Rapid fluorescent assay for screening drugs on Leishmania amastigotes

Shimony, Orly; Jaffe, Charles L.

doi:10.1016/j.mimet.2008.05.026

Cited by 57 publications

(50 citation statements)

References 28 publications

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“…Derivatives (12,18,19,21 and 25) that showed the best antileishmanial activity on microplate assay at 50 mM against L. donovani axenic amastigotes were selected for further investigations: GI 50 values, cytotoxicity to the macrophage cell line THP-1 and anti-leishmanial activity against the L. donovani-infected macrophage cell line THP-1 were evaluated. Betulonic acid 18 showed the best GI 50 value of 14.6 mM on microplate assay against L. donovani axenic amastigotes, followed by L-aspartyl amide derivative 21 and oxime derivative 25, with GI 50 values of 21.2 and 22.8 mM, respectively (Table 1).…”

Section: Resultsmentioning

confidence: 99%

“…18 Anti-leishmanial activities of betulin derivatives were screened using a fluorescent viability microplate assay with L. donovani (MHOM/SD/1962/1S-Cl2d) axenic amastigotes and alamarBlue (resazurin, AbD Serotec, Oxford, UK) as described previously. [19][20][21] Initial screening was carried out by assessing the inhibition of amastigote growth at 50 mM of betulin derivative. All compounds were tested at least twice in triplicate.…”

Section: Methodsmentioning

confidence: 99%

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Anti-leishmanial activity of betulin derivatives

et al. 2010

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Leishmanicidal activity of 24 derivatives of naturally occurring and abundant triterpenes belonging to the lupane series, betulin, betulinic acid and betulonic acid, is described in this study. The easily modified positions of the lupane skeleton, the hydroxy groups of C-3 and C-28, as well as the carbon-carbon double bond C-20-C-29 were used as a starting point to prepare a library of triterpenoid derivatives for bioactivity studies. The compounds were evaluated against Leishmania donovani axenic amastigotes on a microplate assay at 50 lM. GI 50 values of the most effective compounds were evaluated, as well as their cytotoxicity on the human macrophage cell line THP-1, and anti-leishmanial activity against L. donovani-infected THP-1 macrophages was determined. Betulonic acid was the most potent derivative, yielding a GI 50 value of 14.6 lM. Promising and distinct structure-activity relationships were observed, and these compounds can be regarded as significant lead molecules for further improvement and optimization.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Anti-leishmanial activity of betulin derivatives

et al. 2010

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“…Compounds were screened for leishmanicidal activity using an alamarBlue (AbD Serotec) viability assay essentially as described (43); modification and materials are given in SI Materials and Methods.…”

Section: Methodsmentioning

confidence: 99%

Identification of the molecular attributes required for aminoglycoside activity against Leishmania

Shalev

Kondo

Kopelyanskiy³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Leishmaniasis, a parasitic disease caused by protozoa of the genus Leishmania , affects millions of people worldwide. Aminoglycosides are mostly known as highly potent, broad-spectrum antibiotics that exert their antibacterial activity by selectively targeting the decoding A site of the bacterial ribosome, leading to aberrant protein synthesis. Recently, some aminoglycosides have been clinically approved and are currently used worldwide for the treatment of leishmaniasis; however the molecular details by which aminoglycosides induce their deleterious effect on Leishmaina is still rather obscure. Based on high conservation of the decoding site among all kingdoms, it is assumed that the putative binding site of these agents in Leishmania is the ribosomal A site. However, although recent X-ray crystal structures of the bacterial ribosome in complex with aminoglycosides shed light on the mechanism of aminoglycosides action as antibiotics, no such data are presently available regarding their binding site in Leishmania . We present crystal structures of two different aminoglycoside molecules bound to a model of the Leishmania ribosomal A site: Geneticin (G418), a potent aminoglycoside for the treatment of leishmaniasis at a 2.65-Å resolution, and Apramycin, shown to be a strong binder to the leishmanial ribosome lacking an antileishmanial activity at 1.4-Å resolution. The structural data, coupled with in vitro inhibition measurements on two strains of Leishmania , provide insight as to the source of the difference in inhibitory activity of different Aminoglycosides. The combined structural and physiological data sets the ground for rational design of new, and more specific, aminoglycoside derivatives as potential therapeutic agents against leishmaniasis.

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“…A quantitative colorimetric assay using the oxidation-reduction indicator Alamar Blue was developed to measure cytotoxicity of the synthesized compounds against the protozoan parasite Leishmania donovani. The Alamar Blue assay permits a simple, reproducible and reliable method for screening antileishmanial drugs (Judith and Dietmar, 2001;Shimony and Jaffe, 2008;Nakayama et al, 1997).…”

Section: In Vitro Antipromastigote Assaymentioning

confidence: 99%