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Metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin by human hepatic CYP isoforms: evidence for selectivity towards CYP3A4.
Xenobiotica 2001; 31(4):187-204X

Abstract

1. The metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin (BFBFC) to 7-hydroxy-4-trifluoromethylcoumarin (HFC) was studied in human liver microsomes and in cDNA-expressed human liver CYP isoforms. For purposes of comparison, some limited studies were also performed with 7-benzyloxyquinoline (7BQ). 2. Initial interactive docking studies with a homology model of human CYP3A4 indicated that BFBFC was likely to be a selective substrate for CYP3A4 with a relatively high binding affinity, due to the presence of several key hydrogen bonds with active site amino acid residues. 3. Kinetic analysis of NADPH-dependent BFBFC metabolism to HFC in three preparations of pooled human liver microsomes revealed mean (+/- TSEM) Km and Vmax = 4.6 +/- 0.3 microM and 20.0 +/- 3.8 pmol/min/mg protein, respectively. 4. The metabolism of BFBFC to HFC was determined in a characterized bank of 24 individual human liver microsomal preparations employing a BFBFC substrate concentration of lO microM (i.e. around twice Km). Good correlations (r2 = 0.736-0.904) were observed between BFBFC metabolism and markers of CYP3A isoforms. 5. While 10O microM BFBFC was metabolized to HFC by cDNA-expressed CYP3A4, little or no metabolism was observed with cDNA-expressed CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP2E1. 6. The metabolism of 10 microM BFBFC in human liver microsomes was markedly inhibited by 5-50 microM troleandomycin and 0.2-5 microM ketoconazole, but stimulated by 0.2-10 microM alpha-naphthoflavone. The metabolism of 10 microM BFBFC in human liver microsomes was also markedly inhibited by an antibody to CYP3A4. 7. Kinetic analysis of NADPH-dependent 7BQ metabolism to 7-hydroxyquinoline (7HQ) in human liver microsomes revealed Km and Vmax = 70 microM and 3.39 nmol/min/mg protein, respectively. 8. While 80 microM 7BQ was metabolized to 7HQ by cDNA-expressed CYP3A4, only low rates of metabolism were observed with cDNA-expressed CYPIA2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP2E1. 9. In summary, by correlation analysis, the use of cDNA-expressed CYP isoforms, chemical inhibition and inhibitory antibodies, BFBFC metabolism in human liver microsomes appears to be primarily catalysed by CYP3A4. BFBFC may be a useful fluorescent probe substrate for human hepatic CYP3A4, but compared with 7BQ has only a low rate of metabolism in human liver microsomes.

Authors+Show Affiliations

TNO BIBRA International Ltd, Carshalton, Surrey, UK.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

11465405

Citation

Renwick, A B., et al. "Metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin By Human Hepatic CYP Isoforms: Evidence for Selectivity Towards CYP3A4." Xenobiotica; the Fate of Foreign Compounds in Biological Systems, vol. 31, no. 4, 2001, pp. 187-204.
Renwick AB, Lewis DF, Fulford S, et al. Metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin by human hepatic CYP isoforms: evidence for selectivity towards CYP3A4. Xenobiotica. 2001;31(4):187-204.
Renwick, A. B., Lewis, D. F., Fulford, S., Surry, D., Williams, B., Worboys, P. D., ... Evans, D. C. (2001). Metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin by human hepatic CYP isoforms: evidence for selectivity towards CYP3A4. Xenobiotica; the Fate of Foreign Compounds in Biological Systems, 31(4), pp. 187-204.
Renwick AB, et al. Metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin By Human Hepatic CYP Isoforms: Evidence for Selectivity Towards CYP3A4. Xenobiotica. 2001;31(4):187-204. PubMed PMID: 11465405.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin by human hepatic CYP isoforms: evidence for selectivity towards CYP3A4. AU - Renwick,A B, AU - Lewis,D F, AU - Fulford,S, AU - Surry,D, AU - Williams,B, AU - Worboys,P D, AU - Cai,X, AU - Wang,R W, AU - Price,R J, AU - Lake,B G, AU - Evans,D C, PY - 2001/7/24/pubmed PY - 2002/1/18/medline PY - 2001/7/24/entrez SP - 187 EP - 204 JF - Xenobiotica; the fate of foreign compounds in biological systems JO - Xenobiotica VL - 31 IS - 4 N2 - 1. The metabolism of 2,5-bis(trifluoromethyl)-7-benzyloxy-4-trifluoromethylcoumarin (BFBFC) to 7-hydroxy-4-trifluoromethylcoumarin (HFC) was studied in human liver microsomes and in cDNA-expressed human liver CYP isoforms. For purposes of comparison, some limited studies were also performed with 7-benzyloxyquinoline (7BQ). 2. Initial interactive docking studies with a homology model of human CYP3A4 indicated that BFBFC was likely to be a selective substrate for CYP3A4 with a relatively high binding affinity, due to the presence of several key hydrogen bonds with active site amino acid residues. 3. Kinetic analysis of NADPH-dependent BFBFC metabolism to HFC in three preparations of pooled human liver microsomes revealed mean (+/- TSEM) Km and Vmax = 4.6 +/- 0.3 microM and 20.0 +/- 3.8 pmol/min/mg protein, respectively. 4. The metabolism of BFBFC to HFC was determined in a characterized bank of 24 individual human liver microsomal preparations employing a BFBFC substrate concentration of lO microM (i.e. around twice Km). Good correlations (r2 = 0.736-0.904) were observed between BFBFC metabolism and markers of CYP3A isoforms. 5. While 10O microM BFBFC was metabolized to HFC by cDNA-expressed CYP3A4, little or no metabolism was observed with cDNA-expressed CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP2E1. 6. The metabolism of 10 microM BFBFC in human liver microsomes was markedly inhibited by 5-50 microM troleandomycin and 0.2-5 microM ketoconazole, but stimulated by 0.2-10 microM alpha-naphthoflavone. The metabolism of 10 microM BFBFC in human liver microsomes was also markedly inhibited by an antibody to CYP3A4. 7. Kinetic analysis of NADPH-dependent 7BQ metabolism to 7-hydroxyquinoline (7HQ) in human liver microsomes revealed Km and Vmax = 70 microM and 3.39 nmol/min/mg protein, respectively. 8. While 80 microM 7BQ was metabolized to 7HQ by cDNA-expressed CYP3A4, only low rates of metabolism were observed with cDNA-expressed CYPIA2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP2E1. 9. In summary, by correlation analysis, the use of cDNA-expressed CYP isoforms, chemical inhibition and inhibitory antibodies, BFBFC metabolism in human liver microsomes appears to be primarily catalysed by CYP3A4. BFBFC may be a useful fluorescent probe substrate for human hepatic CYP3A4, but compared with 7BQ has only a low rate of metabolism in human liver microsomes. SN - 0049-8254 UR - https://www.unboundmedicine.com/medline/citation/11465405/Metabolism_of_25_bis_trifluoromethyl__7_benzyloxy_4_trifluoromethylcoumarin_by_human_hepatic_CYP_isoforms:_evidence_for_selectivity_towards_CYP3A4_ L2 - http://www.tandfonline.com/doi/full/10.1080/00498250110043526 DB - PRIME DP - Unbound Medicine ER -