| Summary: | 碩士 === 中國醫藥大學 === 藥學系藥物安全碩士班 === 100 === Opiate addiction is an important socioeconomic issue and methadone is the milestone of treatment of opiate addiction. Interindividual variability of methadone maintenance therapy has been widely studied and P-glycoprotein (P-gp) was considered to be one of the major contributors. To investigate the mechanism of methadone and P-gp interaction, cells expressing human P-gp were utilized in the present study to exam whether methadone is a substrate or inhibitor of human P-gp.
HEK293 cells were stably transfected with human P-gp carrying common polymorphisms of human ABCB1 gene. Real-time RT-PCR was conducted to confirm the expression of human P-gp and the function of P-gp was evaluated by rhodamine123 efflux assay and calcein-AM uptake assay. Interaction between methadone and the human P-gp was examed by MDR1 shift assay and ATPase assay.
Results of the MDR1 shift assay and ATPase assay demonstrate that methadone is not a substrate of human P-gp but an ATPase stimulator. Both results of rhodamine123 efflux assay and calcein-AM uptake assay revealed that methadone is an inhibitor of human P-gp and IC50 of wild type, 1236T-2677T-3435T and 1236T-2677A-3435T P-gp were 2.17, 2.97 and 4.43μM, respectively. Further analyses of the type of inhibition demonstrated that methadone inhibited wild-type human P-gp via non-competitive inhibition, while P-gp carrying 1236T-2677T-3435T and 1236T-2677A-3435T genotypes via uncompetitive inhibition.
These results indicated that methadone may inhibit human P-gp under clinical concentration and common polymorphisms in ABCB1 gene may influence the potency and mechanism of the inhibition. Therefore, combination of methadone with drugs which are P-gp substrates may result in drug-drug interaction. Further studies regarding other polymorphisms in ABCB1 gene may provide more complete information for methadone-P-gp interaction
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