title: Towards prediction of in vivo intestinal absorption using a 96-well Caco-2 assay.
abstract: We systematically validated a robust 96-well Caco-2 assay via an extended set of 93 marketed drugs with diverse transport mechanisms and quantified by LC/MS/MS, to investigate its predictive utility while dealing with challenging discovery compounds. Utilizing nonlinear fit, the validation led to a good correlation (R(2) = 0.76) between absorptive permeability, log P(app)(A-B), from in vitro Caco-2 assay and reported human fraction of dose absorbed. We observed that paracellular compounds could be flagged by log P(app)(A-B) (<-5.5 cm/s) and physicochemical property space (c log P < 1). Of 8000 Novartis discovery compounds examined 13% were subject to low recovery (<30%). Compound loss was investigated by comparing cell monolayer and artificial membrane, while 0.5% bovine serum albumin (in both donor and acceptor compartments) was utilized to improve recovery. The second focus of this study was to investigate the advantages and limitations of the current Caco-2 assay for predicting in vivo intestinal absorption. Caco-2 measurements for compounds with high aqueous solubility and low in vitro metabolic clearance were compared to 88 in vivo rat bioavailability studies. Despite the challenges posed by discovery compounds with suboptimal physicochemical properties, Caco-2 data successfully projected low intestinal absorption. This platform sets the stage for mechanistically evaluating compounds towards improving in vitro-in vivo correlations.
authors.affiliation: Metabolism and Pharmacokinetics Platform, Novartis Institutes for BioMedical Research, 250 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Skolnik
authors.forename: Xuena
authors.initials: X
authors.lastname: Lin
authors.forename: Jianling
authors.initials: J
authors.lastname: Wang
authors.forename: Xiao-Hui
authors.initials: XH
authors.lastname: Chen
authors.forename: Timothy
authors.initials: T
authors.lastname: He
authors.forename: Bailin
authors.initials: B
authors.lastname: Zhang
mesh_terms: D018938:Caco-2 Cells; D002463:Cell Membrane Permeability; D006801:Humans; D007408:Intestinal Absorption; D008954:Models, Biological; D010599:Pharmacokinetics