title: Binding and transport of methotrexate and its derivative, MX-68, across the brush-border membrane in rat kidney.
abstract: Binding and transport properties of methotrexate (MTX) and its novel derivative, MX-68, were examined in brush-border membrane vesicles (BBMVs) isolated from rat kidneys. The uptake of MTX, MX-68 and folic acid by BBMVs was stimulated by an inwardly-directed H(+) gradient. Such H(+)-dependent uptake of folic acid is compatible with a previous report (Bhandari et al., Biochim Biophys Acta 1988; 937: 211). The MTX uptake exhibits saturation with a K(m) of 0.834 microM. Although the uptake of these three compounds at optimal pH depended on the osmolarity of the medium, a substantial portion of the uptake was osmolarity-insensitive. By changing the medium osmolarity, the uptake by BBMVs could be separately discriminated as osmolarity-sensitive and insensitive portions, representing transport into the intravesicular space and binding to the surface of BBMVs, respectively. For all three compounds, the binding increased in a time-dependent manner, while the amount transported reached a maximum after a relatively short incubation period. The transport of folic acid, but not its binding, exhibited an overshoot phenomenon under conditions of an inward H(+) gradient. The present results suggest that reabsorption of MTX and MX-68 in the kidney is governed by both their binding and transport mechanisms, with a similar kinetic profile to that of folic acid. The involvement of a transport system seems to make a relatively small contribution to the reabsorption of MTX assessed in BBMVs compared with MX-68 and folic acid.
authors.affiliation: Graduate School of Pharmaceutical Sciences, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, Japan.
authors.forename: Y H
authors.initials: YH
authors.lastname: Han
authors.forename: Y
authors.initials: Y
authors.lastname: Kato
authors.forename: Y
authors.initials: Y
authors.lastname: Sugiyama
mesh_terms: D015074:2-Aminoadipic Acid / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D000818:Animals; D001692:Biological Transport; D004305:Dose-Response Relationship, Drug; D005492:Folic Acid / Q000493:pharmacokinetics; D005493:Folic Acid Antagonists / Q000493:pharmacokinetics*; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000493:pharmacokinetics*; D008871:Microvilli / Q000378:metabolism; D009994:Osmolar Concentration; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D005493:Folic Acid Antagonists; C104193:N-(1-((2,4-diamino-6-pteridinyl)methyl)-3,4-dihydro -2H-1,4-benzothiazine-7-carbonyl)-L-2-aminoadipic acid; D015074:2-Aminoadipic Acid; D005492:Folic Acid; D008727:Methotrexate