DACH1Methylation In Esophageal Cancer Carcinogenesis And Expression Regulation

BackgroundEsophageal cancer is one of the most malignant human cancers. In recent years,the incidence of esophageal cancer has been increasing. In China, the morbility andmortality are in the highest level of the world. It is a complicated process of developingesophageal cancer from normal esophageal epithelia. It includes activation of oncogeneand inactivation of tumor suppressor gene. Abnormality of Genetics and epigeneticsplays an important role on esophageal carcinogenesis. Gene expression regulated byEpigenetics is not involved in DNA sequence changes. Abnomal epigenetic changesexist in almost all types of human tumors. Silencing of tumor suppressor genes bypromoter region CpG island hypermethylation was found in different signaling pathwayin different cancer. Recent studies have shown that DACH1functions as a tumorsuppressor gene. Reduction of DACH1expression was found in several tumor tissues.But the epigenetic changes of DACH1were not reported in esophageal cancer.ObjectiveOur study is to find esophageal cancer early detection marker by examiningmethylation status of DACH1during human esophageal carcinogenesis from normalesophageal epithelia, to different grade of dysplasia and primary squamous cancer. Theregulation of DACH1in esophageal cancer cell lines is also involved in this study toexplore the esophageal cancer prevention and therapeutic methods.Methods1. Methylation specific PCR (MSP) was used to detect DNA methylation of DACH1in human normal esophageal mucosa, mild esophageal dysplasia, moderate dysplasia,severe dysplasia, and primary human esophageal squamous cancer tissues.2. Methylation specific PCR(MSP) was used to detect DNA methylation of DACH1in6esophageal cancer cell lines: KYSE30、KYSE70、KYSE140、TE1、TE3and KYSE150.3. DACH1expression in mRNA was detected by RT-PCR before and after5-aza-dc(5-aza-2’-deoxycytidine) treatment in6esophageal cancer cell lines.Results 1. No methylation of DACH1promoter region was found in6cases of normalesophageal mucosa tissues. DACH1methylation was found18.8%(6/32),36.4%(4/11),50%(4/8) of mild, miderate and severe esophageal dysplasia respectively, and67.2%(43/64) of primary esophageal squamous esophageal cancer. DACH1ismethylated in a progression tendency during esophageal carcinogenesis（P<0.001）.2. No DACH1methylation was found in KYSE30, KYSE70and KYSE140cell lines,but complete methylation was found in TE1, TE3and KYSE150cell lines.3. DACH1expression was found in KYSE30, KYSE70and KYSE140cell lines, and lossof DACH1expression was found in TE1, TE3and KYSE150cell lines. Restoration ofDACH1expression was revealed by5-aza-dc treatment for96h in TE1, TE3andKYSE150cell lines.ConclusionMethylation of DACH1in promoter region was found frequently in humanprimary esophageal squamous cancer. The methylation status was inprogression tendency during esophageal cancer carcinogenesis from normalesophageal epithelia, different grade of dysplasia to invasive cancer. It suggeststhat DACH1methylation may serve as esophageal cancer early detection marker.DACH1expression was regulated by promoter region hypermethylation in esophagealcancer cell lines. It indicate that demethylation of DACH1may become a therapeuticmethod of esophageal cancer.