Reduced folate derivatives are endogenous substrates for cMOAT in rats.
We examined the role of the canalicular multispecific organic anion transporter (cMOAT) in the biliary excretion of reduced folate derivatives in vivo and in vitro using normal [Sprague-Dawley rats (SDR)] and mutant [Eisai hyperbilirubinemic rats (EHBR)] rats whose cMOAT is hereditarily deficient. In vivo, the biliary excretion of endogenous tetrahydrofolate (H4PteGlu), 5-methyltetrahydrofolate (5-CH3-H4PteGlu), and 5,10-methylenetetrahydrofolate (5, 10-CH2-H4PteGlu) in EHBR was reduced to 8.2%, 1.9%, and 5.5% of those in SDR, respectively, whereas that of 10-formyltetrahydrofolate (10-HCO-H4PteGlu) was detected only in SDR and not in EHBR. Bile drainage caused reduction of endogenous plasma folate concentrations in SDR but not in EHBR. In vitro, significant ATP-dependent uptake of 3H-labeled 5-CH3-H4PteGlu into canalicular membrane vesicles was observed only in SDR. This ATP-dependent uptake was saturable with a Michaelis constant (Km) value of 126 microM, which was comparable with its inhibitor constant (Ki) value of 121 microM for the ATP-dependent uptake of a typical cMOAT substrate, 2,4-dinitrophenyl-S-glutathione (DNP-SG). Vice versa, DNP-SG inhibited the uptake of 5-CH3-H4PteGlu with a Ki of 35 microM, which was similar to its Km value. In addition, H4PteGlu and 5, 10-CH2-H4PteGlu also inhibited the ATP-dependent uptake of DNP-SG. These results indicate that 5-CH3-H4PteGlu and other derivatives are transported via cMOAT. Therefore, reduced folate derivatives are the first endogenous substrates for cMOAT that do not contain glutathione, glucuronide, or sulfate moieties.
Kusuhara
H
H
Graduate School of Pharmaceutical Sciences, University of Tokyo, Bunkyo-ku, Tokyo 113, Japan.
Han
Y H
YH
Shimoda
M
M
Kokue
E
E
Suzuki
H
H
Sugiyama
Y
Y
D000818:Animals; D027321:Anion Transport Proteins; D001646:Bile / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D005492:Folic Acid / Q000031:analogs & derivatives* / Q000097:blood / Q000493:pharmacokinetics*; D006932:Hyperbilirubinemia / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008657:Metabolic Clearance Rate; D015394:Molecular Structure; D010084:Oxidation-Reduction; D051381:Rats; D011922:Rats, Mutant Strains; D017207:Rats, Sprague-Dawley; D013379:Substrate Specificity; D013997:Time Factors
D027321:Anion Transport Proteins; D002352:Carrier Proteins; D005492:Folic Acid