Effects of buthionine sulfoximine nifurtimox and benznidazole upon trypanothione and metallothionein proteins in Trypanosoma cruzi.

Maya, Juan Diego; Rodríguez, Andrés; Pino, Laura; Pabón, Adriana; Ferreira, Jorge; Pavani, Mario; Repetto, Yolanda; Morello, Antonio

doi:10.4067/s0716-97602004000100007

Cited by 19 publications

(16 citation statements)

References 25 publications

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“…However, since these effects were observed to the same extent in CL Brener and Nicaragua, we may infer (i) that the IC 50 differences between the two strains are not due to these alterations and (ii) that Bz reduction by NADH and NADPH-dependent reductases may not generate ROS. This result is in agreement with previous reports demonstrating that the trypanocidal effect of Bz is exerted by reduced metabolites of the drug interacting with macromolecules from the parasite rather than by ROS formation and ⌬⌿m (17,18,28).…”

Section: Discussionsupporting

confidence: 83%

Putative Role of the Aldo-Keto Reductase from Trypanosoma cruzi in Benznidazole Metabolism

Garavaglia

Laverrière

Cannata

et al. 2016

Antimicrob Agents Chemother

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b Benznidazole (Bz), the drug used for treatment of Chagas' disease (caused by the protozoan Trypanosoma cruzi), is activated by a parasitic NADH-dependent type I nitroreductase (NTR I). However, several studies have shown that other enzymes are involved. The aim of this study was to evaluate whether the aldo-keto reductase from T. cruzi (TcAKR), a NADPH-dependent oxido-reductase previously described by our group, uses Bz as the substrate. We demonstrated that both recombinant and native TcAKR enzymes reduce Bz by using NADPH, but not NADH, as a cofactor. TcAKR-overexpressing epimastigotes showed higher NADPH-dependent Bz reductase activity and a 50% inhibitory concentration (IC 50 ) value for Bz 1.8-fold higher than that of the controls, suggesting that TcAKR is involved in Bz detoxification instead of activation. To understand the role of TcAKR in Bz metabolism, we studied TcAKR expression and NADPH/NADH-dependent Bz reductase activities in two T. cruzi strains with differential susceptibility to Bz: CL Brener and Nicaragua. Taking into account the results obtained with TcAKR-overexpressing epimastigotes, we expected the more resistant strain, Nicaragua, to have higher TcAKR levels than CL Brener. However, the results were the opposite. CL Brener showed 2-fold higher TcAKR expression and 5.7-fold higher NADPH-Bz reduction than the Nicaragua strain. In addition, NADH-dependent Bz reductase activity, characteristic of NTR I, was also higher in CL Brener than in Nicaragua. We conclude that although TcAKR uses Bz as the substrate, TcAKR activity is not a determinant of Bz resistance in wild-type strains and may be overcome by other enzymes involved in Bz activation, such as NADPH-and NADH-dependent reductases.C hagas' disease, caused by the protozoan Trypanosoma cruzi, affects 11 million people in Latin America (1). The parasite is usually transmitted to humans and other mammals by triatomine bugs. However, transmission can also be oral (through contaminated food), congenital, or transfusional or by organ transplantation, even in countries where the disease is not endemic (2). The course of infection includes an acute phase followed by an indeterminate phase without symptoms, which lasts throughout life in most infected people. Approximately 30% of patients can develop a chronic phase characterized by cardiac and/or digestive pathologies that can lead to death (3).Benznidazole (Bz) and, to a lesser extent, nifurtimox (Nx) are the drugs currently used for treatment of T. cruzi infection (4). Although the mechanism of action of these nitroheterocyclic compounds is not clearly understood, it is known that, to exert their trypanocidal effects, they need to be activated through the reduction of their nitro group (5). Several studies have proposed that an NADH-dependent trypanosomal type I nitroreductase (NTR I) is a key enzyme which catalyzes Bz and Nx activation in vivo (6,7,8,9). However, evidence indicates that although Bz is reduced to its anion radical, redox cycling may not be relevant for its mode of action, as ...

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Section: Discussionsupporting

confidence: 83%

Putative Role of the Aldo-Keto Reductase from Trypanosoma cruzi in Benznidazole Metabolism

Garavaglia

Laverrière

Cannata

et al. 2016

Antimicrob Agents Chemother

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“…through glutathione and trypanothione-S-transferases) and non-enzymatic production of these thiol conjugates is not certain, but non-enzymatic conjugation to macromolecular thiols, e.g., cysteine in proteins, is also likely to contribute to the mode of action of the drug as has been reported for metronidazole and other nitroimidazoles [55], [56]. Low molecular weight cysteine-rich proteins in T. cruzi were shown to have these residues blocked when exposed to Nfx or Bzn, again indicating binding of metabolites from these drugs to macromolecular cysteine residues [57]. Many enzymes rely on active cysteine residues to exert their functions including the tryparedoxin peroxidase-tryparedoxin system, among others, and could be targeted by Bzn metabolites [58].…”

Section: Discussionmentioning

confidence: 78%

Benznidazole Biotransformation and Multiple Targets in Trypanosoma cruzi Revealed by Metabolomics

et al. 2014

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BackgroundThe first line treatment for Chagas disease, a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi, involves administration of benznidazole (Bzn). Bzn is a 2-nitroimidazole pro-drug which requires nitroreduction to become active, although its mode of action is not fully understood. In the present work we used a non-targeted MS-based metabolomics approach to study the metabolic response of T. cruzi to Bzn.Methodology/Principal findingsParasites treated with Bzn were minimally altered compared to untreated trypanosomes, although the redox active thiols trypanothione, homotrypanothione and cysteine were significantly diminished in abundance post-treatment. In addition, multiple Bzn-derived metabolites were detected after treatment. These metabolites included reduction products, fragments and covalent adducts of reduced Bzn linked to each of the major low molecular weight thiols: trypanothione, glutathione, γ-glutamylcysteine, glutathionylspermidine, cysteine and ovothiol A. Bzn products known to be generated in vitro by the unusual trypanosomal nitroreductase, TcNTRI, were found within the parasites, but low molecular weight adducts of glyoxal, a proposed toxic end-product of NTRI Bzn metabolism, were not detected.Conclusions/significanceOur data is indicative of a major role of the thiol binding capacity of Bzn reduction products in the mechanism of Bzn toxicity against T. cruzi.

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“…In mammalian, the nitro group is reduced to an amino group by a type II nitroreductase, with formation of free radical intermediaries and reactive oxygen species (ROS) [4]–[6]. BZL exerts its trypanocidal effect against all forms of the parasite (intra or extracellular) through these metabolites that likely bind to parasite macromolecules [7], [8].…”

Section: Introductionmentioning

confidence: 99%

Regulation of Biotransformation Systems and ABC Transporters by Benznidazole in HepG2 Cells: Involvement of Pregnane X-Receptor

et al. 2012

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BackgroundBenznidazole (BZL) is the only antichagasic drug available in most endemic countries. Its effect on the expression and activity of drug-metabolizing and transporter proteins has not been studied yet.Methodology/Principal FindingsExpression and activity of P-glycoprotein (P-gp), Multidrug resistance-associated protein 2 (MRP2), Cytochrome P450 3A4 (CYP3A4), and Glutathione S-transferase (GST) were evaluated in HepG2 cells after treatment with BZL. Expression was estimated by immunoblotting and real time PCR. P-gp and MRP2 activities were estimated using model substrates rhodamine 123 and dinitrophenyl-S-glutathione (DNP-SG), respectively. CYP3A4 and GST activities were evaluated through their abilities to convert proluciferin into luciferin and 1-chloro-2,4-dinitrobenzene into DNP-SG, respectively. BZL (200 µM) increased the expression (protein and mRNA) of P-gp, MRP2, CYP3A4, and GSTπ class. A concomitant enhancement of activity was observed for all these proteins, except for CYP3A4, which exhibited a decreased activity. To elucidate if pregnane X receptor (PXR) mediates BZL response, its expression was knocked down with a specific siRNA. In this condition, the effect of BZL on P-gp, MRP2, CYP3A4, and GSTπ protein up-regulation was completely abolished. Consistent with this, BZL was able to activate PXR, as detected by reporter gene assay. Additional studies, using transporter inhibitors and P-gp-knock down cells, demonstrated that P-gp is involved in BZL extrusion. Pre-treatment of HepG2 cells with BZL increased its own efflux, as a consequence of P-gp up-regulation.Conclusions/SignificanceModifications in the activity of biotransformation and transport systems by BZL may alter the pharmacokinetics and efficiency of drugs that are substrates of these systems, including BZL itself.

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Effects of buthionine sulfoximine nifurtimox and benznidazole upon trypanothione and metallothionein proteins in Trypanosoma cruzi.

Cited by 19 publications

References 25 publications

Putative Role of the Aldo-Keto Reductase from Trypanosoma cruzi in Benznidazole Metabolism

Putative Role of the Aldo-Keto Reductase from Trypanosoma cruzi in Benznidazole Metabolism

Benznidazole Biotransformation and Multiple Targets in Trypanosoma cruzi Revealed by Metabolomics

Regulation of Biotransformation Systems and ABC Transporters by Benznidazole in HepG2 Cells: Involvement of Pregnane X-Receptor

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