Background Alveolar echinococcosis (AE) is a fatal zoonotic disease distributed mainly in the Northern Hemisphere. At present, its curative treatment relies on surgery, and the development of effective drugs is needed. We previously demonstrated the anti-echinococcal effect of atovaquone (ATV) as a mitochondrial complex III inhibitor in both in vitro and in vivo experiments. However, the anti-echinococcal effect of ATV in vivo was limited, since ATV inhibits only aerobic respiration. In this study, we investigated whether ATV exhibits a synergistic effect when used in combination with other anti-echinococcal drug candidates, including mefloquine (MF), 3-bromopyruvic acid (3BP), crocin, and verapamil (Ver), thereby enhancing their antiparasitic effectiveness. Methods The synergistic effect of anti-echinococcal drug candidates with ATV was examined in culture experiments with Echinococcus multilocularis protoscoleces. Based on the results of these culture experiments, ATV and 3BP were individually and in combination orally administered to BALB/c mice infected with E. multilocularis (dose of 300 eggs). Each drug treatment was started three days prior to infection and continued until day 28 after egg administration, and the number of cysts located in the liver was evaluated (Experiment A). The anti-echinococcal effectiveness of the combination of ATV and 3BP was also evaluated by treating mice with E. multilocularis primary infection for eight weeks (Experiment B) and comparing the effects on cyst growth to those of albendazole (ABZ). Results Culture experiments with E. multilocularis protoscoleces showed that the combined treatments of ATV with 3BP, MF, and Ver were more effective at parasite elimination under both aerobic and anaerobic conditions than the single drug treatments. Crocin was ineffective in the culture assay. In Experiment A, the number of cysts was significantly reduced only in the groups treated with ABZ alone (median 48.0, interquartile range 36.3-58.0) and the combination of ATV and 3BP (median 54.5, interquartile range 46.5-62.8) compared to the control (median 90.0, interquartile range 67.0-100.5). MF and Ver did not exhibit significant in vivo effects on their own. In Experiment B, the group treated with ATV + 3BP showed a similar anti-echinococcal effect as the group treated with ABZ alone. Conclusion In the culture assay, ATV in combination with 3BP, MF, and Ver showed a synergistic effect, enhancing the anti-echinococcal effect under both aerobic and anaerobic conditions. In mice experimentally treated with primary hydatid cysts, co-administration of ATV and 3BP showed a significant preventive effect against infection and demonstrated therapeutic efficacy comparable to that of ABZ. These findings are anticipated to contribute to the development of more effective therapeutic agents for AE.
Keywords: 3-bromopyruvic acid; atovaquone; echinococcosis; echinococcus multilocularis; verapamil.
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