Paromomycin: uptake and resistance in Leishmania donovani
Anupam Jhingran1, Bhavna Chawla, Shailendra Saxena
1School of Life Sciences, Jawaharlal Nehru University, New Delhi, India.
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 1mM 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 72h 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.
Insights
Paromomycin uptake by Leishmania donovani involves initial non-saturable binding to the parasite surface. This leads to decreased mitochondrial potential and inhibited protein synthesis, crucial for its leishmaniasis treatment mechanism.
Area of Science:
- Parasitology
- Molecular Biology
- Pharmacology
Background:
- Leishmaniasis is a significant global health concern.
- Paromomycin is an aminoglycoside antibiotic undergoing clinical trials for leishmaniasis treatment.
- Understanding paromomycin's mechanism of action in Leishmania parasites is critical for optimizing therapy.
Purpose of the Study:
- To elucidate the mechanism of paromomycin uptake in Leishmania donovani.
- To determine the effects of paromomycin on parasite viability and key cellular processes.
- To investigate the basis of drug resistance in Leishmania parasites.
Main Methods:
- In vitro sensitivity testing of Leishmania donovani promastigotes and amastigotes.
- Drug association assays at varying drug concentrations, temperatures, and in the presence of metabolic inhibitors.
- Measurement of mitochondrial membrane potential and protein synthesis inhibition.
- Characterization of drug-resistant parasite strains.
Main Results:
- Paromomycin association with L. donovani exhibited a rapid, non-saturable initial phase, attributed to binding with the negatively charged glycocalyx.
- Prolonged exposure (72h) significantly decreased mitochondrial membrane potential and inhibited both cytoplasmic and mitochondrial protein synthesis.
- Resistant strains displayed reduced paromomycin accumulation, decreased initial surface binding, and diminished drug-induced mitochondrial and protein synthesis impairment.
Conclusions:
- Paromomycin's anti-leishmanial effect is linked to its uptake via non-specific surface binding, subsequent mitochondrial targeting, and inhibition of protein synthesis.
- Reduced drug accumulation and surface binding are key mechanisms underlying paromomycin resistance in Leishmania donovani.
- These findings provide insights into paromomycin's therapeutic action and resistance development, informing future treatment strategies.
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