naphthoquinones and benzonidazole

Chagas disease, which is caused by the protozoan Trypanosoma cruzi, is endemic to Latin America and mainly affects low-income populations. Chemotherapy is based on two nitrocompounds, but their reduced efficacy encourages the continuous search for alternative drugs. Our group has characterised the trypanocidal effect of naphthoquinones and their derivatives, with naphthoimidazoles derived from β-lapachone (N1, N2 and N3) being the most active in vitro.. In the present work, the effects of N1, N2 and N3 on acutely infected mice were investigated.. in vivo activity of the compounds was assessed by parasitological, biochemical, histopathological, immunophenotypical, electrocardiographic (ECG) and behavioral analyses.. Naphthoimidazoles led to a decrease in parasitaemia (8 dpi) by reducing the number of bloodstream trypomastigotes by 25-50% but not by reducing mortality. N1 protected mice from heart injury (15 dpi) by decreasing inflammation. Bradycardia was also partially reversed after treatment with N1 and N2. Furthermore, the three compounds did not reverse hepatic and renal lesions or promote the improvement of other evaluated parameters.. N1 showed moderate trypanocidal and promising immunomodulatory activities, and its use in combination with benznidazole and/or anti-arrhythmic drugs as well as the efficacy of its alternative formulations must be investigated in the near future.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Chagas Disease; Disease Models, Animal; Electrocardiography; Male; Mice; Naphthoquinones; Nitroimidazoles; Parasitemia; Time Factors; Trypanocidal Agents

The obligate intracellular protozoan Trypanosoma cruzi is the causative agent of Chagas disease, a neglected illness affecting millions of people in Latin America that recently entered non-endemic countries through immigration, as a consequence of globalization. The chemotherapy for this disease is based mainly on benznidazole and nifurtimox, which are very efficient nitroderivatives against the acute stage but present limited efficacy during the chronic phase. Our group has been studying the trypanocidal effects of naturally occurring quinones and their derivatives, and naphthoimidazoles derived from β-lapachone N1, N2 and N3 were the most active. To assess the molecular mechanisms of action of these compounds, we applied proteomic techniques to analyze treated bloodstream trypomastigotes, which are the clinically relevant stage of the parasite.. The approach consisted of quantification by 2D-DIGE followed by MALDI-TOF/TOF protein identification. A total of 61 differentially abundant protein spots were detected when comparing the control with each N1, N2 or N3 treatment, for 34 identified spots. Among the differentially abundant proteins were activated protein kinase C receptor, tubulin isoforms, asparagine synthetase, arginine kinase, elongation factor 2, enolase, guanine deaminase, heat shock proteins, hypothetical proteins, paraflagellar rod components, RAB GDP dissociation inhibitor, succinyl-CoA ligase, ATP synthase subunit B and methionine sulfoxide reductase.. Our results point to different modes of action for N1, N2 and N3, which indicate a great variety of metabolic pathways involved and allow for novel perspectives on the development of trypanocidal agents.

Topics: Animals; Chagas Disease; Electrophoresis, Gel, Two-Dimensional; Mice; Naphthoquinones; Nifurtimox; Nitroimidazoles; Proteomics; Protozoan Proteins; Trypanocidal Agents; Trypanosoma cruzi

Incubation of Trypanosoma cruzi culture (epimastigote) forms with nifurtimox (10 or 100 microM), benznidazole (38 or 380 microM) and beta-lapachone (1.6 or 7.8 microM) produced damage of nuclear DNA, as shown by the increased rate of the "unscheduled DNA synthesis" in epimastigotes arrested at phase S (9-, 3-, and 6-fold, respectively). alpha-lapachone, a position isomer of beta-lapachone, was completely ineffective. In order to demonstrate the "unscheduled repair of DNA", the semiconservative replication was inhibited by preincubating the epimastigotes for 16 hours with 10 mM hydroxyurea and 0.3 mM cycloheximide. Kinetoplast DNA (kDNA) extracted from epimastigotes pretreated with the trypanocidal agents revealed an increased number of single-strand breaks. After alkaline agarose-gel electrophoresis, a fast moving DNA fraction was detected in the kDNA from nifurtimox, benznidazole and beta-lapachone-treated parasites, while trapping of alkali-denatured kDNA by nitrocellulose filters, was significantly increased after treating the epimastigotes with the same drugs. Reincubation of these epimastigotes in fresh medium for 24 h, reestablished kDNA electrophoretic and filtration patterns to normality, except with 7.8 microM beta-lapachone, thus proving the reversibility of DNA lesions. Redox-cycling of nifurtimox and beta-lapachone in T. cruzi generates oxygen radicals, and accordingly, the higher effectiveness of these drugs (as compared with benznidazole and alpha-lapachone) supports the role of oxygen radicals for the trypanocidal action.

Topics: Animals; DNA; DNA Damage; DNA Repair; Free Radicals; Naphthoquinones; Nifurtimox; Nitroimidazoles; Oxidation-Reduction; Trypanocidal Agents; Trypanosoma cruzi

Nifurtimox and benznidazole have trypanostatic actions in vitro and inhibit the incorporation of [3H] thymidine, [3H] uridine and L-[3H] leucine in T. cruzi macromolecules. The effect of nifurtimox may be explained by (a) direct inhibition of nucleic acid biosynthesis, or (b) generation of the oxygen radicals in T. cruzi and therefore, only mechanism (a) should be valid. In order to obtain more information on the action of these drugs on T. cruzi, in the present study we examined the effect of nifurtimox and benznidazole on DNA, RNA and protein turnover in epimastigote (culture) forms of the parasite. Complementary experiments were performed with beta-lapachone that, like nifurtimox, generates oxygen radicals in T. cruzi. Epimastigotes (Tulahuen strain) at the exponential-phase of growth were cultured with [3H] thymidine, [3H] uridine or L-[3H] leucine to label DNA, RNA and protein, respectively. After incubation, the cells were washed free of radioactive precursor, resuspended in fresh medium and reincubated at 30 degrees C with nifurtimox (10 or 100 microM), benznidazole (38 or 380 microM) or beta-lapachone (1.6 or 7.8 microM), for 1-3 hours. Controls were incubated without drug. At one hour time intervals, sampler were taken, washed free of medium and filtered through 0.45 microns Metricel filters. The filters were washed with 10% trichloroacetic acid to remove the acid soluble material, and after drying, the radioactivity incorporated in DNA, RNA and protein was counted with a scintillation counter. The results show that after elimination of the labelled precursors, 3H activity in DNA, RNA and protein decayed as a function of the time of incubation. Nifurtimox, benznidazole and beta-lapachone, stimulated in all cases they decay of the incorporated radioactivity. Calculation of "half-life" values for DNA, RNA and protein(s) indicated that nifurtimox and beta-lapachone exerted their greatest effects on DNA while benznidazole increased the decay of DNA, RNA and protein to about the same extent. Taking into account the effects of nifurtimox and beta-lapachone on DNA stability, specific lesions (single-strand breaks) were investigated in DNA from control, nifurtimox, benznidazole or beta-lapachone treated epimastigotes. The number of single-strand breaks was (per 10(6)b) 25 with 100 microM nifurtimox, 1.4 with 380 microM benznidazole and 45 with 7.8 microM g-lapachone. Interestingly enough, after reincubation of nifurtimox-damaged epimastigotes in fresh me

Topics: Animals; Depression, Chemical; DNA; DNA Damage; Free Radicals; Naphthoquinones; Nifurtimox; Nitrofurans; Nitroimidazoles; Oxygen; Protein Biosynthesis; RNA; Trypanocidal Agents; Trypanosoma cruzi