title: Development, validation, and application of a novel 7-day Caco-2 cell culture system.
abstract: BACKGROUND
  Monolayers of Caco-2 cells have been widely accepted as one of the well-established in vitro models to predict intestinal drug permeability and absorption in humans. However, the procedure for culturing the traditional 21-day Caco-2 model is labor intensive and time consuming, which limits its wide application in drug development. The objective of the present study was to develop a rapid Caco-2 model with a 7-day cell culture process.
  
  
  METHODS
  A few modifications of the BIOCOAT® HTS Caco-2 Assay System were introduced including changing the cell seeding density, the composition of cell culture media, and the interval to change media. The monolayer structure was visualized through confocal microscopy. The transepithelial electrical resistance (TEER), apparent permeability coefficients (Papp), and functional activity of P-glycoprotein (P-gp) were determined and compared with the 21-day model. Moreover, pharmacokinetic studies were performed in animal models to evaluate the absolute bioavailability of oral doses (Foral) of different compounds.
  
  
  RESULTS
  Our newly developed 7-day Caco-2 model displayed comparable cellular morphology and integrity with the traditional 21-day model. No significant difference in paracellular and transcellular permeability was observed between the two systems. The efflux ratios of transporting digoxin, the prototypical substrate of P-gp in 21- and 7-day systems were 16.04 and 24.92, respectively. When the P-gp inhibitor verapamil was present, the efflux ratios of 21- and 7-day systems were 1.37 and 0.86, respectively, suggesting the comparability of the P-gp functional activity in both systems. Furthermore, pharmacokinetic studies of several compounds performed in animal models revealed that the absolute bioavailability of oral doses in vivo was well correlated with the Caco-2 permeability in vitro.
  
  
  CONCLUSIONS
  The novel system provides a rapid and economical option for assessing the drug permeability, and is applicable to the studies of intestinal drug absorption.
authors.affiliation: Laboratory of Drug Metabolism and Pharmacokinetics, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, Guangdong 510006, China; Guangdong Food and Drug Administration Center for Evaluation and Certification, Guangzhou, Guangdong 510080, China.
authors.forename: Yike
authors.initials: Y
authors.lastname: Cai
authors.affiliation: Department of Pharmacy, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong 510080, China.
authors.forename: Chenshu
authors.initials: C
authors.lastname: Xu
authors.affiliation: Guangdong Food and Drug Administration Center for Evaluation and Certification, Guangzhou, Guangdong 510080, China.
authors.forename: Peiyi
authors.initials: P
authors.lastname: Chen
authors.affiliation: Laboratory of Drug Metabolism and Pharmacokinetics, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, Guangdong 510006, China.
authors.forename: Jinqing
authors.initials: J
authors.lastname: Hu
authors.affiliation: Laboratory of Drug Metabolism and Pharmacokinetics, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, Guangdong 510006, China.
authors.forename: Rong
authors.initials: R
authors.lastname: Hu
authors.affiliation: Laboratory of Drug Metabolism and Pharmacokinetics, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, Guangdong 510006, China.
authors.forename: Min
authors.initials: M
authors.lastname: Huang
authors.affiliation: Laboratory of Drug Metabolism and Pharmacokinetics, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, Guangdong 510006, China. Electronic address: bihchang@mail.sysu.edu.cn.
authors.forename: Huichang
authors.initials: H
authors.lastname: Bi
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D000284:Administration, Oral; D000818:Animals; D001682:Biological Availability; D018938:Caco-2 Cells; D018929:Cell Culture Techniques / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D004077:Digoxin / Q000008:administration & dosage / Q000493:pharmacokinetics; D004285:Dogs; D006801:Humans; D007408:Intestinal Absorption / Q000187:drug effects; D010539:Permeability; D053719:Tandem Mass Spectrometry; D014700:Verapamil / Q000008:administration & dosage / Q000493:pharmacokinetics
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D004077:Digoxin; D014700:Verapamil
keywords: 7-day culture system; Absorption; Caco-2 cells; P-Glycoprotein; Permeability