Stereoselective transport of amethopterin enantiomers by the proton-coupled folate transporter.
Stereoselective transport of amethopterin enantiomers by the proton-coupled folate transporter.
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DOI：
10.2133/dmpk.25.283
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发表时间：
2010
影响因子：
2.1
通讯作者：
Tomoya Narawa;T. Itoh
中科院分区：
文献类型：
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作者：
Tomoya Narawa;T. Itoh
利用表达PCFT的HEK293细胞（PCFT-HEK293细胞）研究了质子偶联叶酸转运体（PCFT）对甲氨蝶呤（l -阿甲蝶呤，L-MTX）及其对极物（d -阿甲蝶呤，D-MTX）的立体选择性转运。L-MTX和D-MTX的摄取都是pH依赖的，并且随着细胞外pH从5.0到7.4的增加而减少。L-MTX进入PCFT-HEK293细胞的初始摄取速率遵循Michaelis-Menten动力学，K(m)值约为5.0微米。Dixon图显示，未标记的L-MTX、D-MTX和叶酸（FA）竞争性地抑制了L-MTX的摄取，K(i)值分别约为3.6、180和2.1微米。D-MTX进入PCFT-HEK293细胞的初始摄取速率也符合Michaelis-Menten动力学，K(m)值为211微米。D-MTX的V（max）值与L-MTX相似。本研究表明PCFT转运MTX对映体具有高度立体选择性，L-MTX的摄取清除率约为D-MTX的40倍。结果还表明，这种高立体选择性是由于K(m)值的差异，而不是V（max）值的差异。观察到的立体选择性与人体内MTX对映体肠道吸收的差异是一致的。
Stereoselective transport of methotrexate (L-amethopterin, L-MTX) and its antipode (D-amethopterin, D-MTX) by the proton-coupled folate transporter (PCFT) was examined using PCFT-expressing HEK293 cells (PCFT-HEK293 cells). Uptake of both L-MTX and D-MTX was pH-dependent and decreased with an increase in the extracellular pH from 5.0 to 7.4. The initial uptake rate of L-MTX into PCFT-HEK293 cells followed Michaelis-Menten kinetics with a K(m) value of approximately 5.0 microM. Dixon plots revealed that L-MTX uptake was inhibited competitively by unlabeled L-MTX, D-MTX, and folic acid (FA), with K(i) values of approximately 3.6, 180, and 2.1 microM, respectively. The initial uptake rate of D-MTX into PCFT-HEK293 cells also followed Michaelis-Menten kinetics with a K(m) value of 211 microM. The V(max) value of D-MTX was similar to that of L-MTX. The present study revealed that the transport of MTX enantiomers by PCFT is highly stereoselective with the uptake clearance of L-MTX being approximately 40-fold greater than that of D-MTX. It was also revealed that this high stereoselectivity results from the difference in K(m) values, and not V(max) values, between the enantiomers. The observed stereoselectivity was consistent with the differences in the intestinal absorption of MTX enantiomers in humans.