2000 Fiscal Year Final Research Report Summary
Targeted Gene Therapy Using Magnetic Cationic Liposomes for Osteosarcoma
| Project/Area Number |
11671435
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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 |
Orthopaedic surgery
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| Research Institution | Hiroshima University |
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Principal Investigator |
SAGITA Takashi Hiroshima Uni., Faculty of Medicine, Dept.of Orthopaedic Surgery, Associate Professor, 医学部, 助教授 (40235883)
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| Project Period (FY) |
1999 – 2000
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| Keywords | gene therapy / local targeting / magnetic cationic liposomes / p53 gene / osteosarcoma |
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| Research Abstract |
To control the growth of primary tumors effectively, we recently invented intravenously administered magnetic liposomes, which could be effectively delivered to solid tumors where a permanent magnet was implanted. Furthermore, we developed magnetic cationic liposomes as a nonviral vector to facilitate efficient DNA mediated transfection for targeted gene therapy.
Preparation of magnetic cationic liposomes : A mixture of 18 μmol 3 βN-(N'N'-dimetylaminoetane)-carbarmoyl cholesterol (DC cholesterol) and 12 μmol of dioleoil phosphatidylethanol amine (DOPE) in chloroform were evaporated by a rotary evaporator and resuspended in 1 ml sterile buffer (pH7.4) containing 10 mg magnetite particles. After hydration for 24 hours at 4℃, the organic solvent was sonicated for 10 minutes in a bath sonicator.
In vitro study : To observe the expression of transfected gene with magnetic cationic liposomes, a reporter gene containing β-galactosidase gene, pCMV SPORT-β gal was used, pCMV SPORT-β gal (100μg) was complexed with magnetic cationic liposomes in increasing ration of 10 : 1, 1 : 1, 1 : 5, and 1 : 10. The mixture was added to osteosarcoma cells (Os515) in incubation medium and osteosarcoma cells were harvested -18 hours after the incubation at 37C.The expression of pCMV SPORT-β gal was observed by western blotting in the ratio of 5 : 1 and 10 : 1.
Magnetic cationic liposomes efficiently mediated the gene transfection into osteosarcoma cells and these results suggest that magnetic cationic liposomes we developed can be a useful nonviral vector for targeted gene therapy.
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