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Research ArticleArticle

Evaluation of Cryopreserved Human Hepatocytes as an Alternative in Vitro System to Microsomes for the Prediction of Metabolic Clearance

Hayley S. Brown, Michael Griffin and J. Brian Houston
Drug Metabolism and Disposition February 2007, 35 (2) 293-301; DOI: https://doi.org/10.1124/dmd.106.011569
Hayley S. Brown
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Michael Griffin
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J. Brian Houston
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Abstract

Human liver microsomes have typically resulted in marked underprediction of in vivo human intrinsic clearance (CLint); therefore, the utility of cryopreserved hepatocytes as an alternative in vitro system has become an important issue. In this study, 10 compounds (tolbutamide, diclofenac, S-warfarin, S-mephenytoin, dextromethorphan, bufuralol, quinidine, nifedipine, testosterone, and terfenadine) were selected as substrate probes for CYP2C9, 2C19, 2D6, and 3A4, and the kinetics of metabolite formation (n = 14 pathways) were investigated in three individual lots of cryopreserved hepatocytes and in a pool of human liver microsomes. For the majority of the compounds, lower unbound KM or S50 values were observed in hepatocytes compared with microsomes, on average by 50% over a 200-fold range (0.5–140 μM). Expressed on an equivalent liver weight basis, a good correlation between microsomal and hepatocyte Vmax values was observed for most pathways greater than 5 orders of magnitude (0.16–216 nmol/min/g liver). Unbound hepatocyte CLint (CLint,u) values, when scaled to the whole liver (range 0.38–4000 ml/min/kg), were on average 2.5-fold higher than microsomal CLint,u values, with the exception of tolbutamide and diclofenac, for which lower hepatocellular CLint,u values were observed. Hepatocyte predicted CLint values were compared with human in vivo CLint values, and to supplement our data, in vitro data from cryopreserved hepatocytes were collated from four other published sources. These data show that for 37 drugs, there is, on average, a 4.5-fold under-prediction of the in vivo CLint using cryopreserved hepatocytes, representing a significant reduction in prediction bias compared with human microsomes.

Footnotes

  • This work was funded by a consortium of pharmaceutical companies (GlaxoSmithKline, Novartis, Pfizer, and Servier) within the Centre for Applied Pharmacokinetic Research at the University of Manchester. M.G. was financially supported by a GlaxoSmithKline studentship. Part of this study was presented at the 9th European ISSX Meeting, June 4–7, 2006, Manchester, UK (Brown et al., 2006,Brown et al., 2006).

  • Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.

  • doi:10.1124/dmd.106.011569.

  • ABBREVIATIONS: CLint, total intrinsic clearance; CLint,u, unbound intrinsic clearance; CLmax, clearance at maximal activation; CLH, hepatic clearance; P450, cytochrome P450; UGT, UDP-glucuronosyltransferase; fub, fraction unbound in blood; fup, fraction unbound in plasma; RB, blood-to-plasma ratio; QH, hepatic blood flow; Ka, the affinity constant for the drug-protein interaction; fumic, fraction unbound in microsomal incubation; fuinc, fraction unbound in hepatocyte incubation; Kp, tissue-plasma partition coefficient; Vcell, volume of the hepatocyte; Vinc, volume of the incubation.

    • Received June 21, 2006.
    • Accepted November 17, 2006.
  • The American Society for Pharmacology and Experimental Therapeutics
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Drug Metabolism and Disposition: 35 (2)
Drug Metabolism and Disposition
Vol. 35, Issue 2
1 Feb 2007
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Research ArticleArticle

Evaluation of Cryopreserved Human Hepatocytes as an Alternative in Vitro System to Microsomes for the Prediction of Metabolic Clearance

Hayley S. Brown, Michael Griffin and J. Brian Houston
Drug Metabolism and Disposition February 1, 2007, 35 (2) 293-301; DOI: https://doi.org/10.1124/dmd.106.011569

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Research ArticleArticle

Evaluation of Cryopreserved Human Hepatocytes as an Alternative in Vitro System to Microsomes for the Prediction of Metabolic Clearance

Hayley S. Brown, Michael Griffin and J. Brian Houston
Drug Metabolism and Disposition February 1, 2007, 35 (2) 293-301; DOI: https://doi.org/10.1124/dmd.106.011569
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