| Co-Investigator(Kenkyū-buntansha) |
IKEDA Teruo Azabu University School of Veterinary Medicine, Assistant Professor, 獣医学部, 助教授 (60151297)
KATAKURA Ken Gunma University School of Medicine, Assistant Professor, 医学部, 助教授 (10130155)
OCHIAI Hideharu Azabu University School of Veterinary Medicine, Instructor, 生物科学総合研究所, 助手 (20247307)
INOMATA Tomoo Azabu University School of Veterinary Medicine, Assistant Professor, 獣医学部, 助教授 (10147978)
SHIROTA Kinji Azabu University School of Veterinary Medicine, Professor, 生物科学総合研究所, 教授 (70147974)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2001: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2000: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1999: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Research Abstract |
Though P-glycoprotein (PGP) which transport coplanar PGB (Co-PCB) has not been found, it was revealed that Co-PCB inhibited the function of PGP. Followings were the findings in this experiment. Drug transport pump, mdr1 and mdr2, were cloned from leishmania amazonensis (La) ; La-mdr1 exerted multi drug resistance, and La-mdr2 transported 5FU. Drug accumulation was examined in the transformant human carcinoma cell, KB3-1, expressed human PGP. Drug accumulation was decreased in the KB3-1 cells expressing wild PGP (KB3-His^<61>) and mutant PGP in which the 61st amino acid was substituted for serine (KB3-Ser^<61>) or phenylalanine (KB3-Phe^<61>), compared to wild KB3-1. Drug tolerance and transport ability for small molecule chemical, colchicine, were greater in KB3-Phe^<61> compared to KB3-Ser^<61>, thus inverse correlation was indicated between the bulk of the side chain of 61st amino acid and the molecular weight of the substrates. Mutant human PGP gene, in which His61 was substituted with Ser^<61> or Phe^<61>, were transfected in porcine kidney cell, LLC-PK1 , and prepared transformant cells, LLC-His^<61>, LLC-Phe^<61> and LLC-Ser^<61>. In the transformant cells, the abilities of the drug tolerance and the decrease of the accumulation were same as in PGP expressing KB3-1 cells, and transepithelial transport was increased. Co-PCB was not transported through the epithelial monolayer in the PGP expressing LLC-PK1. However, the most toxic congenor of Co-PCBs, 3, 3', 4, 4', 5-pentachlorobiphenyl, inhibited transport of the other drugs by PGP, and the inhibiton was remarkable in KB3-Phe^<61> and LLC-Phe^<61>. Drug tolerance cell lines (100 times tolerance) were selected from canine tumor cells. In the cells, clear appearance of the protein and mRNA of PGP were detected by western blot and RT-PCR, respectively.
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