Leucovorin Enhances Chtotoxicity of Trimetrexate/Fluorouracil, But Not Methotrexate/Fluorouracil, in CCRF-CEM Cells

Abstract

Background : Increased response rates in studies of patients with colon cancer have indicated that the cytotoxic effects of fluorouracil (5-FU) are potentiated by leucovorin (LV) and by methotrexate (MTX). However, preliminary studies using a sequential combination of MTX, LV, and 5-FU showed no additional potentiation. Purpose : We hypothesized that the lack of additional cell kill with this combination could be due to competition of LV with MTX for cellular uptake and reduced folate polygluta-mylation. We have tested this possibility by comparing the cytotoxicity of drug combinations containing MTX with that of drug combinations containing trimetrexate (TMTX), an anti-folate that does not compete with LV for uptake or polyglutamylation. Methods : Human lymphocytic leukemia CCRF-CEM cells were exposed to MTX or TMTX for 24 hours and to 5-FU during the last 4 hours of antifo-late exposure. LV was administered 30 minutes before 5-FU. Results : After 20 hours of exposure to TMTX or MTX, intracellular levels of phosphoribosyl pyrophosphate were elevated to a similar degree, and these levels did not decrease after a 30-minute exposure to LV. No additional cell kill was observed when LV was added to the MTX/5-FU combination, but cytotoxicity was enhanced when LV was added to the TMTX/5-FU combination. Conclusions : This study supports the hypothesis that the lack of additional cell kill when high-dose LV is added to the MTX/5-FU combination may be due to competition of MTX with LV for cellular uptake and/or competition of MTX or its polygluta-mates with polyglutamylation of reduced folates. Inasmuch as TMTX does not compete with LV and reduced folates for uptake and polyglutamylation, the synergy obtained with the combination of TMTX plus 5-FU and high-dose LV further supports this hypothesis. [J Natl Cancer Inst 84:1033–1038, 1992]