| Project/Area Number |
17591352
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General surgery
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| Research Institution | Tokyo Women's Medical University |
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Principal Investigator |
YAMADA Osamu Tokyo Women's Medical University, Medical Research Institute, Associate Professor, 医学部, 助教授 (30167712)
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| Project Period (FY) |
2005 – 2006
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| Project Status |
Completed (Fiscal Year 2006)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2006: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2005: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | cancer cell / suicide bomb / telomerase / gene transfection |
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| Research Abstract |
The promoter of telomerase is active in virtually all types of tumors but is silent in most adult somatic cells. We placed the suicide genes under the control of the telomerase promoter with the aim of restricting their expression to tumor cells. As the suicide genes, we used 1. the herpes simplex virus thymidine kinase gene which may confer ganciclovir sensitivity to all tumor cells and 2. bovine α1-3 galactosyltransferase (α1-3GT) cDNA which produces the αGal epitope. The carbohydrate epitope, αGal epitope, is known as a major xenoantigen and the epitope exists abundantly in non-primate mammals. Human and Old world monkeys have anti-αGal antibody, as a natural antibody. The transfection of the functional α1-3GT gene driven by telomerase promoter into human cancer cells may lead to their transformation, making them susceptible to lysis by natural antibodies.
(1) We made the construct of the herpes simplex virus thymidine kinase gene under the control of the telomerase promoter with the
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aim of restrinting its expression to tumor cells. In transfecton experiments, the telomerase promoter driven tymidine kinase gene (hTERTp-TK) conferred ganciclovir sensitivity to leukemic cell line tested, whereas nomal somatic cells remained largely unaffected. Human hTERTp-TK positive K562 cells implanted in nude mice developed into tumors that could be eradicated by ganciclovir treatment.
(2) The fragment encompassing the whole coding sequence of bovine α1-3GT cDNA including the tanslation start site was subcloned to PEGFP basic vector which had been inserted with telomerase promoter. Bovine α1-3GT cDNA under the control of telomerase promoter (hTERTp-α1-3GT) was electrophoretically transfected into the human leukemic cell lines. Stable transfomant was obtained by selection with G418 and, limiting dilution technique. The expression of the αGal epitope was confirmed by flow cytometry, using specific binding with IB4 lectin conjugated with fluorescein isothiocyanate. I have been examining whether the leukemic cells expressing the αGal epitope can be lysed by natural antibodies and this is true for another cancer cell lines. Less
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