EFFECTS OF ATOVAQUONE AND OTHER INHIBITORS ON PNEUMOCYSTIS-CARINII DIHYDROOROTATE DEHYDROGENASE

Dihydroorotate dehydrogenase (DHOD) is a pyrimidine biosynthetic enzym e which is usually directly linked to the mitochondrial respiratory ch ain. Antimalarial naphthoquinones such as atovaquone (566c80) inhibit malarial DHOD by inhibiting electron transport. Since atovaquone also has therapeutic activity against Pneumocystis carinii, the P. carinii DHOD may also be an important drug target. Organisms were obtained fro m immunosuppressed rats, incubated for 24 h in a short-term in vitro c ulture system, and then lysed, P. carinii lysates catalyzed the genera tion of orotate from dihydroorotate at a rate of 852 pmol/mg of protei n per min. Control preparations made from uninfected mice showed much less total enzymatic activity and enzyme specific activity. As expecte d, P. carinii DHOD activity was susceptible to respiratory inhibitors such as cyanide, antimycin A, and salicylhydroxamic acid (SHAM). Susce ptibility to SHAM suggests the presence of an alternative oxidase. In contrast, neither pentamidine nor 5-hydroxy-6-demethylprimaquine (5H6D P), a quinone metabolite of primaquine, inhibited the enzyme. Atovaquo ne inhibited DHOD by 76.3% at 100 mu M and 36.5% at 10 mu M. A similar degree of inhibition was found when the organisms were preincubated w ith the drug. Atovaquone inhibited P. carinii growth in vitro at a som ewhat lower concentration (between 0.3 and 3 mu M). In contrast, Plasm odium falciparum growth and enzyme activity are susceptible to nanomol ar concentrations of atovaquone. Thus, while it is possible that atova quone acts by inhibiting the P. carinii electron transport chain, the possibility of another drug target cannot be excluded.