title: Intrinsic Clearance Assay Incubational Binding: A Method Comparison.
abstract: The fraction of unbound drug (fuinc) in in vitro intrinsic clearance (CLint) incubation is an important parameter in the pursuit of accurate clearance predictions and is often predicted using algorithms based on drug lipophilicity measures. However, analysis of an AstraZeneca database suggests that simple lipophilicity alone is a relatively poor predictor of fuincmeasured using equilibrium dialysis. He fuincvalue can also be measured directly in CLintassays using multiple concentrations of hepatocytes or microsomal protein. Since this approach informs of the unbound drug concentration in the assay used to predict in vivo clearance, it should be considered the gold standard method. As a starting point for building better predictive algorithms we aimed to determine if equilibrium dialysis really is an appropriate assay for assessing fuincEmploying a large number of compounds with a wide range of lipophilicities, experiments were performed to measure fuincusing rat hepatocytes (RH) and human liver microsomes (HLM) in both assay formats. A high percentage (94% and 93% for HLM and RH, respectively) of the fuincvalues were within 2-fold when the compound distribution coefficient describing the ratio of compound concentration in octanol and pH 7.4 buffer when the test system is at equilibrium (lipophilicity measure) (logD7.4) values were less than 3.5. However, with logD7.4values greater than these, the agreement was considerably worse. Additional experimental data generated indicated that this discrepancy was likely due to failings in the direct method when drug binding is high. Thus, we conclude that unbound CLintcan be indeed calculated indirectly by incorporating equilibrium dialysis data with measured CLintbut that simple lipophilicity descriptors alone may be inadequate for predicting fuinc.
authors.affiliation: Respiratory, Inflammation and Autoimmunity Department of Drug Metabolism and Pharmacokinetics (K.G., S.C., L.P.G.), Drug Safety and Metabolism (P.N.), and Cardiovascular and Metabolic Diseases Department of Drug Metabolism and Pharmacokinetics (F.B.), Innovative Medicines and Early Development, AstraZeneca, Gothenburg, Sweden.
authors.forename: Sofia
authors.initials: S
authors.lastname: Chen
authors.affiliation: Respiratory, Inflammation and Autoimmunity Department of Drug Metabolism and Pharmacokinetics (K.G., S.C., L.P.G.), Drug Safety and Metabolism (P.N.), and Cardiovascular and Metabolic Diseases Department of Drug Metabolism and Pharmacokinetics (F.B.), Innovative Medicines and Early Development, AstraZeneca, Gothenburg, Sweden.
authors.forename: Luna
authors.initials: L
authors.lastname: Prieto Garcia
authors.affiliation: Respiratory, Inflammation and Autoimmunity Department of Drug Metabolism and Pharmacokinetics (K.G., S.C., L.P.G.), Drug Safety and Metabolism (P.N.), and Cardiovascular and Metabolic Diseases Department of Drug Metabolism and Pharmacokinetics (F.B.), Innovative Medicines and Early Development, AstraZeneca, Gothenburg, Sweden.
authors.forename: Fredrik
authors.initials: F
authors.lastname: Bergström
authors.affiliation: Respiratory, Inflammation and Autoimmunity Department of Drug Metabolism and Pharmacokinetics (K.G., S.C., L.P.G.), Drug Safety and Metabolism (P.N.), and Cardiovascular and Metabolic Diseases Department of Drug Metabolism and Pharmacokinetics (F.B.), Innovative Medicines and Early Development, AstraZeneca, Gothenburg, Sweden.
authors.forename: Pär
authors.initials: P
authors.lastname: Nordell
authors.affiliation: Respiratory, Inflammation and Autoimmunity Department of Drug Metabolism and Pharmacokinetics (K.G., S.C., L.P.G.), Drug Safety and Metabolism (P.N.), and Cardiovascular and Metabolic Diseases Department of Drug Metabolism and Pharmacokinetics (F.B.), Innovative Medicines and Early Development, AstraZeneca, Gothenburg, Sweden ken.grime@astrazeneca.com.
authors.forename: Ken
authors.initials: K
authors.lastname: Grime
mesh_terms: D000465:Algorithms; D000818:Animals; D003956:Dialysis / Q000379:methods; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D008657:Metabolic Clearance Rate; D008862:Microsomes, Liver / Q000378:metabolism*; D008954:Models, Biological; D004364:Pharmaceutical Preparations / Q000737:chemistry; D010599:Pharmacokinetics; D011485:Protein Binding; D051381:Rats
chemical_list: D004364:Pharmaceutical Preparations