Paromomycin: uptake and resistance in Leishmania donovani

Abstract

Paromomycin is currently in phase IV clinical trials against leishmaniasis. In the present work we elucidate the effect and mechanism of uptake of paromomycin in Leishmania donovani. The in vitro sensitivities of both promastigotes and amastigotes were determined to this aminoglycoside. Association of paromomycin with L. donovani involved a rapid initial phase that was non-saturable up to 1 mM of the drug. This initial phase was largely independent of temperature and not affected by metabolic inhibitors. Poly-lysine, a membrane impermeant polycation, caused profound inhibition of this association of the drug with the parasite indicating that it represented a binding of the cationic paromomycin to the negatively charged leishmanial glycocalyx. After 72 h of exposure to the drug the mitochondrial membrane potential was significantly decreased, indicating that this organelle might be the ultimate target of the drug. Both cytoplasmic and mitochondrial protein synthesis were inhibited following paromomycin exposure. A line selected for resistance to the drug showed reduced paromomycin accumulation associated with a significant reduction in the initial binding to the cell surface. The drug induced reduction in membrane potential and inhibition of protein synthesis were less pronounced in the resistant strain in comparison to the wild-type.