Abstract
We tested the hypothesis that primary cultures of human hepatocytes could predict potential drug interactions with methadone and buprenorphine. Hepatocytes (five donors) were preincubated with dimethyl sulfoxide (DMSO) (vehicle), rifampin, or nelfinavir before incubation with methadone or buprenorphine. Culture media (0–60 min) was analyzed by liquid chromatography-tandem mass spectrometry for R- and S-methadone and R- and S-2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) or for buprenorphine, norbuprenorphine, and their glucuronides [buprenorphine-3-glucuronide (B-3-G) and norbuprenorphine-3-glucuronide (N-3-G)]. R- and S-EDDP were detected in three of five, four of five, and five of five media from cells pretreated with DMSO, nelfinavir, and rifampin. R-EDDP increased 3.1- and 26.5-fold, and S-EDDP increased 2.5- and 21.3-fold after nelfinavir and rifampin. The rifampin effect was significant. B-3-G production was detected in media of all cells incubated with buprenorphine and accounted for most of the buprenorphine loss from culture media; it was not significantly affected by either pretreatment. Norbuprenorphine and N-3-G together were detected in three of five, four of five, and five of five donors pretreated with DMSO, nelfinavir and rifampin, and norbuprenorphine in one of five, one of five, and two of five donors. Although there was a trend for norbuprenorphine (2.8- and 4.9-fold) and N-3-G (1.7- and 1.9-fold) to increase after nelfinavir and rifampin, none of the changes were significant. To investigate low norbuprenorphine production, buprenorphine was incubated with human liver and small intestine microsomes fortified to support both N-dealkylation and glucuronidation; N-dealkylation predominated in small intestine and glucuronidation in liver microsomes. These studies support the hypothesis that methadone metabolism and its potential for drug interactions can be predicted with cultured human hepatocytes, but for buprenorphine the combined effects of hepatic and small intestinal metabolism are probably involved.
Footnotes
This work was supported in part by the National Institutes of Health National Institute on Drug Abuse [Grant RO1-DA10100]; and the National Institutes of Health National Institute of General Medical Sciences [Grant GM066411]. Normal human hepatocytes were obtained through the Liver Tissue Cell Distribution System, Pittsburgh, PA, which was funded by the National Institutes of Health [Contract N01-DK-7–004/HHSN267200700004C]. For projects unrelated to the current study, D.E.M. has received research funding and consultant fees from Reckitt Benckiser, the manufacturer of buprenorphine.
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.109.028605
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- AUC
- area under the time-concentration curve
- EDDP
- 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine
- P450
- cytochrome P450
- UGT
- UDP-glucuronosyl transferase
- HLM
- human liver microsomes
- HSIM
- human small intestine microsomes
- UDPGA
- uridine 5′-diphosphoglucuronic acid
- DMSO
- dimethylsulfoxide
- LC-ESI-MS/MS
- liquid chromatography-electrospray ionization-tandem mass spectrometry
- QC
- quality control sample
- AUIC
- area under the incubation time-concentration curve.
- Received May 27, 2009.
- Accepted September 17, 2009.
- Copyright © 2009 by The American Society for Pharmacology and Experimental Therapeutics
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