Antileishmanial activity of MDL 28170, a potent calpain inhibitor

d’Avila-Levy, Claudia M.; Marinho, Fernanda A.; Santos, Lívia O.; Martins, Juliana L.; Santos, André Luis Souza dos; Branquinha, Marta H.

doi:10.1016/j.ijantimicag.2006.03.021

Cited by 29 publications

(47 citation statements)

References 21 publications

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“…Our results showed that this calpain inhibitor promoted cellular alterations and arrested the growth of the proliferative forms of both parasites in a dose-dependent manner [7], [14]. Previous works from our group also showed that MDL28170 acted against all the morphological stages found in T. cruzi , without displaying any relevant cytotoxic effect on mammalian host cells [14], [15].…”

Section: Introductionsupporting

confidence: 69%

The Calpain Inhibitor MDL28170 Induces the Expression of Apoptotic Markers in Leishmania amazonensis Promastigotes

et al. 2014

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BackgroundHuman cutaneous leishmaniasis is caused by distinct species, including Leishmania amazonensis. Treatment of cutaneous leishmaniasis is far from satisfactory due to increases in drug resistance and relapses, and toxicity of compounds to the host. As a consequence for this situation, the development of new leishmanicidal drugs and the search of new targets in the parasite biology are important goals.Methodology/Principal FindingsIn this study, we investigated the mechanism of death pathway induced by the calpain inhibitor MDL28170 on Leishmania amazonensis promastigote forms. The combined use of different techniques was applied to contemplate this goal. MDL28170 treatment with IC50 (15 µM) and two times the IC50 doses induced loss of parasite viability, as verified by resazurin assay, as well as depolarization of the mitochondrial membrane, which was quantified by JC-1 staining. Scanning and transmission electron microscopic images revealed drastic alterations on the parasite morphology, some of them resembling apoptotic-like death, including cell shrinking, surface membrane blebs and altered chromatin condensation pattern. The lipid rearrangement of the plasma membrane was detected by Annexin-V labeling. The inhibitor also induced a significant increase in the proportion of cells in the sub-G0/G1 phase, as quantified by propidium iodide staining, as well as genomic DNA fragmentation, detected by TUNEL assay. In cells treated with MDL28170 at two times the IC50 dose, it was also possible to observe an oligonucleossomal DNA fragmentation by agarose gel electrophoresis.Conclusions/SignificanceThe data presented in the current study suggest that MDL28170 induces apoptotic marker expression in promastigotes of L. amazonensis. Altogether, the results described in the present work not only provide a rationale for further exploration of the mechanism of action of calpain inhibitors against trypanosomatids, but may also widen the investigation of the potential clinical utility of calpain inhibitors in the chemotherapy of leishmaniases.

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Section: Introductionsupporting

confidence: 69%

The Calpain Inhibitor MDL28170 Induces the Expression of Apoptotic Markers in Leishmania amazonensis Promastigotes

et al. 2014

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“…The effects of pepstatin A (Sigma Chemical Co., USA) on epimastigotes of T. cruzi were assessed by a method similar to that previously described elsewhere (d' Avila-Levy et al 2006). Briefly, epimastigotes were counted using a Neubauer chamber and resuspended in fresh BHI+FBS medium to a final concentration of 5×10 6 viable epimastigotes per milliliter.…”

Section: Effects Of Pepstatin a On The Growth Rate And Cell Morphologymentioning

confidence: 99%

Multiple effects of pepstatin A on Trypanosoma cruzi epimastigote forms

et al. 2011

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Herein, we have aimed to explore the effects of pepstatin A, a powerful aspartic protease inhibitor, on Trypanosoma cruzi, the etiologic agent of Chagas' disease. Pepstatin A arrested the proliferation of epimastigotes of T. cruzi (clone Dm28c, TcI lineage), in both dose- and time-dependent manner. The IC(50) value was calculated to be 36.2 μM after 96 h of parasite-drug contact. The parasite treatment with pepstatin A resulted in significant morphological alterations, including parasites becoming round in shape, reduction (≈25%) of the parasite size, and parasites presenting parts or the whole flagellum detached from the cell body. Cell lysis was not observed, resulting in a trypanostatic effect. The treatment of different T. cruzi strains, belonging to distinct phylogenetic lineages, with pepstatin A at 36.2 μM resulted in growth inhibition as follows: 28% to Y (TcII), 45% to CL Brener (TcII), 45.4% to 4167 (Z3), and 26.4% to 3663 (Z3) strains. The hydrolysis of a cathepsin D fluorogenic substrate (7-methoxycoumarin-4-acetyl-Gly-Lys-Pro-Ile-Leu-Phe-Phe-Arg-Leu-Lys(DNP)-D: -Arg-amide) by T. cruzi epimastigote extract was inhibited (≈65%) by pepstatin A at 10 μM, suggesting that an aspartic protease may be the intracellular target of this inhibitor. Curiously, pepstatin A induced an increase of 54% and 98%, respectively, in the surface expression of gp63- and calpain-related molecules in epimastigotes, but not in the cruzipain level, as well as stimulated the epimastigote-to-trypomastigote differentiation in a dose-dependent manner. However, approximately 45% of the trypomastigotes had their flagellum detached from the cell body. These results contribute to understand the possible role of aspartic proteases in the physiology of T. cruzi cells, adding new in vitro insights into the possibility of exploiting aspartic protease as promising targets to treat Chagas' disease.

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“…In L. major, it was demonstrated that a calpain related protein (LmCALP20.2) is up regulated in the promastigote insect stage and LmCALP20.1, coded by the adjacent gene, is up regulated in the subsequent metacyclic insect stage [149]. Our group demonstrated that MDL28170, a potent calpain inhibitor, is capable of reducing promastigote growth in culture and induces cell death [150], probable through apoptosis (unpublished data). A proteomics screen implicated a calpain-related protein in drug resistance in L. donovani clinical field isolates, probably by modulating drug-induced apoptosis [151].…”

Section: Cysteine Peptidasesmentioning

confidence: 83%

Biological Roles of Peptidases in Trypanosomatids~!2009-11-26~!2010-02-15~!2010-03-18~!

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Branquinha²,

d’Avila-Levy³

et al. 2010

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Abstract:In this review, we report the recent developments in the characterization of peptidases and their possible biological functions in the Trypanosomatidae family. The focus will be on peptidases from Trypanosoma cruzi, Leishmania spp., African trypanosomes and plant and insect trypanosomatids. There are numerous events in parasite development where the involvement of peptidases has been established, and they will be approached in the present review. Also in this review we will discuss the central roles have been proposed for peptidases in diverse processes such as virulence, host cell interaction and invasion, catabolism of host proteins, differentiation, cell cycle progression and both stimulation and evasion of host immune responses.

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Antileishmanial activity of MDL 28170, a potent calpain inhibitor

Cited by 29 publications

References 21 publications

The Calpain Inhibitor MDL28170 Induces the Expression of Apoptotic Markers in Leishmania amazonensis Promastigotes

The Calpain Inhibitor MDL28170 Induces the Expression of Apoptotic Markers in Leishmania amazonensis Promastigotes

Multiple effects of pepstatin A on Trypanosoma cruzi epimastigote forms

Biological Roles of Peptidases in Trypanosomatids~!2009-11-26~!2010-02-15~!2010-03-18~!

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