Inhibition of Synthesis of Pyrimidine Nucleotides by 2-Hydroxy-3-(3,3-dichloroallyl)-1,4-naphthoquinone

Abstract Dichloroallyl lawsone [2-hydroxy-3-(3,3-dichloroallyl)-1,4-naphthoquinone, NSC 126771] is of interest as an antitumor agent. Like certain other naphthoquinones, it is known to be a respiratory poison as a result of interference with electron transport, but it has not been shown that this action is responsible for its toxicity or antitiumor activity. Studies in cultured L1210 cells showed that dichloroallyl lawsone ( a ) stimulated the incorporation of [ 14 C]uridine into macromolecules, ( b ) specifically decreased the pool sizes of uridine triphosphate, ( c ) prevented the pyrazofurin-induced accumulation of orotidine and orotic acid, and ( d ) exerted cytotoxicity that could be prevented or reversed almost completely by uridine. This evidence that dichloroallyl lawsone specifically inhibited biosynthesis of pyrimidine nucleotides in intact cells was confirmed by studies in homogenates; these showed that this agent inhibited the conversion of [ 14 C]carbamylphosphate to orotate and uridine monophosphate but not its conversion to dihydroorotate and that it was without effect on the conversion of [6- 14 C]orotate to uridine monophosphate. That the site of inhibition was dihydroorotate dehydrogenase was confirmed in studies with isolated mitochondria from mouse liver. An enzyme from this source catalyzed the oxidation of [6- 14 C]dihydroorotate (apparent K m , 6.7 × 10 −6 m), and dichloroallyl lawsone functioned as an inhibitor uncompetitive with dihydroorotate (K i , 2.7 × 10 −8 m). Addition of ubiquinone was without effect on the reaction. Dichloroallyl lawsone, however, did not inhibit dihydroorotate dehydrogenase from Zymobacterium oroticum . These results indicate that the blockade of pyrimidine biosynthesis resulting from inhibition of dihydroorotate dehydrogenase is primarily responsible for the cytotoxicity of dichloroallyl lawsone to L1210 cells. More limited studies with the structurally related quinone, lapachol [2-hydroxy-3-(3-methyl-2-butenyl)-1,4-naphthoquinone], indicated that it exerts similar effects on biosynthesis of pyrimidines.