Antichagasic Agents Research Articles

ChemInform Abstract: Novel Quinoxaline 1,4‐Di‐N‐oxide Derivatives as New Potential Antichagasic Agents.

AbstractThe title products are obtained in very low yields.

Complexes of Trypanosoma cruzi Sterol 14α-Demethylase (CYP51) with Two Pyridine-based Drug Candidates for Chagas Disease: STRUCTURAL BASIS FOR PATHOGEN SELECTIVITY

Chagas disease, caused by the eukaryotic (protozoan) parasite Trypanosoma cruzi, is an alarming emerging global health problem with no clinical drugs available to treat the chronic stage. Azole inhibitors of sterol 14α-demethylase (CYP51) were proven effective against Chagas, and antifungal drugs posaconazole and ravuconazole have entered clinical trials in Spain, Bolivia, and Argentina. Here we present the x-ray structures of T. cruzi CYP51 in complexes with two alternative drug candidates, pyridine derivatives (S)-(4-chlorophenyl)-1-(4-(4-(trifluoromethyl)phenyl)-piperazin-1-yl)-2-(pyridin-3-yl)ethanone (UDO; Protein Data Bank code 3ZG2) and N-[4-(trifluoromethyl)phenyl]-N-[1-[5-(trifluoromethyl)-2-pyridyl]-4-piperi-dyl]pyridin-3-amine (UDD; Protein Data Bank code 3ZG3). These compounds have been developed by the Drugs for Neglected Diseases initiative (DNDi) and are highly promising antichagasic agents in both cellular and in vivo experiments. The binding parameters and inhibitory effects on sterol 14α-demethylase activity in reconstituted enzyme reactions confirmed UDO and UDD as potent and selective T. cruzi CYP51 inhibitors. Comparative analysis of the pyridine- and azole-bound CYP51 structures uncovered the features that make UDO and UDD T. cruzi CYP51-specific. The structures suggest that although a precise fit between the shape of the inhibitor molecules and T. cruzi CYP51 active site topology underlies their high inhibitory potency, a longer coordination bond between the catalytic heme iron and the pyridine nitrogen implies a weaker influence of pyridines on the iron reduction potential, which may be the basis for the observed selectivity of these compounds toward the target enzyme versus other cytochrome P450s, including human drug-metabolizing P450s. These findings may pave the way for the development of novel CYP51-targeted drugs with optimized metabolic properties that are very much needed for the treatment of human infections caused by eukaryotic microbial pathogens.

Novel 3-Nitro-1 H -1,2,4-Triazole-Based Compounds as Potential Anti-Chagasic Drugs: In Vivo Studies

Chagas disease is caused by the parasite Trypanosoma cruzi, is endemic in Latin America and leads to an estimated 14,000 deaths per year and around 100 million people at risk of infection. Drugs currently used in the treatment of Chagas are old, partially effective and have numerous side effects. We have previously reported that 3-nitro-1H-1,2,4-triazole-based compounds demonstrate significant and selective activity against T. cruzi amastigotes in infected L6 cells via activation of a type I nitroreductase, specific to trypanosomatids. In the present work we evaluated in vivo 13 of these compounds based on their high in vitro potency against T. cruzi (IC50 < 1 µM) and selectivity (SI: toxicity to L6 cells/toxicity against T. cruzi amastigotes > 200). Representative compounds of different chemical classes were included. A fast luminescence assay with transgenic parasites that express luciferase, and live imaging techniques were used. A total of 11 out of 13 compounds demonstrated significant antichagasic activity when administered intraperitoneally for 5-10 days at relatively small doses. The best in vivo activity was demonstrated by amides and sulfonamide derivatives. ADMET studies were performed for specific compounds. At least three compounds were identified as effective, non-toxic antichagasic agents suitable for further development.

Novel quinoxaline 1,4-di-N-oxide derivatives as new potential antichagasic agents

As a continuation of our research and with the aim of obtaining new agents against Chagas disease, an extremely neglected disease which threatens 100 million people, eighteen new quinoxaline 1,4-di-N-oxide derivatives have been synthesized following the Beirut reaction. The synthesis of the new derivatives was optimized through the use of a new and more efficient microwave-assisted organic synthetic method. The new derivatives showed excellent in vitro biological activity against Trypanosoma cruzi. Compound 17, which was substituted with fluoro groups at the 6- and 7-positions of the quinoxaline ring, was the most active and selective in the cytotoxicity assay. The electrochemical study showed that the most active compounds, which were substituted by electron-withdrawing groups, possessed a greater ease of reduction of the N-oxide groups.

Novel 3-Nitro-1H-1,2,4-triazole-Based Aliphatic and Aromatic Amines as Anti-Chagasic Agents

A series of novel 2-nitro-1H-imidazole- and 3-nitro-1H-1,2,4-triazole-based aromatic and aliphatic amines were screened for antitrypanosomal activity and mammalian cytotoxicity by the Drugs for Neglected Diseases initiative (DNDi). Out of 42 compounds tested, 18 3-nitro-1,2,4-triazoles and one 2-nitroimidazole displayed significant growth inhibitory properties against T. cruzi amastigotes (IC(50) ranging from 40 nM to 1.97 μM), without concomitant toxicity toward the host cells (L6 cells), having selectivity indices (SI) 44-1320. Most (16) of these active compounds were up to 33.8-fold more potent than the reference drug benznidazole, tested in parallel. Five novel 3-nitro-1,2,4-triazoles were active against bloodstream-form (BSF) T. b. rhodesiense trypomastigotes (IC(50) at nM levels and SI 220-993). An NADH-dependent nitroreductase (TbNTR) plays a role in the antiparasitic activity because BSF T. b. brucei trypomastigotes with elevated TbNTR levels were hypersensitive to tested compounds. Therefore, a novel class of affordable 3-nitro-1,2,4-triazole-based compounds with antitrypanosomal activity has been identified.

A Decade of Targets and Patented Drugs for Chemotherapy of Chagas Disease

Chagas disease, a parasitic infection typically spread by triatomine bugs, affects millions of people throughout Latin America. Current chemotherapy based on the nitroaromatic compounds, benzonidazole and nifurtimox provides unsatisfactory results and suffers from considerable side effects. Therefore, there is still an urgent need for new drugs to treat this neglected disease. During the last decade, the advances and understanding in the biology and biochemistry of Trypanosoma cruzi have allowed the identification of multiple new targets for Chagas' disease chemotherapy. Among the most promising targets for antiparasitic drugs are: cruzipain, the main cysteine protease of T. cruzi, essential for parasite survival and proliferation in mammalian host; ergosterol biosynthesis pathway; trypanothione synthesis and thiol-dependant redox metabolism. Specific enzymes of the glycolytic, pentose phosphate, polyisoprenoid (farnesylpyrophosphate synthase) and other particular biosynthetic pathways as well as enzymes from purine salvage (hypoxanthine-guanine phosphoribosyl-transferase, dihydrofolate reductase) have also been intensively studied in T. cruzi. In particular, trypanocidal agents that target the validated biochemical pathways of the parasite including cysteine proteinase inhibitors and inhibitors capable to block ergosterol biosynthesis are currently in the pipeline. Among the latter, posaconazole and ravuconazole, are planned to enter in clinical trials for trypanocidal chemotherapy in the near future. This review will summarize advances on antichagasic agents directed to specific parasite targets such as metabolic pathways or specific enzymes. Related patents filed and issued from 2000 to 2010 claiming inhibitors for specific parasite targets will be also discussed. Among them, the most represented were those related with cysteine proteinase inhibitors.

Synthesis, characterization and anti- Trypanosoma cruzi evaluation of ferrocenyl and cyrhetrenyl imines derived from 5-nitrofurane

A series of new cyrhetrenyl and ferrocenylimines complexes derived from 5-nitrofurane were designed, synthesized and characterized. The 1H and 13C NMR spectra indicate that these compounds adopt an anti-( E) conformation in solution, and confirmed for 1b by the X-ray crystal crystallography. The electronic effects of cyrhetrenyl ( 1b) and ferrocenyl ( 1a) bound directly to nitrogen have been correlated with the chemical shift of the iminic carbon. The trypanocidal activity (Tulahuen strain of Trypanosoma cruzi) of these compounds has been studied with respect to the substituent on the nitrogen atom of the 5-nitrofurfurylideneamino pharmacophore. Even though all the resulting derivatives were less active than Nifurtimox, the cyrhetrenyl complex ( 1b), compared with ferrocenyl ( 1a) or purely organic ( 4a and 4b) analogues, was more efficient as antichagasic agent. This result is likely due to the enhanced lipophilic character of the molecules or from a possible synergy between the cyrhetrenyl and 5-nitrofurane groups.

5-Nitro-2-furyl derivative actives against Trypanosoma cruzi: Preliminary in vivo studies

Ten 5-nitro-2-furyl derivatives, with good to excellent in vitro anti- Trypanosoma cruzi activity, and nifurtimox were tested oral and intraperitoneally on healthy animals for its acute toxicity on murine models. According to animals' survival percentage, organ histological results, biochemical and haematological findings, three new derivatives, with toxicity like nifurtimox, were selected to test in vivo as antichagasic agents. Clearly, dependences between chemical structure and both acute toxicity and in vivo anti- T. cruzi activity were observed. 4-Hexyl-1-[3-(5-nitro-2-furyl)-2-propenylidene]semicarbazide displayed good profile as anti- T. cruzi agent and better acute toxicity profile than nifurtimox.

Trypanosoma cruzi: Antiproliferative effect of indole phytoalexins on intracellular amastigotes in vitro

American trypanosomiasis (Chagas disease) continues to be a significant public health problem, and the therapeutic potential of current antichagasic agents (nifurtimox and benznidazole) is rather limited. Here we report on the antitrypanosomal effect of 1-methoxyspirobrassinol and other indole phytoalexins—secondary metabolites produced by Cruciferous plants. These compounds, that previously demonstrated antimicrobial and anticancer properties, displayed significant antiproliferative effects on intracellular amastigotes of Trypanosoma cruzi and may be prospective candidates for antichagasic drug design and development.

Antichagasic Activities of Natural Products against Trypanosoma cruzi

Trypanosoma cruzi (T. cruzi) is a parasitic protozoan transmitted to mammalian hosts by blood-sucking tri- atomine bugs. Infections by T. cruzi, known as Chagas' disease, pose a major public health problem in endemic countries in Central and South America. New chemotherapeutic agents are desired because of the lack of effective vaccines, undesirable side effects of anti-chagasic drugs in use such as nifurtimox and benznidazole, and the emer- gence of parasite resistance to these drugs. In the past two decades, novel advances and an improved understanding of the biology and biochemistry of T. cruzi have led to the identification of various targets for chemotherapy to treat Chagas' disease. In addition, many efforts have been undertaken to develop antichagasic agents, such as designed and synthesized compounds, natural products, and their derivatives, against a number of targets. Here, I mainly review recent studies on the antichagasic activities of natural products.

An Insight on Targets and Patented Drugs for Chemotherapy of Chagas Disease

Chagas disease or American Trypanosomiasis, a parasitic infection typically spread by triatomine bugs, affects millions of people throughout Latin America. Current chemotherapy based on the nitroaromatic compounds, benznidazole and nifurtimox provides unsatisfactory results and suffers from considerable side effects and low efficacy. Therefore, there is an urgent need for new drugs to treat this neglected disease. Over the last two decades, new advances and understanding in the biology and the biochemistry of Trypanosoma cruzi has allowed the identification of multiple targets for Chagas disease chemotherapy. This review summarizes antichagasic agents obtained based on i) target metabolic biochemical pathways or parasite specific enzymes, ii) natural products and its derivatives, iii) design and synthesis of lead compounds. Related patents filed and issued from 2000 to early 2006 are also discussed. Most of them claimed inhibitors on specific parasite targets such as cysteine proteinase, sterol biosynthesis, protein farnesyltransferase, etc. Particularly, those related to cysteine proteinase inhibitors were the most represented. Natural products also displayed many anti-T cruzi lead compounds. In addition, a few patents claiming natural or synthetic compounds with antichagasic activity, disclosed no specific target. However, only a small proportion of all these patents displayed specific data of biological trypanocidal activity.

Synthesis, in vitro evaluation, and SAR studies of a potential antichagasic 1 H-pyrazolo[3,4- b]pyridine series

The development of new drugs against Trypanosoma cruzi is still required since the only two drugs currently used cause severe side effects. In this work we described the synthesis, the in vitro biological evaluation, and the SAR results of 1 H-pyrazolo[3,4- b]pyridine derivatives, a new antichagasic agent series. The presence of fluorine, hydroxyl or nitro group at Y position resulted in at least one or two promising compounds in each set of derivatives ( 6f, 6g, 6i, 6l, and 6m). The SAR study showed that trypanocidal activity observed depends on both geometric and stereoelectronic parameters (MEP and frontier molecular orbitals HOMO and LUMO). We also used the Osiris program for calculating and comparing the fragment based druglikeness of the most active derivative ( 6g) (IC 50 = 1.9 μg/mL), the inactive compound ( 6o), and the current toxic antichagasic drugs (nifurtimox and benznidazole). Interestingly 6g presented a potential druglikeness higher than nifurtimox and benznidazole while 6o presented the lowest value among them.

Synthesis and in vitro evaluation of potential antichagasic hydroxymethylnitrofurazone (NFOH-121): a new nitrofurazone prodrug

The synthesis of mutual prodrugs of nitrofurazone with primaquine, using specific and nonspecific spacer groups, has been previously attempted seeking selective antichagasic agents. The intermediate reaction product, hydroxymethylnitrofurazone (NFOH-121), was isolated and tested in LLC-MK 2 culture cells infected with trypomastigotes forms of Trypanosoma cruzi showing higher trypanocidal activity than nitrofurazone and benznidazol in all stages. The mutagenicity tests showed that the prodrug was less toxic than the parent drug. Degradation assays were carried out in pH 1.2 and 7.4.

Design, synthesis, and biological evaluation of aryloxyethyl thiocyanate derivatives against Trypanosoma cruzi.

As a continuation of our project aimed at the search for new and safe chemotherapeutic and chemoprophylactic agents against American trypanosomiasis (Chagas' disease), several drugs structurally related to 4-phenoxyphenoxyethyl thiocyanate (4) were designed, synthesized, and evaluated as antiproliferative agents against the parasite responsible for this disease, the hemoflagellated protozoan Trypanosoma cruzi. This thiocyanate derivative was previously shown to be an effective and potent agent against T. cruzi proliferation. Several drugs possessing thiocyanate groups proved to be effective growth inhibitors of T. cruzi growth. Among the designed compounds, it is important to point out the extremely potent activity shown by 11, 23, 38, 53, 90, 99, and 117 against the epimastigote forms of the parasite. All of them exhibited IC(50) values in the low micromolar range, and these values were comparable with those presented by our lead drug 4 and ketokonazole, a well-known antiparasitic agent. The activity displayed by the nitrogen-containing derivative 90 was very promising with IC(50) values of 3.3 microM. Several other thiocyanate derivatives also proved to be very potent inhibitors of the multiplication of T. cruzi epimastigotes, such as compounds 28, 33, 43, 48, 56, 61, 66, 71, 76, and 124. Compound 43 resulted in being a promising drug because it was also very effective against amastigotes, the clinically more relevant form of the parasite. This compound was 3-fold more potent than 4, while 11 showed nearly the same activity as our lead drug against intracellular T. cruzi. It was very surprising that the experimental juvenoid 124, although fairly devoid of activity against epimastigotes, was very effective against intracellular amastigotes growing in myoblasts. The rest of the designed compounds showed a broad degree of inhibitory action, from moderately active drugs to drugs almost devoid of antiparasitic activity. Compound 43 is an interesting example of an effective antichagasic agent that presents excellent prospectives not only as a lead drug but also to be used for further in vivo studies.

Bisphosphonates Are Potent Inhibitors of Trypanosoma cruzi Farnesyl Pyrophosphate Synthase

We report the cloning and sequencing of a gene encoding the farnesyl pyrophosphate synthase of Trypanosoma cruzi. The protein (T. cruzi farnesyl pyrophosphate synthase, TcFPPS) is an attractive target for drug development, since the growth of T. cruzi is inhibited by carbocation transition state/reactive intermediate analogs of its substrates, the nitrogen-containing bisphosphonates currently in use in bone resorption therapy. The protein predicted from the nucleotide sequence of the gene has 362 amino acids and a molecular mass of 41.2 kDa. Several sequence motifs found in other FPPSs are present in TcFPPS. Heterologous expression of TcFPPS in Escherichia coli produced a functional enzyme that was inhibited by the nitrogen-containing bisphosphonates alendronate, pamidronate, homorisedronate, and risedronate but was less sensitive to the non-nitrogen-containing bisphosphonate etidronate, which, unlike the nitrogen-containing bisphosphonates, does not affect parasite growth. The protein contains a unique 11-mer insertion located near the active site, together with other sequence differences that may facilitate the development of novel anti-Chagasic agents.