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Altered drug membrane permeability in a multidrug-resistant Leishmania tropica line
M J Chiquero1, J M Pérez-Victoria, F O'Valle
1Instituto de Parasitología y Biomedicina Lopez-Neyra, Consejo Superior de Investigaciones Científicas, Granada, Spain.
Abstract:
We selected a Leishmania tropica cell line resistant to daunomycin (DNM) that presents a multidrug-resistant (MDR) phenotype characterized by overexpression of a P-glycoprotein of 150 kDa. The resistant line overexpressed an MDR-like gene, called ltrmdr1, located in an extrachromosomal circular DNA. DNM uptake experiments using laser flow cytometry showed a significant reduction in drug accumulation in the resistant parasites. The initial stages of the interaction of DNM with membranes from wild-type and DNM-resistant parasites were defined by a rapid kinetic stopped-flow procedure which can be described by two kinetic components. On the basis of a previous similar kinetic study with tumor cells, we ascribed the fast component to rapid interaction of DNM with membrane surface components and the slow component to passive diffusion of the drug across the membranes. The results reported here indicate that entrance of DNM into wild-type parasites was facilitated in respect to the resistant ones. We propose that resistance to DNM in L. tropica is a multifactorial event involving at least two complementary mechanisms. an altered drug membrane permeability and the overexpression of a protein related to P-glycoprotein that regulates drug efflux.
Insights
Leishmania tropica parasites resistant to daunomycin (DNM) exhibit multidrug resistance (MDR) due to altered membrane permeability and P-glycoprotein overexpression, reducing drug accumulation.
Area of Science:
- Parasitology
- Molecular Biology
- Biochemistry
Background:
- Leishmania tropica is a parasite causing leishmaniasis.
- Multidrug resistance (MDR) is a significant challenge in treating parasitic infections.
- Daunomycin (DNM) is an anthracycline antibiotic with potential antiparasitic activity.
Purpose of the Study:
- To investigate the mechanisms of daunomycin (DNM) resistance in Leishmania tropica.
- To characterize the molecular basis of the multidrug-resistant (MDR) phenotype in L. tropica.
- To elucidate the role of P-glycoprotein and membrane permeability in DNM resistance.
Main Methods:
- Selection of a DNM-resistant Leishmania tropica cell line.
- Analysis of gene expression and localization (ltrmdr1 gene on extrachromosomal DNA).
- Quantification of DNM uptake using laser flow cytometry.
- Kinetic analysis of DNM-membrane interaction via stopped-flow spectroscopy.
Main Results:
- The resistant L. tropica line overexpressed a 150 kDa P-glycoprotein and an MDR-like gene (ltrmdr1) on extrachromosomal DNA.
- DNM accumulation was significantly reduced in resistant parasites compared to wild-type.
- Kinetic studies revealed altered DNM-membrane interactions, suggesting reduced passive diffusion in resistant cells.
Conclusions:
- DNM resistance in L. tropica is a multifactorial phenomenon.
- Mechanisms include altered drug membrane permeability and overexpression of a P-glycoprotein-related efflux pump.
- These findings contribute to understanding and potentially overcoming drug resistance in Leishmania parasites.