Mechanism of Action of Miltefosine on Leishmania donovani Involves the Impairment of Acidocalcisome Function and the Activation of the Sphingosine-Dependent Plasma Membrane Ca ²⁺ Channel

Abstract

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 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 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 Ca homeostasis represents an important object for the action of drugs in trypanosomatids. Recently, we have described a plasma membrane Ca channel in , which is similar to the L-type voltage-gated Ca channel (VGCC) present in humans. Remarkably, the parasite Ca 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 Ca channel from Interestingly, nifedipine, the classical antagonist of the human channel, was not able to fully block the parasite plasma membrane Ca 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 , inducing the rapid alkalinization of these important organelles. In conclusion, we demonstrate two new mechanisms of action of miltefosine in, both related to disruption of parasite Ca homeostasis.

Identification and electrophysiological properties of a sphingosine‐dependent plasma membrane Ca2+ channel in Trypanosoma cruzi

“…In Fig. , the residues involved in the binding of dihydropyridines (DHPs) are shown, and their essentialness were identified . The colored amino acids show the conserved aligned residues of the three studied sequences and in black the substitution of a nonconserved change in at least one of the sequences in a specific position.…”

Lipase Precursor-Like Protein Promotes Miltefosine Tolerance in Leishmania donovani by Enhancing Parasite Infectivity and Eliciting Anti-inflammatory Responses in Host Macrophages

“…Briefly, primary peritoneal macrophages derived from 6-week-old female BALB/c mice were infected with 6-day-old LdLip ϩϩ , LdNeo, or LdWT promastigotes at a ratio of 10:1 (parasites/macrophages) in 200 l complete RPMI 1640 medium in 8-well chambered slides and incubated for 16 h at 37°C in 5% CO 2 . Infected macrophages were washed and incubated further for 48 h, with increasing concentrations of either MIL (1, 5, 10, 20, and 30 M), AmB (0.027, 0.054, 0.108, 0.539, and 2.0 M), or SAG (1,5,10,20,30, and 40 g/ml). Macrophages were stained with Diff-Quik solutions and subsequently examined for intracellular amastigotes.…”

“…MIL induces apoptosis-like cell death in Leishmania, and inhibition of apoptotic cell death has been documented in MIL-resistant parasites (7)(8)(9). MIL plays a role in impairment of acidocalcisome function, activation of the sphingosinedependent plasma membrane Ca 2ϩ channel, and inhibition of cytochrome c oxidase in L. donovani (10,11). Leishmania undergoes metabolic reconfiguration during oxidative stress to resist reactive oxygen species (ROS) (12).…”

Aktuelle Kenntnisse zu Therapie und Prävention der kaninen Leishmaniose

“…While they do not have similar action mechanisms, they both induce leishmanicidal activity (39). The action mechanism of miltefosine is not clearly understood and multiple mechanisms are proposed (7), the most recent of which being parasite calcium homeostasis (45). This can reduce the parasite burden and improve clinical signs, although relapses have been reported when miltefosine has been administered alone, and therefore combination with allopurinol is preferable (27,28,32).…”