title: Prediction of Drug Clearance and Drug-Drug Interactions in Microscale Cultures of Human Hepatocytes.
abstract: Accurate prediction of in vivo hepatic drug clearance using in vitro assays is important to properly estimate clinical dosing regimens. Clearance of low-turnover compounds is especially difficult to predict using short-lived suspensions of unpooled primary human hepatocytes (PHHs) and functionally declining PHH monolayers. Micropatterned cocultures (MPCCs) of PHHs and 3T3-J2 fibroblasts have been shown previously to display major liver functions for several weeks in vitro. In this study, we first characterized long-term activities of major cytochrome P450 enzymes in MPCCs created from unpooled cryopreserved PHH donors. MPCCs were then used to predict the clearance of 26 drugs that exhibit a wide range of turnover rates in vivo (0.05-19.5 ml/min per kilogram). MPCCs predicted 73, 92, and 96% of drug clearance values for all tested drugs within 2-fold, 3-fold, and 4-fold of in vivo values, respectively. There was good correlation (R(2) = 0.94, slope = 1.05) of predictions between the two PHH donors. On the other hand, suspension hepatocytes and conventional monolayers created from the same donor had significantly reduced predictive capacity (i.e., 30-50% clearance values within 4-fold of in vivo), and were not able to metabolize several drugs. Finally, we modulated drug clearance in MPCCs by inducing or inhibiting P450s. Rifampin-mediated CYP3A4 induction increased midazolam clearance by 73%, while CYP3A4 inhibition with ritonavir decreased midazolam clearance by 79%. Similarly, quinidine-mediated CYP2D6 inhibition reduced clearance of dextromethorphan and desipramine by 71 and 22%, respectively. In conclusion, MPCCs created using cryopreserved unpooled PHHs can be used for drug clearance predictions and to model drug-drug interactions.
authors.affiliation: School of Biomedical Engineering (C.L., S.R.K.) and Department of Mechanical Engineering (S.R.K.), Colorado State University, Fort Collins, Colorado; Department of Bioengineering (C.L., S.R.K.), University of Illinois at Chicago, Chicago, Illinois; Hepregen Corporation, Medford, Massachusetts (J.S., A.M.).
authors.forename: Christine
authors.initials: C
authors.lastname: Lin
authors.affiliation: School of Biomedical Engineering (C.L., S.R.K.) and Department of Mechanical Engineering (S.R.K.), Colorado State University, Fort Collins, Colorado; Department of Bioengineering (C.L., S.R.K.), University of Illinois at Chicago, Chicago, Illinois; Hepregen Corporation, Medford, Massachusetts (J.S., A.M.).
authors.forename: Julianne
authors.initials: J
authors.lastname: Shi
authors.affiliation: School of Biomedical Engineering (C.L., S.R.K.) and Department of Mechanical Engineering (S.R.K.), Colorado State University, Fort Collins, Colorado; Department of Bioengineering (C.L., S.R.K.), University of Illinois at Chicago, Chicago, Illinois; Hepregen Corporation, Medford, Massachusetts (J.S., A.M.).
authors.forename: Amanda
authors.initials: A
authors.lastname: Moore
authors.affiliation: School of Biomedical Engineering (C.L., S.R.K.) and Department of Mechanical Engineering (S.R.K.), Colorado State University, Fort Collins, Colorado; Department of Bioengineering (C.L., S.R.K.), University of Illinois at Chicago, Chicago, Illinois; Hepregen Corporation, Medford, Massachusetts (J.S., A.M.) Skhetani@uic.edu.
authors.forename: Salman R
authors.initials: SR
authors.lastname: Khetani
mesh_terms: D000328:Adult; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D065693:Cytochrome P-450 Enzyme Inducers / Q000494:pharmacology*; D065607:Cytochrome P-450 Enzyme Inhibitors / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004347:Drug Interactions; D005260:Female; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology*; D006801:Humans; D007527:Isoenzymes; D007700:Kinetics; D008297:Male; D008657:Metabolic Clearance Rate; D008875:Middle Aged; D008954:Models, Biological; D018570:Risk Assessment; D013379:Substrate Specificity; D055815:Young Adult
chemical_list: D065693:Cytochrome P-450 Enzyme Inducers; D065607:Cytochrome P-450 Enzyme Inhibitors; D007527:Isoenzymes; D003577:Cytochrome P-450 Enzyme System