NUCLEOSIDE-MEDIATED MITIGATION OF 5-FLUOROURACIL-INDUCED TOXICITY IN SYNCHRONIZED MURINE ERYTHROLEUKEMIC CELLS

5-Fluorouracil (5-FU) is a chemotherapeutic agent known to retard embr yonic growth and induce cleft palate and limb deformities. The predomi nant mechanism underlying its toxic action is thought to be inhibition of thymidylate synthetase (TS), and hence thymidine triphosphate (dTT P) synthesis, resulting in alteration of the balance of deoxynucleotid e (dNTP) pools and disruption of DNA synthesis. Indeed, previously we demonstrated retarded cell-cycle progression concurrent with a 60% dec rease in TS activity in rat whole embryos following maternal exposure to 40 mg/kg 5-FU on Gestational Day 14 and in the murine erythroleukem ic cell (MELC) suspension culture following exposure to 5-25 mu M 5-FU for 2 hr. In the study described herein, we used high-performance liq uid chromatography (HPLC) to demonstrate in both of these model system s that 5-FU exposure results in similar patterns of dNTP perturbations : a prolonged decrease in dTTP and dGTP levels and an increase in dCTP and dATP. In addition, we used centrifugal elutriation to synchronize MELC in the phases of the cell cycle (G(0)/G(1) and early S) most sen sitive to 5-FU to investigate the ability of nucleoside supplementatio n to mitigate 5-FU-induced toxicity. Our data indicate that following a 2-hr exposure to 5-25 mu M 5-FU, supplementation with 1-10 mu M thym idine (TdR) for 24 hr partially reverses 5-FU-induced toxicity as evid enced by increased cellular proliferation and cell-cycle progression a nd amelioration of 5-FU-induced perturbations of protein synthesis and cellular membrane permeability compared to unsupplemented 5-FU-expose d cells. However, TdR concentrations greater than or equal to 100 mu M inhibited growth or were cytotoxic. In comparison, supplementation wi th 10 mu M-10 mM of deoxycytidine (CdR) was not toxic, but effected a dose-dependent recovery from 5-FU-induced toxicity. At 1-100 mu M, nei ther deoxyadenosine nor deoxyguanosine supplementation reduced 5-FU-in duced toxicity; at higher concentrations, both purine nucleotides inhi bited cell growth. Although these results support the hypothesis that 5-FU disrupts the MELC cell cycle by depleting dTTP (a perturbation th at is reversible by TdR supplementation), they also indicate that CdR supplementation offers an additional recovery pathway. (C) 1997 Academ ic Press.