| Project/Area Number |
05404040
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| Research Category |
Grant-in-Aid for General Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Research Field |
Hematology
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| Research Institution | NIIGATA UNIVERSITY |
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Principal Investigator |
SHIBATA Akira NIIGATA UNIVERSITY,SCHOOL OF MEDICINE,PROFESSOR, 医学部, 教授 (10004772)
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| Co-Investigator(Kenkyū-buntansha) |
KISHI Kenji NIIGATA UNIVERSITY,SCHOOL OF MEDICINE,FELLOW, 医学部付属病院, 助手 (30186209)
KOIKE Tadashi NIIGATA UNIVERSITY,SCHOOL OF MEDICINE,FELLOW, 医学部付属病院, 助手 (30170161)
TAKAHASHI Masuhiro NIIGATA UNIVERSITY,SCHOOL OF MEDICINE,FELLOW, 医学部, 助手 (90179531)
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| Project Period (FY) |
1993 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥11,000,000 (Direct Cost: ¥11,000,000)
Fiscal Year 1994: ¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1993: ¥8,900,000 (Direct Cost: ¥8,900,000)
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| Keywords | HOMOLOGOUS RECOMBINATION / GENE THERAPY / PGD210 / THYMIDINE KINASE / GANCICLOVIR / FDC-P2 / 脱白血病化 / BCR / ABL融合遺伝子 / 相同遺伝子組み替え / 電気パルス法 / Neo遺伝子 / HSV-tk遺伝子 |
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| Research Abstract |
The essential gene therapy for malignant neoplasias including leukemia is to normalize the tumor cells by replacing the abnormal gene with normal relevant gene. In order to repair the abnormal oncogene, the procedure of homologous recombination is inevitable. We generated the plasmid for homologous recombination and performed the preliminary study using the plasmid. The plamid for homologous recombination (pGDTKbcrNeoablreverse) was generated by reversing the direction of bcr-Neo-abl (homologous region), which was obtained from pGD210 by inserting Neo gene, in the plasmid containing TK region, in order to make the homologous region more specific. Autonomous proliferation was acquired in the IL-3 dependent mouse hematopoictic cell line FDC-P2, which was transfected with bcr-abl gene. The copy number of transfected bcr-abl was one in all clones established. G418 resistance and the susceptibility for ganciclovir were observed in the FDC-P2 cells, which were transfected with pGDTKbcrNeoablreverse and demonstrated to express the messenger of Neo and TK.The present study suggested the applicability of the plasmid for knocking out the bcr-abl oncogene in the fashion of homologous recombination, since the cells with homologous recombination were demonstrated to be successfully selected by using G418 and ganciclovir. The pivotal issue of the gene introduction in this fashion is how to increase the rate of homologous recombination in the mammalian cells. For this purpose, the new strategy such as using Rad51 will be necessary to be developed.
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