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Cell-cidal effect of the combination therapy of p53 gene under the control of radiation- and hypoxia-sensitized promoter and heavy particle irradiation in glioma

放疗缺氧增敏启动子控制下的p53基因与重粒子线照射联合治疗胶质瘤细胞杀伤作用

基本信息

批准号：
16591466
负责人：
KOSHIKAWA Nobuko
金额：
$ 1.41万
依托单位：
Chiba cancer center research institute
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

项目摘要

AIM : Malignant gliomas, especially the glioma cells under hypoxic microenvironments, are refractory to conventional radiotherapy and chemotherapy. The aims of this study were to examine the efficacy of combination of radiation/hypoxia dual-sensitive promoter-regulated p53 gene and high-LET radiation for killing glioma cells with mutant p53 under normoxic and hypoxic conditions and to compare the effectiveness as a 'trigger' to activate therapeutic gene expression between high-LET and low-LET radiation. METHODS : The chimeric promoter Egr-HRE was generated by insertion of CArG elements derived from Egr-1 gene and hypoxia response elements(HREs) derived from erythropoietin gene. p53 expression vector was constructed by cloning of the Egr-HRE promoter upstream of p53 gene. After transfecting the plasmid into p53-mutant U373 human glioma cell line, the cells were exposed to hypoxia (0.1% O_2, 24 h) and high-LET (1 Gy) or low-LET radiation (1 Gy). The expression level of p53 and the cell killing effect was determined by Western blotting and colony formation assay, respectively. RESULTS : The results showed that the exposure of the cells to high-LET carbon ions and hypoxia greatly enhanced the expression of p53 and accordingly was most effective in killing the glioma cells. When compared with low-LET radiation, high-LET carbon ions were more effective in enhancing p53 expression and killing the cells. CONCLUSION : The results suggest that the combination of radiation/hypoxia dual promoter-regulated p53 gene and high-LET carbon ions is a promising strategy for effective gene therapy of malignant gliomas and that high-LET radiation is more effective to activate therapeutic gene expression than low-LET radiation.

目的：恶性胶质瘤，尤其是缺氧微环境下的胶质瘤细胞，是常规放化疗难以治疗的肿瘤。本研究的目的是检查组合的辐射/缺氧双敏感启动子调控的p53基因和高LET辐射的疗效杀死胶质瘤细胞与突变p53在常氧和缺氧条件下，并比较有效性作为一个“触发”激活治疗基因的表达之间的高LET和低LET辐射。方法：采用正交试验设计嵌合启动子Egr-HRE是通过插入Egr-1基因的CArG元件和促红细胞生成素基因的缺氧反应元件（HRE）而产生的。通过克隆p53基因上游的Egr-HRE启动子构建p53表达载体。将该质粒转染p53突变型人脑胶质瘤细胞系U373后，将细胞暴露于低氧（0.1%O_2，24 h）和高LET（1戈伊）或低LET（1戈伊）照射。Western blotting法检测p53蛋白表达水平，克隆形成实验检测细胞杀伤作用。研究结果：结果表明，高LET碳离子和低氧暴露显著增强了p53的表达，从而对胶质瘤细胞的杀伤作用最强。与低LET辐射相比，高LET碳离子在增强p53表达和杀死细胞方面更有效。结论：结果提示，高LET碳离子与辐射/低氧双启动子调控的p53基因联合应用是一种有效的恶性胶质瘤基因治疗策略，高LET辐射比低LET辐射更能有效激活治疗基因的表达。

项目成果

期刊论文数量（36）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

A calcium binding protein, S100A4, mediates T cell dependent cytotoxicity as a transformation-associated antigen

钙结合蛋白 S100A4 作为转化相关抗原介导 T 细胞依赖性细胞毒性

DOI：
发表时间：
2004
期刊：
Microbiology and Immunology 49・1
影响因子：
0
作者：
Kondo N;et al.
通讯作者：
et al.

A procedure for culturing astrocytes from white matter and the application of the siTNA technique for silencing the expression of their specific marker, S100A4

从白质培养星形胶质细胞的程序以及应用 siTNA 技术沉默其特定标记 S100A4 的表达

DOI：
发表时间：
2006
期刊：
Brain ResBrain Res Protoc. 15
影响因子：
0
作者：
Shigenori Y;Katayama Y;Mori T;Maeda T;Kawamata T;Tohyama H;Brain ResBrain Res Protoc.
通讯作者：
Brain ResBrain Res Protoc.

Hypoxia-regulated expression of attenuated diphtheria toxin A fused with hypoxia-inducible factor-lalpha oxygen-dependent degradatiott domain preferentially induces apoptosis ofhypoxic cells in solid tumor.

与缺氧诱导因子-lα氧依赖性降解结构域融合的减毒白喉毒素A的缺氧调节表达优先诱导实体瘤中缺氧细胞的凋亡。