title: Effect of ketocholate derivatives on methotrexate uptake in Caco-2 cell monolayers.
abstract: OBJECTIVE
  The bile salts (BS) cholate (C) and 12-monoketocholate (12-MKC) have been shown to inhibit the transcellular permeation of methotrexate (MTX) across Caco-2 cell monolayers. The aim of this study was to investigate the mechanism of this inhibition by comparing the effects of C, 7-MKC, 12-MKC, 3,7-diketocholate (DKC) and triketocholate (TKC) on MTX uptake by Caco-2 cells.
  
  
  METHODS
  Critical micelle concentrations (CMCs) and cytotoxicities of BS and their effects on membrane fluidity Caco-2 cells were determined by standard methods. MTX uptake by Caco-2 cell monolayers was determined using LC-MS/MS.
  
  
  RESULTS
  Replacing hydroxyl groups in C with keto groups and changing from 7-MKC to 12-MKC resulted in BS with lower cytotoxicity, higher CMC and decreased ability to inhibit the uptake of MTX. 7- and 12-MKC increased membrane fluidity of hydrophilic regions of Caco-2 cell membranes, DKC and TKC increased membrane fluidity of hydrophobic regions and C had little effect on membrane fluidity of either region.
  
  
  CONCLUSIONS
  Replacing hydroxyl groups in C with keto groups produces BS with different physicochemical properties and biological effects. Since ketocholates (but not C) decrease MTX uptake in parallel with increasing membrane fluidity, it is suggested that ketocholates inhibit MTX influx transporters indirectly through disturbing their lipid environment.
authors.affiliation: School of Pharmacy, University of Otago, PO Box 56, Dunedin, New Zealand. gongchenmel@gmail.com
authors.forename: Gong
authors.initials: G
authors.lastname: Chen
authors.forename: Lin
authors.initials: L
authors.lastname: Yang
authors.forename: Hu
authors.initials: H
authors.lastname: Zhang
authors.forename: Ian G
authors.initials: IG
authors.lastname: Tucker
authors.forename: J Paul
authors.initials: JP
authors.lastname: Fawcett
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000008:administration & dosage / Q000493:pharmacokinetics*; D018938:Caco-2 Cells; D020355:Cholates / Q000737:chemistry* / Q000633:toxicity; D002853:Chromatography, Liquid; D005079:Excipients / Q000737:chemistry* / Q000633:toxicity; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008560:Membrane Fluidity / Q000187:drug effects; D008727:Methotrexate / Q000008:administration & dosage / Q000493:pharmacokinetics*; D008823:Micelles; D010539:Permeability; D053719:Tandem Mass Spectrometry
chemical_list: D000964:Antimetabolites, Antineoplastic; D020355:Cholates; D005079:Excipients; D008823:Micelles; D008727:Methotrexate