The Effects of Bovine Serum Albumin on Kinetic Characterization of Human Liver Microsomal CYP2C19 and CYP2E1 Activities
Main Article Content
Abstract
The addition of bovine serum albumin (BSA) to incubations of HLM or recombinant enzymes enhances the activities of several human cytochrome P450 (CYP) and UDP-glucuronosyltransferase (UGT) enzymes, including CYP1A2, CYP2C8, CYP2C9, UGT1A8, UGT1A9, UGT1A10, UGT2A1, UGT2B4, UGT2B7, and UGT2B15. Available evidence suggests that BSA increases the activities of these drug metabolizing enzymes by sequestering polyunsaturated fatty acids that are released from membranes, especially when using human liver microsomes (HLM) as the enzyme source. Despite the importance of the ‘albumin effect’ for the accurate determination of kinetic parameters, the effects of BSA on CYP enzymes other than CYP1A2, CYP2C8, and CYP2C9 are yet to be investigated. This study aimed to investigate the effects of BSA on kinetic characterization of human liver microsomal omeprazole (OMP) 5¢-hydroxylation and chlorzoxazone (CZX) 6-hydroxlation pathways which are catalyzed by CYP2C19 and CYP2E1, respectively. When BSA (2% w/v) was added to incubations, the Km and Vmax valuesfor CYP2C19-catalyzed OMP 5¢-hydroxylation were decreased by approximately 75% and 50%, resulting in a 2-fold increase in CLint. By contrast, BSA had a minimal effect on the Km, Vmax, and CLint values for human liver microsomal CYP2E1-catalyzed CZX 6-hydroxylation. These data confirm that BSA selectively affects human liver microsomal CYP activities.
Article Details
Upon acceptance of an article, the Pharmacological and Therapeutic Society of Thailand will have exclusive right to publish and distribute the article in all forms and media and grant rights to others. Authors have rights to use and share their own published articles.
References
Boase S and Miners JO. In vitro-in vivo correlations for drugs eliminated by glucuronidation: investigations with the model substrate zidovudine. Br J Clin Pharmacol. 2002 Nov;54(5):493-503.
Riley RJ, McGinnity DF and Austin RP. A unified model for predicting human hepatic, metabolic clearance from in vitro intrinsic clearance data in hepatocytes and microsomes. Drug Metab Dispos. 2005 Sep;33(9):1304-11.
Rowland A, Elliot DJ, Knights KM, Mackenzie PI and Miners JO. The "albumin effect" and in vitro-in vivo extrapolation: sequestration of long-chain unsaturated fatty acids enhances phenytoin hydroxylation by human liver microsomal and recombinant cytochrome P450 2C9. Drug Metab Dispos. 2008 May;36(5):870-77.
Rowland A, Gaganis P, Elliot DJ, Mackenzie PI, Knights KM and Miners JO. Binding of inhibitory fatty acids is responsible for the enhancement of UDP-glucuronosyltransferase 2B7 activity by albumin: implications for in vitro-in vivo extrapolation. J Pharmacol Exp Ther. 2007 Apr;321(1):137-47.
Tsoutsikos P, Miners JO, Stapleton A, Thomas A, Sallustio BC and Knights KM. Evidence that unsaturated fatty acids are potent inhibitors of renal UDP-glucuronosyltransferases (UGT): kinetic studies using human kidney cortical microsomes and recombinant UGT1A9 and UGT2B7. Biochem Pharmacol. 2004 Jan;67(1):191-99.
Wattanachai N, Polasek TM, Heath TM, Uchaipichat V, Tassaneeyakul W, Tassaneeyakul W and Miners JO. In vitro-in vivo extrapolation of CYP2C8-catalyzed paclitaxel 6alpha-hydroxylation: effects of albumin on in vitro kinetic parameters and assessment of interindividual variability in predicted clearance. Eur J Clin Pharmacol. 2011 Feb;67(8):815–24.
Wattanachai N, Tassaneeyakul W, Rowland A, Elliot DJ, Bowalgaha K, Knights KM and Miners JO. Effect of albumin on human liver microsomal and recombinant CYP1A2 activities: impact on in vitro-in vivo extrapolation of drug clearance. Drug Metab Dispos. 2012 May;40(5):982-89.
Yamazaki H and Shimada T. Effects of arachidonic acid, prostaglandins, retinol, retinoic acid and cholecalciferol on xenobiotic oxidations catalysed by human cytochrome P450 enzymes. Xenobiotica. 1999 Mar;29(3):231-41.
Yao HT, Chang YW, Lan SJ, Chen CT, Hsu JT and Yeh TK. The inhibitory effect of polyunsaturated fatty acids on human CYP enzymes. Life Sci. 2006 Nov;79(26):2432-40.
Kilford PJ, Stringer R, Sohal B, Houston JB and Galetin A. Prediction of drug clearance by glucuronidation from in vitro data: use of combined cytochrome P450 and UDP-glucuronosyltransferase cofactors in alamethicin-activated human liver microsomes. Drug Metab Dispos. 2009 Jan;37(1):82-89.
Tang C, Lin Y, Rodrigues AD and Lin JH. Effect of albumin on phenytoin and tolbutamide metabolism in human liver microsomes: an impact more than protein binding. Drug Metab Dispos. 2002 Jun;30(6):648-54.
Rowland A, Knights KM, Mackenzie PI and Miners JO. The "albumin effect" and drug glucuronidation: bovine serum albumin and fatty acid-free human serum albumin enhance the glucuronidation of UDP-glucuronosyltransferase (UGT) 1A9 substrates but not UGT1A1 and UGT1A6 activities. Drug Metab Dispos. 2008 Jun;36(6):1056-62.
Raungrut P, Uchaipichat V, Elliot DJ, Janchawee B, Somogyi AA and Miners JO. In vitro-in vivo extrapolation predicts drug-drug interactions arising from inhibition of codeine glucuronidation by dextropropoxyphene, fluconazole, ketoconazole, and methadone in humans. J Pharmacol Exp Ther. 2010 Aug;334(2):609-18.
Rowland A, Knights KM, Mackenzie PI and Miners JO. Characterization of the binding of drugs to human intestinal fatty acid binding protein (IFABP): potential role of IFABP as an alternative to albumin for in vitro-in vivo extrapolation of drug kinetic parameters. Drug Metab Dispos. 2009 Jul;37(7):1395-403.
Manevski N, Troberg J, Svaluto-Moreolo P, Dziedzic K, Yli-Kauhaluoma J and Finel M. Albumin stimulates the activity of the human UDP-glucuronosyltransferases 1A7, 1A8, 1A10, 2A1 and 2B15, but the effects are enzyme and substrate dependent. PLoS ONE. 2013 Feb;8(1):e54767.
Rowland A, Elliot DJ, Williams JA, Mackenzie PI, Dickinson RG and Miners JO. In vitro characterization of lamotrigine N2-glucuronidation and the lamotrigine-valproic acid interaction. Drug Metab Dispos. 2006 Jun;34(6):1055-62.
Andersson T, Miners JO, Veronese ME, Tassaneeyakul W, Meyer UA and Birkett DJ. Identification of human liver cytochrome P450 isoforms mediating omeprazole metabolism. Br J Clin Pharmacol. 1993 Dec;36(6):521-30.
Tassaneeyakul W, Vannaprasaht S and Yamazoe Y. Formation of omeprazole sulphone but not 5-hydroxyomeprazole is inhibited by grapefruit juice. Br J Clin Pharmacol. 2000 Feb;49(2):139-44.
Chiba K, Kobayashi K, Manabe K, Tani M, Kamataki T and Ishizaki T. Oxidative metabolism of omeprazole in human liver microsomes: cosegregation with S-mephenytoin 4'-hydroxylation. J Pharmacol Exp Ther. 1993 Jul;266(1):52-59.
Karam WG, Goldstein JA, Lasker JM and Ghanayem BI. Human CYP2C19 is a major omeprazole 5-hydroxylase, as demonstrated with recombinant cytochrome P450 enzymes. Drug Metab Dispos. 1996 Oct;24(10):1081-87.
Peter R, Bocker R, Beaune PH, Iwasaki M, Guengerich FP and Yang CS. Hydroxylation of chlorzoxazone as a specific probe for human liver cytochrome P-450IIE1. Chem Res Toxicol. 1990 Nov;3(6):566-73.
Tassaneeyakul W, Birkett DJ, McManus ME, Tassaneeyakul W, Veronese ME, Andersson T, Tukey RH and Miners JO. Caffeine metabolism by human hepatic cytochromes P450: contributions of 1A2, 2E1 and 3A isoforms. Biochem Pharmacol. 1994 May;47(10):1767-76.
McLure JA, Miners JO and Birkett DJ. Nonspecific binding of drugs to human liver microsomes. Br J Clin Pharmacol. 2000 May;49(5):453-61.
Li XQ, Weidolf L, Simonsson R and Andersson TB. Enantiomer/enantiomer interactions between the S- and R- isomers of omeprazole in human cytochrome P450 enzymes: major role of CYP2C19 and CYP3A4. J Pharmacol Exp Ther. 2005 Nov;315(2):777-87.
Shu Y, Wang LS, Xu ZH, He N, Xiao WM, Wang W, Huang SL and Zhou HH. 5-hydroxylation of omeprazole by human liver microsomal fractions from Chinese populations related to CYP2C19 gene dose and individual ethnicity. J Pharmacol Exp Ther. 2000 Nov;295(2):844-51.
Manevski N, Svaluto Morelo P, Yli-Kauhaluoma J and Finel M. Bovine Serum Albumin Decreases the Km Values of the Human UDP-glucuronosyltransferases 1A9 and 2B7, but only in UGT1A9 it also largely increases the Vmax value. Drug Metab Dispos. 2011 Aug;39(11):2117-29.
Xu BQ, Ishii M, Ding LR, Fischer NE and Inaba T. Interaction of serum proteins with CYP isoforms in human liver microsomes: inhibitory effects of human and bovine albumin, alpha-globulins, alpha-1-acid glycoproteins and gamma-globulins on CYP2C19 and CYP2D6. Life Sci. 2003 Mar;72(17):1953-62.
Draper AJ, Madan A, Latham J and Parkinson A. Development of a non-high pressure liquid chromatography assay to determine [14C]chlorzoxazone 6-hydroxylase (CYP2E1) activity in human liver microsomes. Drug Metab Dispos. 1998 Apr;26(4):305-12.
Kim RB, Yamazaki H, Chiba K, O'Shea D, Mimura M, Guengerich FP, Ishizaki T, Shimada T and Wilkinson GR. In vivo and in vitro characterization of CYP2E1 activity in Japanese and Caucasians. J Pharmacol Exp Ther. 1996 Oct;279(1):4-11.
Uchaipichat V, Winner LK, Mackenzie PI, Elliot DJ, Williams JA and Miners JO. Quantitative prediction of in vivo inhibitory interactions involving glucuronidated drugs from in vitro data: the effect of fluconazole on zidovudine glucuronidation. Br J Clin Pharmacol. 2006 Apr;61(4):427-39.
Porubsky PR, Battaile KP and Scott EE. Human cytochrome P450 2E1 structures with fatty acid analogs reveal a previously unobserved binding mode. J Biol Chem. 2010 Jul;285(29):22282-90.
Matsumoto S, Ding LR, Ishii M, Fischer NE and Inaba T. The interaction of human and bovine serum proteins with CYP3A in human liver microsomes: inhibition of testosterone 6beta-hydroxylation by albumin, alpha-globulins, alpha(1)-acid glycoprotein and gamma-globulins. Toxicol Lett. 2002 Nov;136(1):33-41.