| Project/Area Number |
10670241
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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 |
寄生虫学(含医用動物学)
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| Research Institution | Gunma University, School of Medicine |
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Principal Investigator |
KATAKURA Ken Gunma University School of Medicine, Department of Parasitology Associate Professor, 医学部, 助教授 (10130155)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 1999: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1998: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | 5-fluorouracil / ABC protein / anti-cancer drugs / drug accumulation / Leishmania amazonensis / multidrug resistance (MDR) / P-glycoprotein / transporter / リーシュマニア / MDR / 寄生適応 / 遺伝子導入 |
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| Research Abstract |
We isolated and characterized two different P-glycoprotein genes in a parasitic protozoan, Leishmania amazonensis. The LaMDR1 gene encodes 1341 amino acids for a protein consisting of two similar halves, each containing one ATP-binding and six transmembrane domains. Transfection and over-expression of the LaMDR1 gene in wild-type promastigotes of L. amazonensis conferred resistance to vinblastine, doxorubicin and actinomycin D, indicating that the LaMDR1 gene is a homologue of human multidrung resistance MDR1 gene. The LaMDR2 gene encodes 1267 amino acids, revealing only 47% amino acid identity to the LaMDR1 protein. Transfectants with the LaMDR2 gene showed resistance to 5-fluorouracil (5-FU), a fluoropyrimidine used for cancer chemotherapy, but not to other anti-cancer drugs described above. A slightly higher intracellular accumulation of [ィイD13ィエD1H] vinblastine was observed in the LaMDR1-transfected cells compared with control transfectant cells. In transfectants with the LaMDR2 gene, an increased accumulation of [ィイD13ィエD1H]5-FU was measured in the first 30 min after drug exposure, but the drug accumulation was gradually decreased and reached to 20-30% of the control level at 120 min after the exposure. This 5-FU accumulation kinetics in transfectants with the LaMDR2 gene suggests that the LaMDR2 protein may locate on the membrane of cytoplasmic vesicles and transport 5-FU inside the vesicles during the initial phase, and then drugs were secreted by exocytosis during the later phase. These results revealed that the LaMDR2 gene is a new member of the ATP-binding cassette (ABC) family of protein with the P-glycoprotein structure and play a novel role in survival of the parasites in the host cells. Subcellular localization of the LaMDR2 protein using anti-LaMDR2 antibodies is under study. Functional analysis of the LaMDR2 gene by gene-disruption technology is also in process.
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