title: A new mechanism of methotrexate action revealed by target screening with affinity beads.
abstract: Methotrexate (MTX) is the anticancer and antirheumatoid drug that is believed to block nucleotide synthesis and cell cycle by inhibiting dihydrofolate reductase activity. We have developed novel affinity matrices, termed SG beads, that are easy to manipulate and are compatible with surface functionalization. Using the matrices, here we present evidence that deoxycytidine kinase (dCK), an enzyme that acts in the salvage pathway of nucleotide biosynthesis, is another target of MTX. MTX modulates dCK activity differentially depending on substrate concentrations. 1-beta-D-Arabinofuranosylcytosine (ara-C), a chemotherapy agent often used in combination with MTX, is a nucleoside analog whose incorporation into chromosome requires prior phosphorylation by dCK. We show that, remarkably, MTX enhances incorporation and cytotoxicity of ara-C through regulation of dCK activity in Burkitt's lymphoma cells. Thus, this study provides new insight into the mechanisms underlying MTX actions and demonstrates the usefulness of the SG beads.
authors.affiliation: Aphoenix Inc., Tokyo, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Uga
authors.forename: Chikanori
authors.initials: C
authors.lastname: Kuramori
authors.forename: Akiko
authors.initials: A
authors.lastname: Ohta
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Tsuboi
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Tanaka
authors.forename: Mamoru
authors.initials: M
authors.lastname: Hatakeyama
authors.forename: Yuki
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: Takashi
authors.initials: T
authors.lastname: Takahashi
authors.forename: Masahiro
authors.initials: M
authors.lastname: Kizaki
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Handa
mesh_terms: D000595:Amino Acid Sequence; D001085:Arabinofuranosylcytosine Triphosphate / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002846:Chromatography, Affinity / Q000379:methods*; D002877:Chromosomes, Human / Q000235:genetics; D003842:Deoxycytidine Kinase / Q000737:chemistry / Q000302:isolation & purification; D004354:Drug Screening Assays, Antitumor / Q000295:instrumentation* / Q000379:methods*; D006801:Humans; D019169:Jurkat Cells; D008727:Methotrexate / Q000494:pharmacology*; D008863:Microspheres; D008969:Molecular Sequence Data; D008970:Molecular Weight; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism
chemical_list: D001085:Arabinofuranosylcytosine Triphosphate; D013762:Tetrahydrofolate Dehydrogenase; D003842:Deoxycytidine Kinase; D008727:Methotrexate