Multiple membrane transport systems for the uptake of folate-based thymidylate synthase inhibitors.
N10-Propargyl-5,8-dideazafolic acid (CB3717) and 2-desamino-2-methyl-N10-propargyl-5,8-dideazafolic acid (ICI-198,583) are potent folate-based inhibitors of thymidylate synthase. We studied the membrane transport and the growth-inhibitory effects of the two thymidylate synthase inhibitors on human CCRF-CEM leukemia cells with different transport properties for folic acid, reduced folates, and methotrexate (MTX). Membrane transport of [3H]ICI-198,583 can proceed via the high affinity/low capacity reduced folate carrier as supported by findings that (a) uptake of [3H]ICI-198,583 was significantly impaired in CEM cells which have a transport defect for MTX, (b) variants of CEM cells which overproduce the reduced folate carrier system showed a concomitant increase in the uptake of [3H]ICI-198,583 as for [3H]MTX, (c) MTX inhibited transport of [3H]ICI-198,583, and (d) uptake of [3H]ICI-198,583 was inhibited after treatment of CEM cells with an N-hydroxysuccinimide ester of MTX, which is a potent inhibitor of MTX transport. However, a membrane-associated folate-binding protein (FBP) offers another route for entry of CB3717 and ICI-198,583. CEM-FBP cells that have an elevated amount of FBP and do not have a functional reduced folate carrier were 640- and 61-fold more sensitive to CB3717 and ICI-198,583, respectively, compared to control CEM cells expressing the reduced folate/MTX carrier. This high sensitivity was related to a high affinity of the FBP for CB3717 and ICI-198,583 (Kd 2-3 nM), which is only 3-fold lower than for folic acid (Kd 1 nM) but significantly higher than for MTX (Kd 100 nM). Furthermore, after incubation of CEM-FBP cells for 24 h at 10 nM [3H]ICI-198,583, the high affinity binding of the FBP for ICI-198,583 allowed a 600-fold concentrative uptake of [3H]ICI-198,583 and its conversion to polyglutamate forms. These results indicate that multiple folate transport systems may be involved in the uptake of folate-based thymidylate synthase inhibitors.
Jansen
G
G
Department of Internal Medicine, University Hospital Utrecht, The Netherlands.
Schornagel
J H
JH
Westerhof
G R
GR
Rijksen
G
G
Newell
D R
DR
Jackman
A L
AL
D000970:Antineoplastic Agents / Q000493:pharmacokinetics*; D001692:Biological Transport; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004337:Drug Carriers; D005492:Folic Acid / Q000031:analogs & derivatives* / Q000378:metabolism / Q000493:pharmacokinetics; D005493:Folic Acid Antagonists / Q000493:pharmacokinetics; D006801:Humans; D066298:In Vitro Techniques; D007938:Leukemia / Q000378:metabolism*; D008727:Methotrexate; D011799:Quinazolines / Q000493:pharmacokinetics*; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors*
D000970:Antineoplastic Agents; D004337:Drug Carriers; D005493:Folic Acid Antagonists; D011799:Quinazolines; C031662:CB 3717; D005492:Folic Acid; D013940:Thymidylate Synthase; C059966:ICI 198583; D008727:Methotrexate