Mechanism of Action of Miltefosine on Leishmania donovani Involves the Impairment of Acidocalcisome Function and the Activation of the Sphingosine-Dependent Plasma Membrane Ca2+ Channel

ABSTRACT Leishmania donovani is the causing agent of visceral leishmaniasis, a common infection that affects millions of people from the most underdeveloped countries. Miltefosine is the only oral drug to treat infections caused by L. donovani. Nevertheless, its mechanism of action is not well understood. While miltefosine inhibits the synthesis of phosphatidylcholine and also affects the parasite mitochondrion, inhibiting the cytochrome c oxidase, it is to be expected that this potent drug also produces its effect through other targets. In this context, it has been reported that the disruption of the intracellular Ca2+ homeostasis represents an important object for the action of drugs in trypanosomatids. Recently, we have described a plasma membrane Ca2+ channel in Leishmania mexicana, which is similar to the L-type voltage-gated Ca2+ channel (VGCC) present in humans. Remarkably, the parasite Ca2+ channel is activated by sphingosine, while the L-type VGCC is not affected by this sphingolipid. In the present work we demonstrated that, similarly to sphingosine, miltefosine is able to activate the plasma membrane Ca2+ channel from L. donovani. Interestingly, nifedipine, the classical antagonist of the human channel, was not able to fully block the parasite plasma membrane Ca2+ channel, indicating that the mechanism of interaction is not identical to that of sphingosine. In this work we also show that miltefosine is able to strongly affect the acidocalcisomes from L. donovani, inducing the rapid alkalinization of these important organelles. In conclusion, we demonstrate two new mechanisms of action of miltefosine in L. donovani, both related to disruption of parasite Ca2+ homeostasis.