title: Stereoselectivity of the reduced folate carrier in Caco-2 cells.
abstract: Stereoselectivity of the human reduced folate carrier (RFC1) was examined in Caco-2 cells using methotrexate (l-amethopterin or l-MTX) and its antipode (d-amethopterin or d-MTX) as model substrates. The initial uptake rate of folic acid (FA) was concentration dependent, with a K(m) value of approximately 0.6 microM. The Eadie-Hofstee plot of the RFC1-mediated FA uptake revealed a single component for FA uptake into Caco-2 cells, demonstrating that only RFC1 is involved in FA uptake. l-MTX inhibited FA uptake in a competitive manner with a K(i) value of approximately 2 microM, similar to the K(m) value of l-MTX. d-MTX also competitively inhibited FA uptake with a K(i) value being approximately 120 microM, indicating that the affinity of d-MTX is ca. 60-fold less than that of l-MTX. The stereoselectivity of human RFC1 observed in the present study was consistent not only with the stereoselectivity of rabbit RFC1 observed in rabbit intestinal brush border membrane vesicles but also with the reported differences in oral absorption of amethopterin enantiomers in humans.
authors.affiliation: School of Pharmaceutical Sciences, Kitasato University, Tokyo, Japan.
authors.forename: Tomoya
authors.initials: T
authors.lastname: Narawa
authors.forename: Rikako
authors.initials: R
authors.lastname: Shimizu
authors.forename: Shuhei
authors.initials: S
authors.lastname: Takano
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Tsuda
authors.forename: Kiyoko
authors.initials: K
authors.lastname: Ono
authors.forename: Hideo
authors.initials: H
authors.lastname: Yamada
authors.forename: Tomoo
authors.initials: T
authors.lastname: Itoh
mesh_terms: D001692:Biological Transport / Q000187:drug effects*; D018938:Caco-2 Cells; D004305:Dose-Response Relationship, Drug; D005493:Folic Acid Antagonists / Q000493:pharmacokinetics* / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000378:metabolism / Q000493:pharmacokinetics*; D008727:Methotrexate / Q000493:pharmacokinetics* / Q000494:pharmacology; D058978:Reduced Folate Carrier Protein; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D005493:Folic Acid Antagonists; D026901:Membrane Transport Proteins; D058978:Reduced Folate Carrier Protein; C548699:SLC19A1 protein, human; D008727:Methotrexate