Investigating the mode of action of the redox-active antimalarial drug plasmodione using the yeast model

Malaria is caused by protozoan parasites and remains a major public health issue in subtropical areas. Plasmodione (3-[4-(trifluoromethyl)benzyl]-menadione) is a novel early lead compound displaying fast-acting antimalarial activity. Treatment with this redox active compound disrupts the redox balance of parasite-infected red blood cells. In vitro, the benzoyl analogue of plasmodione can act as a subversive substrate of the parasite flavoprotein NADPH-dependent glutathione reductase, initiating a redox cycling process producing ROS. Whether this is also true in vivo remains to be investigated. Here, we used the yeast model to investigate the mode of action of plasmodione and uncover enzymes and pathways involved in its activity. We showed that plasmodione is a potent inhibitor of yeast respiratory growth, that in drug-treated cells, the ROS-sensitive aconitase was impaired and that cells with a lower oxidative stress defence were highly sensitive to the drug, indicating that plasmodione may act via an oxidative stress. We found that the mitochondrial respiratory chain flavoprotein NADH-dehydrogenases play a key role in plasmodione activity. Plasmodione and metabolites act as substrates of these enzymes, the reaction resulting in ROS production. This in turn would damage ROS-sensitive enzymes leading to growth arrest. Our data further suggest that plasmodione is a pro-drug whose activity is mainly mediated by its benzhydrol and benzoyl metabolites. Our results in yeast are coherent with existing data obtained in vitro and in Plasmodium falciparum, and provide additional hypotheses that should be investigated in parasites.