Association between FOXA2 promoter methylation and metastasis in invasive esophageal squamous cell carcinoma

• Main Authors: Ruei-Siang Wu, 吳睿庠
• Other Authors: Ruei-Nian Li
• Format: Others
• Language: zh-TW
• Published: 2014
• Online Access: http://ndltd.ncl.edu.tw/handle/859any

碩士 === 高雄醫學大學 === 生物醫學暨環境生物學研究所 === 102 === Esophageal squamous cell carcinoma (ESCC) is the sixth leading cause of cancer death among men in Taiwan. Because of high level of metastasis, the five-year survival rate is below 10%. Forkhead box A2 (FOXA2, also known as HNF3β), can directly upregulate the metastatic suppressor gene E-cadherin, and inhibit cell migration in many kinds of cancer. The mechanism of metastasis regulation in ESCC is yet unknown. Preliminarily, many CpG sites distribute in FOXA2 promoter with MethPrimer software analysis. We considered low FOXA2 expression owing to hypermethylation in cancer cells. Methylation level of FOXA2 was analyzed by methylation-specific polymerase chain reaction (MSP) in metastatic and non-metastatic ESCC samples. Higher methylation level of FOXA2 promoter in metastatic ESCC was discovered. Therefore, we selected different ESCC cell lines based on metastasis ability to confirm the relationship between FOXA2 expression and metastasis. First, 5-Aza-2’-deoxycitidine (5-aza-dC) was used to demethylate FOXA2 promoter in ESCC cell lines. FOXA2 and E-cadherin reexpression were confirmed by western blot. Furthermore, low cell migration level was confirmed by wound healing assay and Boyden chamber assay after treating with 5-aza-dC. In the meantime, the stronger cell-cell adhesion was discovered. Based on the above results, we constructed E-cadherin promoter luciferase reporter system to confirm FOXA2 influence on E-cadherin promoter. The induction of promoter activity was observed after treating with 5-aza-dC in ESCC cell lines. On the other hand, reexpression of FOXA2 by 5-aza-dC in ESCC cell lines was inhibited by FOXA2 siRNA and E-cadherin expression was also inhibited. According to our results, reexpression of FOXA2 can upregulate E-cadherin expression and has the potential to inhibit metastasis in esophageal cancer.