Regulatory volume decrease in Leishmania mexicana: effect of anti-microtubule drugs

Dagger, Fracehuli; Valdivieso, Elizabeth; Marcano, Ana Karina; Ayesta, C.

doi:10.1590/s0074-02762013000100014

Cited by 3 publications

(3 citation statements)

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“…Quantification demonstrated a ∼1.5-fold increase in flagella length (Figure S1B, p < 0.05 by ANOVA compared to DMSO control) when promastigotes were treated with MMV676477, paclitaxel, or active analogs for 24 h at their respective EC 50 72h . We also found that promastigotes treated with paclitaxel, MMV676477, or active analogs at their respective EC 50 72h for 48 h were more rounded or pear-shaped (defined as parasites with a cell body width that was ≥70% of the cell body length 29 ) than DMSO controls or promastigotes treated with the unrelated drug, miltefosine (Figure S1C). Quantification demonstrated a 2-fold increase in pear-shaped promastigotes in paclitaxel, MMV676477, and active analog-treated groups (Figure S1D, p < 0.05 by ANOVA compared to DMSO control).…”

Section: Screening the Pathogen Box Identifies Multiple Hitmentioning

confidence: 79%

“…Since we obtained MMV676477 from a phenotypic screen, our next goal was to identify its likely target. Upon review of the literature, we noted that MMV676477-treated L. amazonensis microscopically resembled Leishmania treated with the anticancer drug paclitaxel (“Taxol”, EC 50 72h for La = 760 ± 8.3 nM) . Paclitaxel stabilizes microtubules and prevents mitosis in cancer cells .…”

Section: Resultsmentioning

confidence: 99%

“…Upon review of the literature, we noted that MMV676477-treated L. amazonensis microscopically re-sembled Leishmania treated with the anticancer drug paclitaxel ("Taxol", EC 50 72h for La = 760 ± 8.3 nM). 29 Paclitaxel stabilizes microtubules and prevents mitosis in cancer cells. 30 We therefore used paclitaxel as a benchmark positive control for antimitotic activity and an unrelated antileishmanial drug, miltefosine, as a negative control.…”

Section: Screening the Pathogen Box Identifies Multiple Hitmentioning

confidence: 99%

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An Antiparasitic Compound from the Medicines for Malaria Venture Pathogen Box Promotes Leishmania Tubulin Polymerization

et al. 2020

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The few frontline antileishmanial drugs are poorly effective and toxic. To search for new drugs for this neglected tropical disease, we tested the activity of compounds in the Medicines for Malaria Venture (MMV) “Pathogen Box” against Leishmania amazonensis axenic amastigotes. Screening yielded six discovery antileishmanial compounds with EC50 values from 50 to 480 nM. Concentration–response assays demonstrated that the best hit, MMV676477, had mid-nanomolar cytocidal potency against intracellular Leishmania amastigotes, Trypanosoma brucei, and Plasmodium falciparum, suggesting broad antiparasitic activity. We explored structure–activity relationships (SAR) within a small group of MMV676477 analogs and observed a wide potency range (20–5000 nM) against axenic Leishmania amastigotes. Compared to MMV676477, our most potent analog, SW41, had ∼5-fold improved antileishmanial potency. Multiple lines of evidence suggest that MMV676477 selectively disrupts Leishmania tubulin dynamics. Morphological studies indicated that MMV676477 and analogs affected L. amazonensis during cell division. Differential centrifugation showed that MMV676477 promoted partitioning of cellular tubulin toward the polymeric form in parasites. Turbidity assays with purified Leishmania and porcine tubulin demonstrated that MMV676477 promoted leishmanial tubulin polymerization in a concentration-dependent manner. Analogs’ antiparasitic activity correlated with their ability to facilitate purified Leishmania tubulin polymerization. Chemical cross-linking demonstrated binding of the MMV676477 scaffold to purified Leishmania tubulin, and competition studies established a correlation between binding and antileishmanial activity. Our studies demonstrate that MMV676477 is a potent antiparasitic compound that preferentially promotes Leishmania microtubule polymerization. Due to its selectivity for and broad-spectrum activity against multiple parasites, this scaffold shows promise for antiparasitic drug development.

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Section: Screening the Pathogen Box Identifies Multiple Hitmentioning

confidence: 79%

Section: Resultsmentioning

confidence: 99%

Section: Screening the Pathogen Box Identifies Multiple Hitmentioning

confidence: 99%

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An Antiparasitic Compound from the Medicines for Malaria Venture Pathogen Box Promotes Leishmania Tubulin Polymerization

et al. 2020

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Anti-leishmanial effect of spiro dihydroquinoline-oxindoles on volume regulation decrease and sterol biosynthesis of Leishmania braziliensis

Leañez

Núñez

García-Marchán

et al. 2019

Experimental Parasitology

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Morphogenesis Dynamics in Leishmania Differentiation

2022

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Leishmania, the causative agent of leishmaniasis, is an obligatory intracellular parasite that cycles between phagolysosome of mammalian macrophages, where it resides as round intracellular amastigotes, and the midgut of female sandflies, where it resides as extracellular elongated promastigotes. This protozoan parasite cytoskeleton is composed of stable and abundant subpellicular microtubules (SPMT). This study aims to determine the kinetics of developmental morphogenesis and assess whether microtubules remodelling is involved in this process. Using image-streaming technology, we observed that rounding of promastigotes during differentiation into amastigotes was initiated promptly after exposure to the differentiation signal. Stabilizing microtubules with taxol sped rounding, but later killed differentiating parasites if taxol was not removed. Microtubule destabilizers such as vinblastine had no effect on the rate of rounding, nor on the viability of differentiating parasites. In the reverse process, elongation is initiated after a delay of 7.5 and completed 72 h after exposure to the amastigote to the promastigote differentiation signal. During the delay, parasites became highly sensitive to treatment with microtubule destabilizers. The addition of vinblastine during the first 7.5 h halted differentiation and killed parasites. Between 8 and 24 h, parasites gradually became resistant to vinblastine and, in parallel, started to elongate. In contrast, taxol had no effect on parasite elongation, nor on the viability of these cells. In a parallel study, we showed that the Leishmania-specific protein kinase A (PKA) holoenzyme containing the LdPKAR3-C3 complex is essential for promastigote elongation. Mutant promastigotes lacking either of these proteins are round, but maintain their flagella. Here, we observed that during differentiation into amastigotes, these mutants round at the same rate as the wild type, but never exceed the WT density of round amastigotes. In the reverse process, these mutants undergo the same initial delay and then elongate at the same rate as the WT. They stop elongating when they reach 20% of elongated cells in mature promastigotes. Our analysis indicates that while promastigote rounding into amastigotes did not require microtubule remodelling, morphogenesis of round amastigotes into elongated promastigotes required microtubule rearrangement before elongation was initiated. This is the first study that investigates the dynamics of microtubules during parasite development.

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Regulatory volume decrease in Leishmania mexicana: effect of anti-microtubule drugs

Cited by 3 publications

References 37 publications

An Antiparasitic Compound from the Medicines for Malaria Venture Pathogen Box Promotes Leishmania Tubulin Polymerization

An Antiparasitic Compound from the Medicines for Malaria Venture Pathogen Box Promotes Leishmania Tubulin Polymerization

Anti-leishmanial effect of spiro dihydroquinoline-oxindoles on volume regulation decrease and sterol biosynthesis of Leishmania braziliensis

Morphogenesis Dynamics in Leishmania Differentiation

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