Nitroheterocyclic drugs cure experimental <i>Trypanosoma cruzi</i> infections more effectively in the chronic stage than in the acute stage

The insect-transmitted protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease, and infects 5-8 million people in Latin America. Chagas disease is characterised by an acute phase, which is partially resolved by the immune system, but then develops as a chronic life-long infection. There is a consensus that the front-line drugs benznidazole and nifurtimox are more effective against the acute stage in both clinical and experimental settings. However, confirmative studies have been restricted by difficulties in demonstrating sterile parasitological cure. Here, we describe a systematic study of nitroheterocyclic drug efficacy using highly sensitive bioluminescence imaging of murine infections. Unexpectedly, we find both drugs are more effective at curing chronic infections, judged by treatment duration and therapeutic dose. This was not associated with factors that differentially influence plasma drug concentrations in the two disease stages. We also observed that fexinidazole and fexinidazole sulfone are more effective than benznidazole and nifurtimox as curative treatments, particularly for acute stage infections, most likely as a result of the higher and more prolonged exposure of the sulfone derivative. If these findings are translatable to human patients, they will have important implications for treatment strategies

Development of an ionic-liquid-based dispersive liquid–liquid microextraction method for the determination of antichagasic drugs in human breast milk. Optimization by central composite design

Chagas disease constitutes a major public health problem in Latin America. Human breast milk is a biological sample of great importance for the analysis of therapeutic drugs, as unwanted exposure through breast milk could result in pharmacological effects in the nursing infant. Thus, the goal of breast milk drug analysis is to inquire to which extent a neonate may be exposed to a drug during lactation. In this work, we developed an analytical technique to quantify benznidazole and nifurtimox (the two antichagasic drugs currently available for the medical treatment) in human breast milk, with a simple sample pre-treatment followed by an ionic-liquid-based dispersive liquid–liquid microextraction combined with high-performance liquid chromatography and UV detection. For this technique, the ionic liquid 1-octyl-3-methylimidazolium hexafluorophosphate has been used as the “extraction solvent”. A central composite design was used to find the optimum values for the significant variables affecting the extraction process: volume of ionic liquid, volume of dispersant solvent, ionic strength, and pH. At the optimum working conditions, the average recoveries were 77.5 and 89.7%, the limits of detection were 0.06 and 0.09 μg mL-1 and the inter-day reproducibilities were 6.25 and 5.77% for benznidazole and nifurtimox, respectively. The proposed methodology can be considered sensitive, simple, robust, accurate, and green

In Vitro and In Vivo High-Throughput Assays for the Testing of Anti-Trypanosoma cruzi Compounds

The treatment of Trypanosoma cruzi infection (the cause of human Chagas disease) remains a significant challenge. Only two drugs, both with substantial toxicity, are available and the efficacy of these dugs is often questioned – in many cases due to the limitations of the methods for assessing efficacy rather than to true lack of efficacy. For these reasons relatively few individuals infected with T. cruzi actually have their infections treated. In this study, we report on innovative methods that will facilitate the discovery of new compounds for the treatment of T. cruzi infection and Chagas disease. Utilizing fluorescent and bioluminescent parasite lines, we have developed in vitro tests that are reproducible and facile and can be scaled for high-throughput screening of large compound libraries. We also validate an in vivo screening test that monitors parasite replication at the site of infection and determines the effectiveness of drug treatment in less than two weeks. More importantly, results in this rapid in vivo test show strong correlations with those obtained in long-term (e.g. 40 day or more) treatment assays. The results of this study remove one of the obstacles for identification of effective and safe compounds to treat Chagas disease

Computational Identification of Uncharacterized Cruzain Binding Sites

Chagas disease, caused by the unicellular parasite Trypanosoma cruzi, claims 50,000 lives annually and is the leading cause of infectious myocarditis in the world. As current antichagastic therapies like nifurtimox and benznidazole are highly toxic, ineffective at parasite eradication, and subject to increasing resistance, novel therapeutics are urgently needed. Cruzain, the major cysteine protease of Trypanosoma cruzi, is one attractive drug target. In the current work, molecular dynamics simulations and a sequence alignment of a non-redundant, unbiased set of peptidase C1 family members are used to identify uncharacterized cruzain binding sites. The two sites identified may serve as targets for future pharmacological intervention

Chagas' disease: an update on immune mechanisms and therapeutic strategies

The final decade of the 20th century was marked by an alarming resurgence in infectious diseases caused by tropical parasites belonging to the kinetoplastid protozoan order. Among the pathogenic trypanosomatids, some species are of particular interest due to their medical importance. These species include the agent responsible for Chagas' disease, Trypanosoma cruzi. Approximately 8 to 10 million people are infected in the Americas, and approximately 40 million are at risk. in the present review, we discuss in detail the immune mechanisms elicited during infection by T. cruzi and the effects of chemotherapy in controlling parasite proliferation and on the host immune system.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Instituto Nacional de Biologia Estrutural e QuImica Medicinal em Doencas Infecciosas (INBEQMeDI)Univ São Paulo, Inst Biomed Sci, Dept Parasitol, São Paulo, BrazilMackenzie Presbeterian Univ, Ctr Biomol Sci & Hlth, São Paulo, BrazilNatl Univ Rosario, Sch Med Sci, Inst Immunol, Rosario, Santa Fe, ArgentinaCSIC, Inst Parasitol & Biomed Lopez Neyra, Granada, SpainUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, São Paulo, BrazilFAPESP: 08/57596-4FAPESP: 07/08648-9CNPq: 473906/2008-2Web of Scienc

The biological in vitro effect and selectivity of aromatic dicationic compounds on Trypanosoma cruzi

Trypanosoma cruzi is a parasite that causes Chagas disease, which affects millions of individuals in endemic areas of Latin America. One hundred years after the discovery of Chagas disease, it is still considered a neglected illness because the available drugs are unsatisfactory. Aromatic compounds represent an important class of DNA minor groove-binding ligands that exhibit potent antimicrobial activity. This study focused on the in vitro activity of 10 aromatic dicationic compounds against bloodstream trypomastigotes and intracellular forms of T. cruzi. Our data demonstrated that these compounds display trypanocidal effects against both forms of the parasite and that seven out of the 10 compounds presented higher anti-parasitic activity against intracellular parasites compared with the bloodstream forms. Additional assays to determine the potential toxicity to mammalian cells showed that the majority of the dicationic compounds did not considerably decrease cellular viability. Fluorescent microscopy analysis demonstrated that although all compounds were localised to a greater extent within the kinetoplast than the nucleus, no correlation could be found between compound activity and kDNA accumulation. The present results stimulate further investigations of this class of compounds for the rational design of new chemotherapeutic agents for Chagas disease

Skin lesions, malaise, and heart failure in a renal transplant recipient

A male Caucasian patient developed nodular erythematous skin lesions, malaise, and clinical signs of progressive heart failure 4 months after renal transplantation. Bronchoscopy with bronchoalveolar lavage performed for a small infiltrate seen on a computed tomography scan revealed Trypanosoma, which had at this point not been suspected as a cause. Parasitemia was present, and reactivation rather than transmission of Chagas' disease was established by performing polymerase chain reaction and serology in the donor and recipient. Treatment with benznidazole and allopurinol successfully reduced parasitemia, but the clinical course was fatal owing to progression of severe myocarditis. The patient had never lived in an endemic area, but had an extensive travel history in South America. The last visit was more than 5 years before transplantation. In non-endemic countries (United States, Europe), reactivation after transplantation has only been very rarely reported. Given the rising numbers of transplantations in patients with a migration background and extensive travel histories, specific screening procedures have to be considered