition, 4 out of the 5 human cell lines were also unexpressed. A high frequency of methylation was revealed in 12 (8 unexpressed and 4 lowly expressed) of the 15 (80 %) downregulated cancer tissues and 3 of the 4 unexpressed cell lines. No methylation peak was observed in the two highly expressed normal esophageal epithelia and the methylation frequency was low (3/20) among the 20 cases in the highly expressed adjacent tissues. The methylation status of the samples was consistent with the result of DNA sequencing.

Conclusion: These results indicate that the inactivation of ECRG 4 gene by hypermethylation is a frequent molecular event in ESCC and may be involved in the carcinogenesis of this cancer.

MeSH terms

  • Carcinoma, Squamous Cell / genetics*
  • CpG Islands / genetics*
  • DNA Methylation*
  • Down-Regulation*
  • Esophageal Neoplasms / genetics*
  • Gene Expression*
  • Humans
  • Neoplasm Proteins
  • Proteins / genetics*
  • Tumor Cells, Cultured
  • Tumor Suppressor Proteins


  • ECRG4 protein, human
  • Neoplasm Proteins
  • Proteins
  • Tumor Suppressor Proteins