Synergistic and antagonistic interactions of methotrexate and 1-β- D-arabinofuranosylcytosine in hepatoma cells : The modulating effect of purines

Abstract

A 4-hr pretreatment with methotrexate antagonized the cytotoxic effect of subsequent arabinosylcytosine treatment in rat hepatoma cells of lines N 1S 1 and 3924A, but in the hepatoma line 8999R and the fibroblast line BF5, MTX pretreatment was synergistic with the arabinosylcytosine treatment. Measurement of cellular deoxyribonucleoside triphosphate concentrations showed that in those lines in which antagonism was found the dCTP increased, whereas in the lines where the drugs were synergistic the dCTP pool was decreased. Conversely, dATP levels were high when the drugs were synergistic and low when antagonism was obtained. Although methotrexate pretreatment antagonized arabinosylcytosine in N 1S 1 and 3924A cells, pretreatment of these cells with the combination of methotrexate plus a purine (either hypoxanthine or 2′-deoxyadenosine) resulted in synergism with arabinosylcytosine. Deoxynucleotide pool measurements showed that methotrexate in combination with either hypoxanthine or 2′-deoxyadenosine increased dATP and decreased dCTP in the N 1S 1 and 3924A hepatoma cells. In N 1S 1 cells, pretreatment with 2′-deoxyadenosine alone for 4 hr was synergistic with arabinosylcytosine. It was concluded that elevated dATP pools enhanced arabinosylcytosine cytotoxicity by depleting the dCTP pool, through feedback inhibition of ribonucleotide reductase, thus causing greater inhibition of DNA biosynthesis and greater incorporation of AraCTP into nucleic acid. Methotrexate was synergistic in those cell lines where dATP accumulated and dCTP was decreased, but when methotrexate had a potent antipurine effect dCTP pools increased and arabinosylcytosine was antagonized. The synergistic interaction was more marked at cytocidal drug concentrations than it was at growth-inhibitory doses.