Transduction of suicide gene into malignant mesothelioma with tumor-specific promoter

用肿瘤特异性启动子将自杀基因转导至恶性间皮瘤

• 批准号: 11670571
• 负责人: INASE Naohiko
• 金额: $ 0.9万
• 依托单位: Tokyo Medical and Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670571/
• 关键词: malignant mesothelioma; gene therapy; tumor-specific expression; specific promoter; keratin 19; calretinin; suicide gene; keratin19; calretinin; 腫瘍特異的プロモーター; 悪性胸膜中皮種

To achieve the specific expression of a transfected suicide gene in malignant mesothelioma cells, we applied the enhancer-promoter fusion sequence of the keratin 19(K19)gene. Northern blot analysis of three mesothelioma cell lines demonstrated that K19 mRNA was expressed most abundantly in the H2052 mesothelioma cell line. Subsequently, in a luciferase reporting assay, K19 promoter(260 bp)exhibited higher promoter activity in H2052 cells than in the other two cell lines. In addition, ligation of a 3' enhancer(80 bp)of the K19 gene to upstream of the K19 promoter sufficiently enhanced the promoter activity. After transfecting an expression vector containing the K19 enhancer-promoter bound thymidine kinase gene(pK19-TK)into the H2052 cells, the pK19-TK transfected cells become more sensitive to GCV than non-transfected cells in vitro and in vivo. The K19 enhancer-promoter sequence seemed to be specific and efficient enough for the expression of the transfected suicide gene in malignant mesothelioma cells.

为了实现自杀基因在恶性间皮瘤细胞中的特异性表达，我们应用了角蛋白19(K19)基因的增强子-启动子融合序列。对3个间皮瘤细胞系的Northern印迹分析表明，K19mRNA在H2052间皮瘤细胞系中表达最丰富。随后，在荧光素酶报告实验中，K19启动子(260bp)在H2052细胞中显示出比在其他两个细胞系中更高的启动子活性。此外，将K19基因的3‘端增强子(80bp)连接到K19启动子的上游，充分提高了启动子的活性。将含有K19增强子-启动子结合的胸苷激酶基因的表达载体(pK19-TK)导入H2052细胞后，pK19-TK细胞在体内外对GCV的敏感性均高于未转染细胞。K19增强子-启动子序列似乎具有足够的特异性和高效性，可以在恶性间皮瘤细胞中表达自杀基因。