Abstract
In this study we have evaluated the application and reliability of using fluorescence (FLUO)-based high throughput screening assays with recombinant CYPs (rCYP). This was accomplished by screening 29 clinically important antiparasitic drugs for inhibition of the five major drug-metabolizing CYPs (-1A2, -2C9, -2C19, -2D6, and -3A4). Data from FLUO/rCYP assays were compared with that obtained by conventional HPLC assays using human liver microsomes (HLM) and rCYPs. TheKi values showed good correlations: FLUO/rCYP and HPLC/rCYP (r2 = 0.81), HPLC/rCYP and HPLC/HLM (r2 = 0.82), and FLUO/rCYP and HPLC/HLM (r2 = 0.72). Niclosamide had substrate-dependent contrasting effects on CYP2C9 activity with an apparent activation (400%) of 7-methoxy-4-trifluoromethylcoumarin demethylase activity and potent inhibition (Ki = 6.00 μM) of diclofenac 4-hydroxylase activity. Potent inhibitors of CYP1A2 were artemisinin, dihydroartemisinin, thiabendazole, primaquine, and niclosamide (Ki = 0.43, 3.67, 1.54, 0.22, and 2.70 μM, respectively). Proguanil, cycloguanil, amodiaquine, and desethylamodiaquine inhibited CYP2D6 (Ki = 6.76, 5.97, 2.1, and 4.13 μM, respectively). Considering the Cmax of these drugs, artemisinin, thiabendazole, primaquine, amodiaquine, and desethylamodiaquine may cause clinically important interactions because they are predicted to inhibit 67 to 99% of the activities of the CYPs they interact with. In addition, our results suggest CYP1A2 inhibition as the mechanism behind the observed thiabendazole/theophylline and primaquine/antipyrine interactions in vivo.
Footnotes
-
Send reprint requests to: Dr. Collen Mutowembwa Masimirembwa, Department of DMPK and Bioanalytical Chemistry, AstraZeneca, R & D Mölndal, S-431 83 Mölndal, Sweden. E-mail: collen.masimirembwa{at}astrazeneca.com
-
Tashinga Bapiro is a recipient of a fellowship from International Programme in Chemical Sciences, Uppsala University, Sweden.
- Abbreviations used are::
- CYP cytochrome P450
- HPLC, high-performance liquid chromatography
- HTS
- high throughput screening
- rCYP
- recombinant cytochrome P450
- FLUO
- fluorescence
- HLM
- human liver microsome
- MFC
- 7-methoxy-4-trifluoromethylcoumarin
- Received June 30, 2000.
- Accepted October 17, 2000.
- The American Society for Pharmacology and Experimental Therapeutics
DMD articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|