In vitro‘time-to-kill’ assay to assess the cidal activity dynamics of current reference drugs againstLeishmania donovaniandLeishmania infantum

Maes, Louis; Beyers, Jolien; Mondelaers, Annelies; Kerkhof, Magali Van den; Eberhardt, E.; Caljon, Guy; Hendrickx, Sarah

doi:10.1093/jac/dkw409

Cited by 22 publications

(16 citation statements)

References 16 publications

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“…The high deposition of AmB in both kinds of skin observed in this study could indicate a long availability, as commonly required. The achieved levels of AmB retained in both kinds of skin resulted in a higher value than the IC 50 of AmB for amastigotes, which is considered a goal for successful treatment [45].…”

Section: Discussionmentioning

confidence: 99%

Topical Amphotericin B Semisolid Dosage Form for Cutaneous Leishmaniasis: Physicochemical Characterization, Ex Vivo Skin Permeation and Biological Activity

et al. 2020

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Amphotericin B (AmB) is a potent antifungal successfully used intravenously to treat visceral leishmaniasis but depending on the Leishmania infecting species, it is not always recommended against cutaneous leishmaniasis (CL). To address the need for alternative topical treatments of CL, the aim of this study was to elaborate and characterize an AmB gel. The physicochemical properties, stability, rheology and in vivo tolerance were assayed. Release and permeation studies were performed on nylon membranes and human skin, respectively. Toxicity was evaluated in macrophage and keratinocyte cell lines, and the activity against promastigotes and intracellular amastigotes of Leishmania infantum was studied. The AmB gel remained stable for a period of two months, with optimal properties for topical use and no apparent toxic effect on the cell lines. High amounts of AmB were found in damaged and non-damaged skin (1230.10 ± 331.52 and 2484.57 ± 439.12 µg/g/cm2, respectively) and they were above the IC50 of AmB for amastigotes. Although there were no differences in the in vitro anti-leishmanial activity between the AmB solution and gel, the formulation resulted in a higher amount of AmB being retained in the skin, and is therefore a candidate for further studies of in vivo efficacy.

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Section: Discussionmentioning

confidence: 99%

Topical Amphotericin B Semisolid Dosage Form for Cutaneous Leishmaniasis: Physicochemical Characterization, Ex Vivo Skin Permeation and Biological Activity

et al. 2020

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“…Recently, time–kill experiments for standard anti-leishmanial drugs aimed to identify the minimal exposure time needed to eliminate viable intracellular amastigotes at selected drug concentrations. 10 Here we developed a PD modelling approach, where apparent drug concentrations have been used to estimate EC 50 values. This approach allowed characterization of drug effects over the whole evaluation period, with EC 50 values indicating higher potency of amphotericin B over miltefosine.…”

Section: Discussionmentioning

confidence: 99%

Pharmacodynamics and cellular accumulation of amphotericin B and miltefosine in Leishmania donovani-infected primary macrophages

Voak

Standing

Sepúlveda

et al. 2018

Journal of Antimicrobial Chemotherapy

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ObjectivesWe examined the in vitro pharmacodynamics and cellular accumulation of the standard anti-leishmanial drugs amphotericin B and miltefosine in intracellular Leishmania donovani amastigote–macrophage drug assays.MethodsPrimary mouse macrophages were infected with L. donovani amastigotes. In time–kill assays infected macrophages were exposed to at least six different concentrations of serially diluted drugs and the percentage of infected macrophages was determined after 6, 12, 24, 48, 72 and 120 h of exposure. Cellular drug accumulation was measured following exposure to highly effective drug concentrations for 1, 6, 24, 48 and 72 h. Data were analysed through a mathematical model, relating drug concentration to the percentage of infected cells over time. Host cell membrane damage was evaluated through measurement of lactate dehydrogenase release. The effect of varying the serum and albumin concentrations in medium on the cellular accumulation levels of miltefosine was measured.ResultsAmphotericin B was more potent than miltefosine (EC50 values of 0.65 and 1.26 μM, respectively) and displayed a wider therapeutic window in vitro. The kinetics of the cellular accumulation of amphotericin B was concentration- and formulation-dependent. At an extracellular concentration of 10 μM miltefosine maximum cellular drug levels preceded maximum anti-leishmanial kill. Miltefosine induced membrane damage in a concentration-, time- and serum-dependent manner. Its cellular accumulation levels increased with decreasing amounts of protein in assay medium.ConclusionsWe have developed a novel approach to investigate the cellular pharmacology of anti-leishmanial drugs that serves as a model for the characterization of new drug candidates.

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“…Statistical analysis : Initial scoring system for primary screens consisted of visual examination under microscope and EC 50 values were obtained by nonlinear regression and curves were produced using GraphPad Prism 5.03 (GraphPad, San Diego). Infection indexes for infected BMM were obtained by calculating the average number of parasites per macrophages . At least 100 macrophages were counted per triplicate coverslip.…”

Section: Methodsmentioning

confidence: 99%

“…Infection indexes for infected BMM were obtained by calculating the average number of parasites per macrophages. [21] At least 100 macrophages were counted per triplicate coverslip.…”

Section: Methodsmentioning

confidence: 99%

Fragment‐Based Phenotypic Lead Discovery: Cell‐Based Assay to Target Leishmaniasis

Ayotte

Bilodeau²,

Descoteaux

et al. 2018

ChemMedChem

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A rapid and practical approach for the discovery of new chemical matter for targeting pathogens and diseases is described. Fragment-based phenotypic lead discovery (FPLD) combines aspects of traditional fragment-based lead discovery (FBLD), which involves the screening of small-molecule fragment libraries to target specific proteins, with phenotypic lead discovery (PLD), which typically involves the screening of drug-like compounds in cell-based assays. To enable FPLD, a diverse library of fragments was first designed, assembled, and curated. This library of soluble, low-molecular-weight compounds was then pooled to expedite screening. Axenic cultures of Leishmania promastigotes were screened, and single hits were then tested for leishmanicidal activity against intracellular amastigote forms in infected murine bone-marrow-derived macrophages without evidence of toxicity toward mammalian cells. These studies demonstrate that FPLD can be a rapid and effective means to discover hits that can serve as leads for further medicinal chemistry purposes or as tool compounds for identifying known or novel targets.

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In vitro‘time-to-kill’ assay to assess the cidal activity dynamics of current reference drugs againstLeishmania donovaniandLeishmania infantum

Cited by 22 publications

References 16 publications

Topical Amphotericin B Semisolid Dosage Form for Cutaneous Leishmaniasis: Physicochemical Characterization, Ex Vivo Skin Permeation and Biological Activity

Topical Amphotericin B Semisolid Dosage Form for Cutaneous Leishmaniasis: Physicochemical Characterization, Ex Vivo Skin Permeation and Biological Activity

Pharmacodynamics and cellular accumulation of amphotericin B and miltefosine in Leishmania donovani-infected primary macrophages

Fragment‐Based Phenotypic Lead Discovery: Cell‐Based Assay to Target Leishmaniasis

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