Abstract: Objective : To investigate the methylation of a CpG island in the MT-3 gene in tissues of esophageal can-cer and its clinical significance. Methods : A total of 68 patients with esophageal cancer who underwentsurgery between May and July 2005 were analyzed. There were 49 males and 19 females, ranging from 42 to76 years old. The numbers of cases of esophageal cancer in the upper, middle, and lower part of the esopha-gus were 6, 46, and 16, respectively. There were 3 stage Ⅰ cases, 13 stage IIA cases, 29 stage IIB cases, 22stage Ⅲ cases, and 1 stage Ⅳ case. Eight cases were well differentiated, 54 cases were moderately differen-tiated, and 6 cases were poorly differentiated. All of the patients suffered from squamous cell carcinoma andwere followed up for more than 30 months. Samples of tumor and normal margin tissues were collected. DNAand RNA were extracted from these tissues. Combined bisulfite restriction analysis (COBRA) was employedto investigate the status of MT-3 methylation. Real-time RT-PCR was used to detect the levels of MT-3 mRNAexpression. The results of COBRA and RT-PCR, clinicopathologic parameters, and follow-up data were ana-lyzed. Results : MT-3 methylation was found in 16.2% (11/68) of the tumor tissues. However, no methylationwas detected in the normal tissues from the upper or lower margin. The expression of MT3 mRNA in methylat-ed and unmethylated tumor tissues was 2.54±1.11 and 5.13±1.86, respectively (P=0.027). MT-3 methylationwas correlated with TNM stage, lymph node metastasis, and survival time but was not correlated with gender,tumor localization, differentiation grade or tumor depth. Conclusion : Methylation of the MT-3 gene can impactits mRNA expression in esophageal cancer, especially in advanced esophageal cancer. Methylation of theMT-3 gene is correlated with lymph node metastasis and implies poor prognosis.