Carrier-mediated hepatobiliary transport of a novel antifolate, N-[4-[(2,4-dianninopteridine-6-yl)methyl]-3,4-dihydro-2H-1,4-benzothiazin-7-yl]carbonyl-L-homoglutamic acid, in rats.
The hepatic uptake and biliary excretion of a novel methotrexate derivative, N-[4-[(2,4-diamminopteridine-6-yl)methyl]-3,4-dihydro-2H-1,4-benzothiazin-7-yl]carbonyl-L-homoglutamic acid (MX-68), were examined in rats in vitro using isolated hepatocytes and bile canalicular membrane vesicles (CMVs), respectively. The uptake of MX-68 by isolated rat hepatocytes showed a saturable component (Km = 2.15 microM and Vmax = 2.34 pmol/min/mg of protein) and was inhibited by ATP-depletors and anionic compounds such as taurocholate and probenecid. [3H]MX-68 uptake was also inhibited by folate analogs such as methotrexate and 5CH3-tetrahydrofolate, but the effect of these compounds was slightly less than that of unlabeled MX-68. On replacing Na+ with choline, MX-68 uptake remained unchanged, whereas the methotrexate uptake was reduced. Uptake of MX-68 increased as the extracellular pH fell from 7.5 to 5.5. These results suggest that MX-68 is taken up via active transport systems. The uptake of MX-68 by CMVs prepared from normal rats exhibited clear ATP dependence, whereas ATP had only a minimal effect on the uptake by CMVs from Eisai-hyperbilirubinemic rats with a hereditary deficiency in canalicular multispecific organic anion transporter (cMOAT). The initial uptake rate of ATP-dependent MX-68 transport showed saturation with kinetic parameters similar to those of methotrexate. MX-68 inhibited the ATP-dependent transport of 2,4-dinitrophenyl-S-glutathione, a typical substrate for cMOAT, the inhibition constant (162 microM) being comparable with the Km of ATP-dependent MX-68 transport. These results suggest that the biliary excretion of MX-68 via the bile canalicular membrane is mediated mainly by cMOAT. In conclusion, active transport systems are involved in membrane penetration of MX-68 both at sinusoidal and canalicular sides in the liver, the latter being mainly involved with methotrexate (MTX) whereas the former differs partially from that for MTX.
Graduate School of Pharmaceutical Sciences, University of Tokyo, Bunkyo-ku, Tokyo, Japan.
Y H
YH
Han
Y
Y
Kato
Y
Y
Watanabe
K
K
Terao
Y
Y
Asoh
Y
Y
Sugiyama
D015074:2-Aminoadipic Acid / Q000031:analogs & derivatives* / Q000493:pharmacokinetics*; D000818:Animals; D027321:Anion Transport Proteins; D000838:Anions / Q000494:pharmacology; D000963:Antimetabolites / Q000494:pharmacology; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D004337:Drug Carriers; D005492:Folic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005493:Folic Acid Antagonists / Q000493:pharmacokinetics*; D005978:Glutathione / Q000031:analogs & derivatives / Q000378:metabolism; D022781:Hepatocytes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008297:Male; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000493:pharmacokinetics*; D051381:Rats; D012964:Sodium / Q000494:pharmacology; D013997:Time Factors; D022161:Transport Vesicles / Q000378:metabolism; D014316:Tritium
D027321:Anion Transport Proteins; D000838:Anions; D000963:Antimetabolites; D002352:Carrier Proteins; D004337:Drug Carriers; 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; D014316:Tritium; D015074:2-Aminoadipic Acid; C030190:S-(2,4-dinitrophenyl)glutathione; D005492:Folic Acid; D012964:Sodium; D005978:Glutathione; D008727:Methotrexate