title: Mechanism of dihydrofolate reductase downregulation in melanoma by 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin.
abstract: In our search to improve the stability and cellular absorption of tea polyphenols, we synthesized 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin (TMECG), which showed high antiproliferative activity against melanoma. TMECG downregulates dihydrofolate reductase (DHFR) expression in melanoma cells and we detail the sequential mechanisms that result from this even. TMECG is specifically activated in melanoma cells to form a stable quinone methide (TMECG-QM). TMECG-QM has a dual action on these cells. First, it acts as a potent antifolate compound, disrupting folate metabolism and increasing intracellular oxidized folate coenzymes, such as dihydrofolate, which is a non-competitive inhibitor of dihydropterine reductase, an enzyme essential for tetrahydrobiopterin (H(4)B) recycling. Such inhibition results in H(4)B deficiency, endothelial nitric oxide synthase (eNOS) uncoupling and superoxide production. Second, TMECG-QM acts as an efficient superoxide scavenger and promotes intra-cellular H(2)O(2) accumulation. Here, we present evidence that TMECG markedly reduces melanoma H(4)B and NO bioavailability and that TMECG action is abolished by the eNOS inhibitor N(omega)-nitro-L-arginine methyl ester or the H(2)O(2) scavenger catalase, which strongly suggests H(2)O(2)-dependent DHFR downregulation. In addition, the data presented here indicate that the simultaneous targeting of important pathways for melanoma survival, such as the folate cycle, H(4)B recycling, and the eNOS reaction, could represent an attractive strategy for fighting this malignant skin pathology.
authors.affiliation: Department of Biochemistry and Molecular Biology A, School of Biology, University of Murcia, E-30100 Espinardo, Murcia, Spain.
authors.forename: Luís
authors.initials: L
authors.lastname: Sánchez-del-Campo
authors.forename: Soledad
authors.initials: S
authors.lastname: Chazarra
authors.forename: María F
authors.initials: MF
authors.lastname: Montenegro
authors.forename: Juan
authors.initials: J
authors.lastname: Cabezas-Herrera
authors.forename: José Neptuno
authors.initials: JN
authors.lastname: Rodríguez-López
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D001708:Biopterins / Q000031:analogs & derivatives / Q000378:metabolism; D015153:Blotting, Western; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004093:Dihydropteridine Reductase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects*; D005492:Folic Acid / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008545:Melanoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D016877:Oxidants / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism*
chemical_list: C529447:3-O-(3,4,5-trimethoxybenzoyl)epicatechin; D016877:Oxidants; D001708:Biopterins; C010920:dihydrofolate; D002392:Catechin; D005492:Folic Acid; D006861:Hydrogen Peroxide; D052250:Nitric Oxide Synthase Type III; D013762:Tetrahydrofolate Dehydrogenase; D004093:Dihydropteridine Reductase; C003402:sapropterin