The role of methotrexate and dihydrofolate polyglutamates in the enhancement of fluorouracil action by methotrexate.
The effect of the methotrexate (MTX)-5-fluorouracil (FUra) combination on L1210 cell viability was highly sequence dependent in that a 1.4 log (25-fold) increase in cytotoxicity was observed when the MTX preceded FUra, as compared to the reverse sequence. The formation of ternary complexes of thymidylate synthetase has been examined as a basis for the interaction of FUra and MTX in L1210 cells. L1210 cells converted 39% of the total intracellular MTX into MTX-poly-gamma-glutamates within 4 hours of a 1 microM MTX exposure. MTX-diglutamate (2,4-diamino,N 10-methylpteroyl-diglutamate) and MTX-triglutamate were the predominant metabolites. In contrast to the ternary complexes formed with MTX and MTX-diglutamate, the 5-fluorodeoxy-uridylate (FdUMP)-7,8-dihydropteroylpentaglutamate-enzyme and the FdUMP-5,10-methylenetetrahydropteroylpentaglutamate-enzyme complexes were stable to polyacrylamide gel electrophoresis under nondenaturing conditions. MTX-diglutamate enhanced the extent of tight-binding inhibition of thymidylate synthetase activity by FdUMP in the presence of saturating 5,10-methylenetetrahydropteroylpentaglutamate, suggesting that MTX-diglutamate did not antagonize the formation of FdUMP-5,10-methylenetetrahydropteroylpentaglutamate enzyme complex. We propose that the sequence-dependent effect of MTX plus FUra on L1210 cell viability results from MTX and MTX polyglutamate inhibition of dihydrofolate reductase, and consequently a trapping of intracellular folates as dihydrofolate polyglutamates that could increase the extent of FdUMP binding to thymidylate synthetase.
D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004334:Drug Administration Schedule; D004359:Drug Therapy, Combination; D005468:Fluorodeoxyuridylate / Q000378:metabolism; D005472:Fluorouracil / Q000008:administration & dosage / Q000378:metabolism*; D005492:Folic Acid / Q000031:analogs & derivatives*; D005493:Folic Acid Antagonists; D007939:Leukemia L1210 / Q000378:metabolism* / Q000473:pathology; D008727:Methotrexate / Q000008:administration & dosage / Q000031:analogs & derivatives / Q000096:biosynthesis / Q000378:metabolism*; D051379:Mice; D011099:Polyglutamic Acid / Q000031:analogs & derivatives / Q000096:biosynthesis; D011624:Pteroylpolyglutamic Acids / Q000378:metabolism*; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism
D005493:Folic Acid Antagonists; D011624:Pteroylpolyglutamic Acids; D005468:Fluorodeoxyuridylate; D011099:Polyglutamic Acid; C014085:methotrexate polyglutamate; D005492:Folic Acid; D013940:Thymidylate Synthase; D005472:Fluorouracil; D008727:Methotrexate