1995 Fiscal Year Final Research Report Summary
THE ROLE OF CYP2D SUBFAMILY IN METABOLIC ACTIVATION OF THE DRUGS THAT CONTAIN NITROGEN ATOM IN THEIR CHEMICAL STRUCTURES
| Project/Area Number |
06672166
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Biological pharmacy
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| Research Institution | CHIBA UNIVERSITY |
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Principal Investigator |
NARIMATSU Shizuo CHIBA UNIVERSITY,FACULTY OF PHARMACEUTICAL SCIENCES ASSOCIATE PROFESSOR, 薬学部, 助教授 (20113037)
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| Co-Investigator(Kenkyū-buntansha) |
MASUBUCHI Yasuhiro CHIBA UNIVERSITY,FACULTY OF PHARMACEUTICAL SCIENCES RESEARCH ASSOCIATE, 薬学部, 助手 (10209455)
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| Project Period (FY) |
1994 – 1995
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| Keywords | Human liver micosomes / Propranolol / P450-2D6 / P450-1A2 / Superoxide / 1,4-Naphthoquinone / Covalent binding / Enzyme inhibition |
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| Research Abstract |
Propranolol (PL), a beta-blocker, was oxidized to 4-hydroxylated (4-OH-PL), 5-hydroxylated (5-OH-PL) and N-desisopropylated (NDP) metabolites in human liver microsomes. The PL aromatic hydroxylations were suppressed by anti-rat cytochrome P450-2D2 (P450-2D2) antibody and its specific inhibitor (quinidine) concentration dependently. The PL N-desalkylation was decreased by alpha-naphthoflavone (a P450-1A inhibitor) and highly correlated with phenacetin O-deethylation, an index of P450-1A2 activity. These results suggest that the naphthalene ring hydroxylations and the side chain N-desalkylation were mediated by P450-2D6 and P450-1A2, respectively.
Purified rat P450-2D2 was incubated with [3H]-PL in the presence of NADPH,and relationship between radioactivity binding and decrease in enzyme activity was examined. Radioactivity bound to P450-2D2 was suppressed by anti-P450-2D2 antibody in a concentration dependent manner, and the enzyme activity was decereased time-dependently, which showed
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first-order kinetics. Radioactivity was localized at P450-2D2 apoprotein, and corresponded well to the decreased enzyme activity. From these results, it is thought that PL is biotransformed by P450-2D2 to a chemically reactive metabolite, which covalently binds to the enzyme itself, resulting in its inhibition.
4-OH-PL strongly suppressed P450-2D enzyme activity after preincubation with rat liver microsomes in the presence of NADPH.Further metabolism of 4-OH-PL was examined in rat liver microsomes, and the possibilityarose that superoxide was involved in the elimination of 4-OH-PL in the microsomes. It was found that 4-OH-PL was converted to 1,4-naphthoquinone in xanthine-xanthine oxidase system, a well known superoxide-generating system. The quinone was identified as an metabolite of 4-OH-PL also in the system containing rat liver microsomes. Taking these results and findings into account, it seems likely that PL is converted to 4-OH-PL by P450-2D enzymes in human and rat liver microsomes. 4-OH-PL thus obtained is converted by superoxide supplied from microsomal electron transfer system to 1,4-naphthoquinone, which covalently binds to P450-2D enzymes, resulting in the inhibition. Less
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