title: Stereoselective transport of human His27- and Arg27-reduced folate carrier.
abstract: The stereoselective transport of methotrexate (L-amethopterin, L-MTX) and its antipode (D-amethopterin, D-MTX) by human reduced folate carrier (hRFC) has been examined in HEK293 cells expressing H27-hRFC and R27-hRFC. The uptake of both L-MTX and D-MTX increased as the extracellular pH increased from 6.0 to 7.4. The initial uptake rate of L-MTX into the H27- and R27-hRFCs of the HEK293 cells followed Michaelis-Menten kinetics with Km values of approximately 0.24 and 0.47 µM, respectively. Dixon plots revealed that the [(3)H]-L-MTX uptake mediated by the H27- and R27-hRFCs was inhibited competitively by unlabeled L-MTX with Ki values of approximately 0.1 and 0.5 µM, respectively. D-MTX also competitively inhibited the H27- and R27-hRFC mediated uptake of [(3)H]-L-MTX with Ki values of approximately 3.4 and 3.2 µM, respectively. The RFC-mediated uptake clearance of L-MTX was approximately 15-fold greater than that of D-MTX in the H27-hRFC-HEK293 cells, and was more than 30-fold greater than that of D-MTX in the R27-hRFC-HEK293 cells. The results of the current study have revealed that the enantiomers of MTX enantiomers can be transported in a stereoselective manner by the H27- and R27-hRFCs because of significant differences in the affinities of the enantiomers to the hRFC.
authors.affiliation: School of Pharmacy, Kitasato University.
authors.forename: Tomoya
authors.initials: T
authors.lastname: Narawa
authors.forename: Hideaki
authors.initials: H
authors.lastname: Yanagisawa
authors.forename: Tomoo
authors.initials: T
authors.lastname: Itoh
mesh_terms: D000328:Adult; D001692:Biological Transport / Q000235:genetics; D005493:Folic Acid Antagonists / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D008727:Methotrexate / Q000378:metabolism*; D058978:Reduced Folate Carrier Protein / Q000235:genetics* / Q000378:metabolism; D013237:Stereoisomerism; D014162:Transfection; D055815:Young Adult
chemical_list: D005493:Folic Acid Antagonists; D058978:Reduced Folate Carrier Protein; D008727:Methotrexate