TFF1 targeting therapeutic strategy for dissemination of gastric cancer

三叶因子1（TFF 1）被认为是胃癌的抑癌基因。然而，TFF 1表达及其调控在胃癌患者中的作用仍不清楚。本研究的目的是阐明TFF 1的临床意义，并确定其调节机制。我们评估了182例胃癌患者中TFF 1的免疫组化表达，并研究了TFF 1是否与临床病理因素和患者生存相关。使用TFF 1敲低的胃癌细胞的体外研究评估了TFF 1在癌症侵袭中的作用。进行亚硫酸氢盐测序以评估细胞和切除组织中TFF 1的DNA甲基化。TFF 1低表达患者的肿瘤浸润深度明显高于高表达患者（p=0.037）。TFF 1的低表达也与108例仅接受手术治疗的患者的生存率低相关（p=0.029）。多因素分析显示，TFF 1表达和淋巴结转移均为疾病特异性生存期的独立预测因素。在体外研究中，与对照细胞相比，TFF 1缺陷细胞的细胞侵袭力显著增加。硫酸氢盐测序结果显示TFF 1在胃癌细胞和组织中的表达强烈依赖于DNA甲基化。有趣的是，位于TATA盒和缺氧反应元件（HRE）附近的两个特定CpG位点的甲基化状态决定了切除组织中TFF 1的表达。TFF 1表达被DNA甲基化沉默，并与胃癌患者的肿瘤侵袭和不良生存相关。因此，TFF 1的表达和/或甲基化状态可以作为预测晚期胃癌患者生存的有用生物标志物。

Trefoil factor 1 (TFF1) is considered to be a tumor suppressor gene in gastric cancer. However, the role of TFF1 expression and its regulation in gastric cancer patients remain unclear. The aims of this study were to clarify the clinical significance of TFF1 and to determine its regulatory mechanisms. We assessed the immunohistochemical expression of TFF1 in 182 gastric cancer patients and examined whether or not TFF1 is associated with the clinicopathological factors and patient survival. In vitro study using TFF1 knockdown gastric cancer cells evaluated the role of TFF1 in cancer invasion. Bisulfite sequencing was performed to assess DNA methylation of TFF1 in cells and resected tissues. Patients with low expression of TFF1 showed a significantly deeper invasion of the tumor than those with high expression (p=0.037). Low expression of TFF1 was also associated with a poor survival (p=0.029) in 108 patients who were treated by surgery alone. Both TFF1 expression and lymph node metastasis are independent predictive factors for disease-specific survival in a multivariate analysis. In an in vitro study, invasive power of the cells was significantly increased in the　TFF1-deficient cells compared with the control cells. Bisulfate sequencing showed that TFF1 expression is strongly dependent on DNA methylation in both gastric cancer cells and tissues. Interestingly, methylation status of two specific CpG sites, which are located close to a TATA box and hypoxia response element (HRE), determined the TFF1 expression in the resected tissues. TFF1 expression is silenced by DNA methylation and is associated with tumor invasion and a poor survival in gastric cancer patients. The expression and　or methylation status of TFF1 may, therefore, serve as a useful biomarker for predicting survival in patients with advanced gastric cancer.