Cytotoxic synergism of methioninase in combination with 5-fluorouracil andfolinic acid

Potentiation of the cytotoxic activity of 5-fluorouracil (FUra) by folinic acid (5-HCO-H(4)folate) is due to elevation of the methylene tetrahydrofola te (CH2-H(4)folate) level, which increases the stability of the ternary com plex of thymidylate synthase (TS), fluorodeoxyuridine monophosphate, and CH 2-H(4)folate that inactivates the TS. Methionine deprivation results in the production of tetrahydrofolate (H(4)folate) and, subsequently, CH2-H(4)fol ate from methyl tetrahydrofolate, as a consequence of the induction of meth ionine synthesis. We hypothesized that the efficacy of FUra could be augmen ted by the combination of high-concentration 5-HCO-H(4)folate and recombina nt methioninase (rMETase), a methionine-cleaving enzyme. Studies in vitro w ere performed with the cell line CCRF-CEM. Cytotoxic synergism of FUra + rM ETase and FUra + 5-HCO-H(4)folate + rMETase was demonstrated with the combi nation index throughout a broad concentration range of FUra and rMETase. A subcytotoxic concentration of rMETase reduced the IC50 of FUra by a factor of 3.6, and by a factor of 7.5, in the absence and in the presence of 5-HCO -H(4)folate, respectively. 5-HCO-H(4)folate increased the intracellular con centrations of CH2-H(4)folate and H(4)folate from their baseline levels. Co ncentrations of folates were not changed by exposure to rMETase. Levels of free TS in cells treated with FUra + 5-HCO-H,folate and with FUra + rMETase were lower than those in cells exposed to FUra alone. The decrease of TS w as still more pronounced in cells treated with FUra + 5-HCO-H(4)folate + rM ETase. The synergism described in this study will be a basis for further ex ploration of combinations of fluoropyrimidines, folates, and rMETase. (C) 2 001 Elsevier Science Inc. All rights reserved.