Promoter selection for the cytosine deaminase suicide gene constructs in gastric cancer

OBJECTIVE: Carcinomas of the digestive tract represent the second most abundant type of carcinomas in the Western world. During the past two decades, studies of genetic alterations in oncogenes, tumor-suppressor genes, and further cancer-related genes led to growing understanding of molecular mechanisms of gastrointestinal cancer resulting in a genetic progression model. Nevertheless, with a few exceptions, our knowledge of participating genes has not been exploited for gene therapy approaches. Therefore, we monitored promoter activity of a variety of genes shown to be significantly expressed in gastric tumor cells to select optimally active promoters for therapeutical recombinant DNA constructs. When driving a suicide gene these genetic elements can exert cytotoxic effects. METHODS: Using promoter-reporter gene (luciferase) constructs we compared the activities of KRT19, TFF1, SEL1L, MUC4, MUC1, CEL and hTERT by transfecting them into the gastrointestinal cell lines MKN45 and DAN-G for transient expression. After choosing strong promoters we tested the expression of the prokaryotic cytosine deaminase and its cytotoxic effect on the cell cultures. RESULTS: The promoters of SEL1L, MUC1 and KRT19 displayed the highest activity levels in reporter gene assays while other genes reported as upregulated in gastric cancer were moderately expressed. When driving cytosine deaminase in MKN45 cells, the SEL1L promoter induced a 66% cytotoxic effect and the TP1 promoter reached 82%. CONCLUSIONS: From a selection of nine promoter constructs three proved to upregulate the reporter gene well above the level of average activity. They also appear highly capable to drive a suicide gene construct, here tested using prokaryotic cytosine deaminase.