Sitamaquine Sensitivity in Leishmania Species Is Not Mediated by Drug Accumulation in Acidocalcisomes

Luque‐Ortega

López-Martín

et al. 2011

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The 8-aminoquinoline analogue sitamaquine (SQ) is an oral antileishmanial drug currently undergoing phase 2b clinical trials for the treatment of visceral leishmaniasis. In the present study, we investigated the mechanism of action of this drug in Leishmania donovani promastigotes. SQ causes a dose-dependent inhibition of complex II (succinate dehydrogenase) of the respiratory chain in digitonin-permeabilized promastigotes, together with a drop in intracellular ATP levels and a decrease of the mitochondrial electrochemical potential. This is associated with increases of reactive oxygen species and intracellular Ca 2؉ levels, a higher percentage of the population with sub-G 1 DNA content, and exposure of phosphatidylserine. Taken together, these results support a lethal mechanism for SQ that involves inhibition of the respiratory chain complex II, which in turn triggers oxidative stress and finally leads to an apoptosis-like death of Leishmania parasites.Leishmaniasis, a protozoal infectious disease caused by a set of 17 species of the genus Leishmania, shows a wide spectrum of clinical manifestations, including, in order of increasing severity, cutaneous (CL), mucocutaneous (MCL), and visceral (VL) leishmaniasis (3). Although chemotherapy is the only current treatment option for leishmaniasis, its efficacy is increasingly limited by growing resistance to first-line drugs, especially antimonials, by the frequent side effects associated with their use, and by the high cost of treatment (30). The paucity of new drugs in the pipeline, together with the poor definition of Leishmania targets for the drugs in current clinical use, represents an additional concern for current chemotherapy. Solutions to curb this pessimistic scenario rely on combination therapy (40) and the rescue of old drugs, such as paromomycin (17, 36) and sitamaquine (SQ) (39), that were previously discarded.8-Aminoquinolines are an important class of antiparasitic agents (37) with broad application and excellent efficacy but with limitations due to their hematological toxicity (primarily metahemoglobinemia and hemolysis). SQ, formerly known as WR6026, is an 8-aminoquinoline that was initially developed by the Walter Reed Army Institute (46). The results of phase 2b clinical trials of this drug against VL in India (16) and Kenya (45) by GlaxoSmithKline were encouraging. These results, together with its oral administration, represent a substantial advantage in terms of its future widespread implementation.The targets for SQ remain elusive. Entry of SQ into the parasite starts with an electrostatic interaction with anionic phospholipids of the plasma membrane (11). In a seminal work, Vercesi and Docampo (43) observed a loss of mitochondrial electrochemical potential in digitonin-permeabilized parasites after SQ addition, together with alkalinization of acidocalcisomes (44), which also underwent a privileged SQ accumulation, although no correlation was found with its toxicity (18).Herein we provide further insight into the leishmanicidal mechanism of S...

Section: Discussionmentioning

confidence: 99%

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confidence: 99%

The 8-Aminoquinoline Analogue Sitamaquine Causes Oxidative Stress in Leishmania donovani Promastigotes by Targeting Succinate Dehydrogenase

Luque‐Ortega

López-Martín

et al. 2011

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“…Late-stage promastigotes from the WT and R4 lines were used to infect mouse peritoneal macrophages from BALB/c mice (Charles River, Ltd.) at a macrophage/parasite ratio of 1:10, as described previously (14). After infection for 6 h, extracellular parasites were removed by washing with serum-free medium.…”

Section: Chemicalmentioning

confidence: 99%

“…Of the new drugs under development, 8-aminoquinolines such as sitamaquine (WR6026; GlaxoSmithKline), which is currently in phase 2b clinical trials (12,34), represent a promising new oral leishmanicidal treatment. Although the mechanism of action of sitamaquine is still unknown, it has been reported that acidocalcisomes play a key role in the accumulation of sitamaquine, although they do not determine the leishmanicidal potency of the drug (14). Other 8-aminoquinolines chemically related to primaquine have been synthesized and evaluated for their antiparasitic activity (1,25).…”

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confidence: 99%

Increased Glycolytic ATP Synthesis Is Associated with Tafenoquine Resistance inLeishmania major

Manzano

Pérez‐Victoria

et al. 2011

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Tafenoquine (TFQ), an 8-aminoquinoline used to treat and prevent Plasmodium infections, could represent an alternative therapy for leishmaniasis. Indeed, TFQ has shown significant leishmanicidal activity both in vitro and in vivo, where it targets Leishmania mitochondria and activates a final apoptosis-like process. In order not to jeopardize the life span of this potential antileishmania drug, it is important to determine the likelihood that Leishmania will develop resistance to TFQ and the mechanisms of resistance induced. To address this issue, a TFQ-resistant Leishmania major promastigote line (R4) was selected. This resistance, which is unstable in a drug-free medium (revertant line), was maintained in intramacrophage amastigote forms, and R4 promastigotes were found to be cross-resistant to other 8-aminoquinolines. A decreased TFQ uptake, which is probably associated with an alkalinization of the intracellular pH rather than drug efflux, was observed for both the R4 and revertant lines. TFQ induces a decrease in ATP synthesis in all Leishmania lines, although total ATP levels were maintained at higher values in R4 parasites. In contrast, ATP synthesis by glycolysis was significantly increased in R4 parasites, whereas mitochondrial ATP synthesis was similar to that in wild-type parasites. We therefore conclude that increased glycolytic ATP synthesis is the main mechanism underlying TFQ resistance in Leishmania.

“…Using a spectrofluorometric assay, we found that the accumulation of TFQ in T. brucei BSF was 50% lower than in promastigotes of L. major (P Ͻ 0.05), this being despite its higher activity against T. brucei. The absence of correlation between susceptibility and uptake level was also reported for the case of sitamaquine, a related 8-aminoquinoline, against different Leishmania species, (25) and for several analogs of the antitrypanosomal diamidines DB75 and DB820 against T. brucei (26).…”

Section: Resultsmentioning

confidence: 99%

The Oral Antimalarial Drug Tafenoquine Shows Activity against Trypanosoma brucei

Martínez‐García

Pérez-Victoria

et al. 2015

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cThe protozoan parasite Trypanosoma brucei causes human African trypanosomiasis, or sleeping sickness, a neglected tropical disease that requires new, safer, and more effective treatments. Repurposing oral drugs could reduce both the time and cost involved in sleeping sickness drug discovery. Tafenoquine (TFQ) is an oral antimalarial drug belonging to the 8-aminoquinoline family which is currently in clinical phase III. We show here that TFQ efficiently kills different T. brucei spp. in the submicromolar concentration range. Our results suggest that TFQ accumulates into acidic compartments and induces a necrotic process involving cell membrane disintegration and loss of cytoplasmic content, leading to parasite death. Cell lysis is preceded by a wide and multitarget drug action, affecting the lysosome, mitochondria, and acidocalcisomes and inducing a depolarization of the mitochondrial membrane potential, elevation of intracellular Ca 2؉ , and production of reactive oxygen species. This is the first report of an 8-aminoquinoline demonstrating significant in vitro activity against T. brucei.