The interaction between fluoropyrimidines and methotrexate, and [4C]-formate incorporation into nucleic acids and protein

Changes are reported in [14C]-formate incorporation into nucleic acids and protein of Ehrlich ascites tumor cells during exposure to methotrexate (MTX) and fluoropyrimidines. The rate of [14C]-formate incorporation into RNA, DNA, and protein in the presence of only MTX was inhibited by 82%, 91%, and 75% respectively, when compared with control rates. However, in the presence of 5-fluorodeoxyuridine (FdUrd) plus MTX, formate incorporation into RNA, DNA, and protein was inhibited by 67%, 85%, and 66%. Incubation of cells in vitro with [3H]-dihydrofolate (DHF) results in its rapid conversion to [3H]-tetrahydrofolate (THF). The THF/DHF ratio from the soluble fraction of cells that were incubated with [3H]-DHF was 43% greater in the presence of FdUrd and MTX than in the presence of MTX alone. As the rate of [3H]-dUrd incorporation into DNA was reduced by 88% and 99% by pretreating cells with 0.1 muM and 1 muM FdUrd, respectively, the inhibitory effect of MTX on [14C]-formate incorporation into (a) RNA was decreased by 63% and 46%; (b) DNA was decreased by 74% and 61%; and (c) protein was decreased by 63% and 32%. These data suggest that fluoropyrimidines can antagonize the effects of MTX on purines or nucleic acid synthesis and protein synthesis by preventing the consumption of THF for dTMP synthesis.