| Project/Area Number |
07557095
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 試験 |
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| Research Field |
Cerebral neurosurgery
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| Research Institution | KUMAMOTO UNIVERSITY |
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Principal Investigator |
USHIO Yukitaka Kumamoto University School of Medicine Professor, 医学部, 教授 (20028583)
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| Co-Investigator(Kenkyū-buntansha) |
SUGIMOTO Yoshikazu Taiho Pharmaceutical Co., Ltd., Researcher Cancer Research Lab., Hanno Research, 創薬センター・第一がん研究所, 研究員
NISHI Toru Kumamoto University Hospital Instructor, 医学部附属病院, 助手 (00264309)
KOCHI Masato Kumamoto University School of Medicine Lecturer, 医学部, 講師 (70178218)
KURATSU Jun-ichi Kumamoto University School of Medicine Associate Professor, 医学部, 助教授 (20145296)
SAYA Hideyuki Kumamoto University School of Medicine Professor, 医学部, 教授 (80264282)
竹島 秀雄 熊本大学, 医学部, 助手 (70244134)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥16,100,000 (Direct Cost: ¥16,100,000)
Fiscal Year 1996: ¥7,300,000 (Direct Cost: ¥7,300,000)
Fiscal Year 1995: ¥8,800,000 (Direct Cost: ¥8,800,000)
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| Keywords | electro-genetherapy (EGT) / in vivo electroporation / brain tumor / MCP-1 / lac Z gene / plasmid DNA / chemokine / MCP-1遺伝子 / hMCP1 |
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| Research Abstract |
In this project, we have developed a novel method for high-efficiency and region-controlled in vivo gene transfer by combining in vivo electroporation and intra-arterial plasmid DNA injection. A mammalian expression plasmid for the Escherichia coli lacZ gene (driven with SV40 early promoter) was injected into the internal carotid artery of rats whose brain tumors (from prior inoculation) had been electroporated between two electrodes. The lacZ gene was efficiently transferred and expressed in the tumor cells three days after plasmid injection. Injected DNA was distributed in the whole tumor tissue, while the transfer efficiency was uneven within tumor tissue. This efficient transfer could be achieved only by intra-vascular, especially intra-arterial, delivery of the gene. The fact that the transgene was highly expressed in the border region between tumor and adjacent brain, where tumor neovascularization is abundant, may support this hypothesis. Neither any gene transfers nor any eleva
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ted lacZ activity was detected in tissues outside the electrodes. Because intra-arterial plasmid injection alone could not transfer the gene into the tumor, in vivo electroporation was essential for the efficient transfer of the naked plasmid DNA.Human Monocyte Chemoattractant Protein-1 cDNA was also transferred by this method and its long-lasting (three weeks) expression was confirmed by using the Epstein-Barr virus episomal replicon system. MCP-1 is chemokine which can gather not only monocyte but also T-lymphocyte and has significant roles in the immunization step. The expressed MCP-1 protein was functional as evident by the presence of a large numbers of monocytes in the tumor tissue. A large number of exudative macrophages and lymphocytes was recognized in the tumor tissue by TRPM-3 monoclonal antibody and monoclonal antibody MCA-55, respectively. The plasmid was also transferred into the subcutaneous C6 tumors on rat flank. The growth of treated tumors were inhibited. It seemed that the inhibition of tumor growth in vivo was mediated by those infiltrated cells. It is suggested that the electro-genetherapy using MCP-1 gene is efficient method for solid tumor treatment. The tracts made by electrode insertions were filled with astrocytes and lymphocytes, but there were no reactive changes in the surroundings. No adverse clinical effects, such as seizure and paresis, were observed in rats during the three week follow-up period. This method, we designated electro-genetherapy, which does not require viral genes or particles, allows genes to be transferred and expressed in desired organs or tissues and it may lead to the development of a new type of highly effective gene therapy. Less
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