Anti-Parasitics with a Triple Threat: Targeting Parasite Enzymes, the Proton Motive Force, and Host Cell-Mediated Killing

We investigated the effects of the tuberculosis drug candidate SQ109 (<b>8a</b>) and of its analog MeSQ109 (<b>8b</b>) against <i>Leishmania mexicana</i> in promastigote and amastigote forms and against host cell macrophages finding potent activity (1.7 nM) for MeSQ109 against the intracellular forms, as well as low toxicity (∼61 μM) to host cells, resulting in a selectivity index of ∼36,000. We then investigated the mechanism of action of MeSQ109, finding that it targeted parasite mitochondria, collapsing the proton motive force, as well as targeting acidocalcisomes, rapidly increasing the intracellular Ca²⁺ concentration. Using an <i>E. coli</i> inverted membrane vesicle assay, we investigated the pH gradient collapse for SQ109 and 17 analogs, finding that there was a significant correlation (on average, <i>R</i> = 0.67, <i>p</i> = 0.008) between pH gradient collapse and cell growth inhibition in <i>Trypanosoma brucei</i>, <i>T. cruzi</i>, <i>L. donovani</i>, and <i>Plasmodium falciparum</i>. We also investigated pH gradient collapse with other antileishmanial agents: azoles, antimonials, benzofurans, amphotericin B, and miltefosine. The enhanced activity against intracellular trypanosomatids is seen with <i>Leishmania</i> spp. grown in macrophages but not with <i>Trypanosoma cruzi</i> in epithelial cells and is proposed to be due in part to host-based killing, based on the recent observation that SQ109 is known to convert macrophages to a pro-inflammatory (M1) phenotype.