Prolonged and enhanced suppression of thymidylate synthase by weekly 24-h infusion of high-dose 5-fluorouracil

摘要

We have recently demonstrated that HDFL (high-dose 5-FU 2600 mg m–2 week–1 and leucovorin 500 mg m–2 week–1, weekly 24-h infusion) is highly active in the treatment of gastric cancer. To further clarify the possible mechanism underlying the improved activity of HDFL compared with conventional 5-FU regimens, we conducted in vitro studies examining the effect of these regimens on the differential regulation of thymidylate synthase (TS) in NCI-N87, a human gastric cancer cell line. The expected serum concentrations of 5-FU are 100–200 mM (lasting for less than 30 min) and 5–10 mM (lasting for 24 h) for the conventional 5-FU regimens (bolus injection or short intravenous infusion of 5-FU 370–500 mg m–2) and the HDFL regimens, respectively. Western blot analysis revealed that 24-h exposure of NCI-N87 to 2.5–10.0 mM of 5-FU resulted in a dose-dependent depletion of free TS, lasting for more than 24 h. In contrast, 30-min exposure of NCI-N87 to 200 mM of 5-FU resulted in a less than 12-h depletion of free TS. Moreover, 24-h exposure to 5-FU resulted in a higher S-phase blockade and enhanced cytotoxicity. In both modes of 5-FU treatment, the initial rapid depletion of free TS was accompanied by a rapid increment of a higher-molecular-weight TS molecule, suggesting that rapid formation of the ternary complex was the key mechanism of 5-FU action during this period. Northern blot analysis showed that the steady-state mRNA of TS was not affected by either of the schedules. We conclude that 24-h exposure of gastric cancer cells to low concentration of 5-FU resulted in better suppression of free TS, a higher degree of S-phase blockade, and enhanced cytotoxicity compared to 30-min exposure to high concentration of 5-FU. These in vitro results may help explain the improved clinical efficacy of HDFL regimens compared to conventional 5-FU regimens. © 2000 Cancer Research Campaign