title: Effect of albumin on human liver microsomal and recombinant CYP1A2 activities: impact on in vitro-in vivo extrapolation of drug clearance.
abstract: Long-chain unsaturated fatty acids inhibit several cytochrome P450 and UDP-glucuronosyltransferase (UGT) enzymes involved in drug metabolism, including CYP2C8, CYP2C9, UGT1A9, UGT2B4, and UGT2B7. Bovine serum albumin (BSA) enhances these cytochrome P450 and UGT activities by sequestering fatty acids that are released from membranes, especially with human liver microsomes (HLM) as the enzyme source. Here, we report the effects of BSA on CYP1A2-catalyzed phenacetin (PHEN) O-deethylation and lidocaine (LID) N-deethylation using HLM and Escherichia coli-expressed recombinant human CYP1A2 (rCYP1A2) as the enzyme sources. BSA (2% w/v) reduced (p < 0.05) the K(m) values of the high-affinity components of human liver microsomal PHEN and LID deethylation by approximately 70%, without affecting V(max). The K(m) (or S(50)) values for PHEN and LID deethylation by rCYP1A2 were reduced to a similar extent. A fatty acid mixture, comprising 3 μM concentrations each of oleic acid and linoleic acid plus 1.5 μM arachidonic acid, doubled the K(m) value for PHEN O-deethylation by rCYP1A2. Inhibition was reversed by the addition of BSA. K(i) values for the individual fatty acids ranged from 4.7 to 16.7 μM. Single-point in vitro-in vivo extrapolation (IV-IVE) based on the human liver microsomal kinetic parameters obtained in the presence, but not absence, of BSA predicted in vivo hepatic clearances of PHEN O-deethylation and LID N-deethylation that were comparable to values reported in humans, although in vivo intrinsic clearances were underpredicted. Prediction of the in vivo clearances of the CYP1A2 substrates observed here represents an improvement on other experimental systems used for IV-IVE.
authors.affiliation: Department of Pharmacology, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.
authors.forename: Nitsupa
authors.initials: N
authors.lastname: Wattanachai
authors.forename: Wichittra
authors.initials: W
authors.lastname: Tassaneeyakul
authors.forename: Andrew
authors.initials: A
authors.lastname: Rowland
authors.forename: David J
authors.initials: DJ
authors.lastname: Elliot
authors.forename: Kushari
authors.initials: K
authors.lastname: Bowalgaha
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: Knights
authors.forename: John O
authors.initials: JO
authors.lastname: Miners
mesh_terms: D000818:Animals; D002384:Catalysis; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D019388:Cytochrome P-450 CYP1A2 / Q000235:genetics / Q000378:metabolism*; D065609:Cytochrome P-450 CYP1A2 Inhibitors; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005231:Fatty Acids, Unsaturated / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008012:Lidocaine / Q000378:metabolism*; D013058:Mass Spectrometry; D008657:Metabolic Clearance Rate; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D008954:Models, Biological; D010615:Phenacetin / Q000378:metabolism*; D011237:Predictive Value of Tests; D012710:Serum Albumin, Bovine / Q000494:pharmacology*; D013379:Substrate Specificity
chemical_list: D065609:Cytochrome P-450 CYP1A2 Inhibitors; D005231:Fatty Acids, Unsaturated; D012710:Serum Albumin, Bovine; D008012:Lidocaine; C504573:CYP1A2 protein, human; D019388:Cytochrome P-450 CYP1A2; D010615:Phenacetin