| Project/Area Number |
14370299
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Hematology
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| Research Institution | Kyushu University |
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Principal Investigator |
TANI Kenzaburo Kyushu University, Medical Institute of Bioregulation, Prof, 生体防御医学研究所, 教授 (00183864)
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| Co-Investigator(Kenkyū-buntansha) |
MUTA Hiromi Kyushu University, Medical Institute of Bioregulation, Clinic Assoc, 生体防御医学研究所, 助手 (40325478)
ASANO Shigetaka The University of Tokyo, The Institute of Medical Science, Prof, 医科学研究所, 教授 (50134614)
NAKAKUMA Hideki Wakayama Medical University, Prof, 教授 (90207746)
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| Project Period (FY) |
2002 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥14,000,000 (Direct Cost: ¥14,000,000)
Fiscal Year 2003: ¥5,400,000 (Direct Cost: ¥5,400,000)
Fiscal Year 2002: ¥8,600,000 (Direct Cost: ¥8,600,000)
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| Keywords | PNH / gene therapy / gene transduction / PIG-A / VSV-G pseudotype lentivirus vector / GPI anchor deficient K562 cells / maxizyme / random ribozyme library / 臍帯血CD34細胞 / GFP遺伝子 / 発作性夜間血色素尿症 / PIG-A遺伝子 / K562細胞株 / Musashi1 / NOD / SCID / 造血幹細胞 / マウス白血病ウィルスベクター / 水疱性口内炎ウィルスG蛋白 / K562白血病細胞 / VSV-Gシュードタイプレンチウィルスベクター |
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| Research Abstract |
Paroxysmal nocturnal hemoglobinuria(PNH) has been demonstrated to be induced by genetic mutation of phosphatidylinositol glycan-class A(PIG-A). As PNH is the disorder of pluripotent hematopoietic stem cells and characterized not only by hemolysis, but also thrombosis, easy-infectibilitu, hypoplasia, leukemogenicity and poor prognosis, the disease is considered to be the target of cell therapy and gene therapy. In this research project, we analyzed PNH from the standpoint of gene therapy. During the last 2 research years, we produced the following results. (1)To transduce PIG-A gene into human hematopoietic cells efficiently, we made a use of VSV-G pseudotype lentivirus vector(V-LV) and successfully demonstrated the efficient transduction of GFP genes into human hematopoietic stem cells. (2)We constructed PIG-A expression vector(PIG-LV) using V-LV and transduced the PIG-A gene into GPI anchor deficient K562 leukemia cells. We demonstrated the recovered expression of CD 55 and CD59. (3)After obtaining the permission of our research protocol by our IRB and the informed consents from two PNH patients, we harvested bone marrow cells from the patients and transplanted to the marrow of immunodeficient NOG(NOD/SCID/γcnull) mice to make PND model mice. (4)We transduced PIG-A gene into the hematopoietic stem cells of PNH patients in vitro using PIG-LV. (5)Recent report on PNH suggested the additional genes besides PIG-A might be involved in the pathogenesis of PNH, we have been constructing random ribozyme expressing V-LV library to determine the pathogenetic gene. (6)Using newly-developed ribozyme of maxizyme, we demonstrated that the maxizyme was expressed efficiently in human hematopoietic cells as well as leukemia cells. Also the maxizyme could express antileukemia effects by the selective destruction of bcr/abl gene, pathogenic gene of acute lymphocytic leukemia.
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