Antitrypanosomal Agents Research Articles

Anti-trypanosomal Potential of Eucalyptus Camaldulensis

Chemotherapy of African trypanosomiasis in both the human and animal forms has been confronted with multidimensional problems that include paucity of drugs, resistance, high cost, prolonged treatment protocol and adverse side effects. The main objective of this study was to explore alternative sources of antitrypanosomal agents from the leaves, stem and root barks of <em>Eucalyptus camaldulensis</em>. The plant parts were sequentially extracted with hexane, ethyl acetate, methanol and water; and the extracts screened for antitrypanosomal activity. Mice infected with <em>Trypanosoma brucei brucei</em> were administered intraperitoneally doses ranging from 200-600 mg/kg body weight/day of the extracts for 21 consecutive days. One control group was treated with 3.5mg/kg bodyweight of berenil while the other control group was left untreated. The methanol extract of <em>E. camaldulensis</em> (leaf) produced complete cure for the animals in the different dose groups and survived as long as those treated with the standard drug, berenil, although the clearance time was faster for the standard drug. Sub inoculation of healthy mice with the blood and Cerebrospinal Fluid (CSF) of the cured mice did not result in infection, thus indicating a complete and permanent cure. Acute toxicity studies of the methanol extract of <em>E. camaldulensis</em> (leaf) confirmed the safety of the extract because no mortality was recorded even at 5000 mg/kg bodyweight. However, the extract had no prophylactic activity. Bioassay-guided fractionation of the crude methanol extract of <em>E. camaldulensis</em> leaf gave 10 fractions, with only fractions 8 and 9 exhibiting minimal antitrypanosomal activities that were not comparable to those of the crude extract and the standard drug (p≤0.05). Phytochemical screening revealed the presence of terpenes, steroids, saponins, tannins, alkaloids and fatty acids in both the crude extract and fraction 9; while fraction 8 contained only terpenes, steroids and fatty acids. Data from GC-MS analysis of the two fractions indicated likely components to be methyl esters, amides, long chain alkenes and alcohols. The result of this study shows that the methanol extract of <em>E. camaldulensis</em> (leaf) has immense potential for the development of drugs against African trypanosomiasis.

2D-QSAR study of 1,4-benzodiazepine-2-ones as potent anti-trypanosomal agents

2D-QSAR is one of the oldest methods of structure activity relationship analysis which provide many breakthroughs in the medicinal field. This method can provide a direct correlation between various physicochemical properties of a molecule with its structural characteristics. The correlations established by this method are based on the relative contribution of various descriptors. In the present study, this technique is utilized to develop a correlation equation between the anti-human African trypanosomiasis activity and the physicochemical parameters depicted by various descriptors utilized within the study. This series consists of 67 compounds with widespread range of activity between 0.78 and 3.13 μM. In addition, an effective correlation was obtained between SaasCE-index, Slogp, NitrogensCount, 0PathCount, Chi4pathCluster, CarbonsCount, and the biologic activity. These contributed descriptors illustrate that various physicochemical properties of the molecules responsible for the activities. The model suggests that the hydrophobicity has positive impact on the activity, while the presence of electronegative atom especially nitrogen and flexibility of the molecules also provide favorable effect on the activity.

Recombinant Arthrobacter β-1, 3-glucanase as a potential effector molecule for paratransgenic control of Chagas disease

BackgroundChagas disease is most often transmitted to humans by Trypanosoma cruzi infected triatomine bugs, and remains a significant cause of morbidity and mortality in Central and South America. Control of Chagas disease has relied mainly on vector eradication. However, development of insect resistance has prompted us to develop a paratransgenic strategy to control vectorial transmission of T. cruzi. Here, the potential role of recombinant endoglucanases as anti-trypanosomal agents for paratransgenic application is examined. The surface of T. cruzi is covered by a thick coat of mucin-like glycoproteins that have been proposed to play a role in the binding of T. cruzi to the membrane surface of the vector gut. We hypothesize that disruption of these glycoconjugates could arrest parasite development in the vector and abort the transmission cycle. In this work, we examine the effects of recombinant Arthrobacter luteus β-1, 3-glucanase expressed via Rhodococcus rhodnii on T. cruzi Sylvio II strain.Methods and resultsThe coding sequence for β-1, 3-glucanase was cloned in-frame to a heterologous promoter/signal sequence from the Mycobacterium kansasii alpha antigen gene resident in an E. coli/R. rhodnii shuttle vector. The resulting construct was confirmed by sequencing, and electroporated into R. rhodnii. Expression products from positive clones were purified from log phase cultures followed by dialysis into physiological buffers. Lysates and media were quantitated by ELISA against rabbit antibody specific to β-1,3-glucanase. Glucanase-positive samples were applied to live T. cruzi parasites in culture and viability accessed by spectrophotometric and fluorescent microscopic measurements. R. rhodnii-expressed β-1,3-glucanase exhibited toxicity against T. cruzi compared to controls when applied at 5 and 10% of the total culture volume. The decrease in cell viability ranged from a maximum of 50% for the media treatments to 80% for the filtered lysates.ConclusionsThese results suggest that recombinant β-glucanase could be a powerful addition to the arsenal of effector molecules for paratransgenic control of Chagas disease. In future studies, the ability of β-glucanase to function in combination with other effector molecules will be explored. Dual targeting of T. cruzi should not only slow resistance but also permit synergistic or additive lethal effects on T. cruzi.

α-Selective glycosylation affords mucin-related GalNAc amino acids and diketopiperazines active on Trypanosoma cruzi

This work addresses the synthesis and biological evaluation of glycosyl diketopiperazines (DKPs) cyclo[Asp-(αGalNAc)Ser] 3 and cyclo[Asp-(αGalNAc)Thr] 4 for the development of novel anti-trypanosomal agents and Trypanosoma cruzi trans-sialidase (TcTS) inhibitors. The target compounds were synthetized by coupling reactions between glycosyl amino acids αGalNAc-Ser 7 or αGalNAc-Thr 8 and the amino acid (O-tBu)-Asp 17, followed by one-pot deprotection-cyclisation reaction in the presence of 20% piperidine in DMF. The protected glycosyl amino acid intermediates 7 and 8 were, in turn, obtained by α-selective, HgBr2-catalysed glycosylation reactions of Fmoc-Ser/Thr benzyl esters 12/14 with αGalN3Cl 11, being, subsequently, fully deprotected for comparative biological assays. The DKPs 3 and 4 showed relevant anti-trypanosomal effects (IC50 282–124μM), whereas glycosyl amino acids 1 and 2 showed better TcTS inhibition (57–79%) than the corresponding DKPs (13–25%).

A rare form of imported infectious heart block

Background: Chagas disease is endemic in the southern cone of Latin America and is becoming more prevalent in the United States with more than 300,000 people infected. It is an important cause of heart block worldwide, but is thought to be rare in the United States, and therefore easily overlooked. Heart block from Chagas disease often occurs in the young, and is permanent; therefore, early diagnosis and treatment is crucial. Case report: A 37-year-old woman from Bolivia presented with decreased exercise capacity and generalized fatigue. Her electrocardiogram revealed right bundle branch block escape rhythm. Her enzyme-linked immunosorbent assay for Trypanosoma cruzi was negative, but given the high level of suspicion, an immunofluorescent antibody assay was performed, which was diagnostic. She was treated with benznidazole and permanent pacemaker placement. Conclusion: Chagas disease is becoming more prevalent in the United States and other regions of the developed world. Patients presenting from an endemic area with suggestive symptoms require investigation to detect this diagnosis because therapy restores patients to full functional capacity in the short term; in the long term, the prognosis is uncertain but cardiac surveillance for progressive ventricular dysfunction, thrombosis, and tachyarrhythmia is indicated. Treatment with anti-trypanosomal agents should be offered to patients with chronic Chagas disease and a permanent pacemaker should be considered in symptomatic patients with bradycardia.

Identification of the NAD(P)H binding site of eukaryotic UDP-galactopyranose mutase.

UDP-galactopyranose mutase (UGM) plays an essential role in galactofuranose biosynthesis in microorganisms by catalyzing the conversion of UDP-galactopyranose to UDP-galactofuranose. The enzyme has gained attention recently as a promising target for the design of new antifungal, antitrypanosomal, and antileishmanial agents. Here we report the first crystal structure of UGM complexed with its redox partner NAD(P)H. Kinetic protein crystallography was used to obtain structures of oxidized Aspergillus fumigatus UGM (AfUGM) complexed with NADPH and NADH, as well as reduced AfUGM after dissociation of NADP(+). NAD(P)H binds with the nicotinamide near the FAD isoalloxazine and the ADP moiety extending toward the mobile 200s active site flap. The nicotinamide riboside binding site overlaps that of the substrate galactopyranose moiety, and thus NADPH and substrate binding are mutually exclusive. On the other hand, the pockets for the adenine of NADPH and uracil of the substrate are distinct and separated by only 6 Å, which raises the possibility of designing novel inhibitors that bind both sites. All 12 residues that contact NADP(H) are conserved among eukaryotic UGMs. Residues that form the AMP pocket are absent in bacterial UGMs, which suggests that eukaryotic and bacterial UGMs have different NADP(H) binding sites. The structures address the longstanding question of how UGM binds NAD(P)H and provide new opportunities for drug discovery.

Synthesis and antitrypanosomal activity of new 6,6,7-trisubstituted thiopyrano[2,3-d][1,3]thiazoles

А series of novel 6,6,7-trisubstituted thiopyrano[2,3-d][1,3]thiazoles-based molecules have been synthesized and evaluated as potential antitrypanosomal agents. The most active analogue 3b inhibited Trypanosoma brucei brucei and Trypanosoma brucei gambiense with an IC50 of 0.26 and 0.42μМ, respectively. They could be considered as potent hits for further antitrypanosomal drug discovery efforts.

Antitrypanosomal and cysteine protease inhibitory activities of alkyldiamine cryptolepine derivatives

Cryptolepine derivatives containing alkyldiamine side-chains, 2, with potent inhibitory activity against Trypanosoma brucei brucei are reported. Compounds 2 showed improved activity and selectivity to T. b. brucei when compared to the lead compound. The most selective compound, 2k, presents a selectivity index value of 6200 and an IC50 of 10nM against the parasite. These derivatives are also potent inhibitors of the trypanosome papain-like cysteine proteases cruzain, which could, at least in part, explain their antitrypanosomal activity. Overall, these compounds with good antitrypanosomal activity and selectivity provide an encouraging starting point for the rational design of new and effective antitrypanosomal agents.

Identification of selective tubulin inhibitors as potential anti-trypanosomal agents

The potency of a series of sulfonamide tubulin inhibitors against the growth of Trypanosoma brucei (T. brucei), as well as human cancer and primary fibroblast cells were evaluated with the aim of determining whether compounds that selectively inhibit parasite proliferation could be identified. Several compounds showed excellent selectivity against T. brucei growth, and have the potential to be used for the treatment of Human African trypanosomiasis. A T. brucei tubulin protein homology model was built based on the crystal structure of the bovine tubulin. The colchicine-binding domain, which is also the binding site of the tested sulfonamide tubulin inhibitors, showed clear differences between the tubulin structures and presumably explained the selectivity of the compounds.

Theoretical models for the antitrypanosomal activity of thiosemicarbazone derivatives

AbstractThiosemicarbazones are cruzain inhibitors which have been identified as potential antitrypanosomal agents. In this work, several molecular properties were calculated at the density functional theory (DFT)/B3LYP/6‐311G* level for a set of 44 thiosemicarbazones. Unsupervised and supervised pattern recognition techniques (hierarchical cluster analysis, principal component analysis, kth‐nearest neighbors, and soft independent modeling by class analogy) were used to obtain structure–activity relationship models, which are able to classify unknown compounds according to their activities. The chemometric analyses performed here revealed that 12 descriptors can be considered responsible for the discrimination between high and low activity compounds. Classification models were validated with an external test set, showing that predictive classifications were achieved with the selected variable set. The results obtained here are in good agreement with previous findings from the literature, suggesting that our models can be useful on further investigations on the molecular determinants for the antichagasic activity. © 2012 Wiley Periodicals, Inc.

Design and synthesis of new (E)-cinnamic N-acylhydrazones as potent antitrypanosomal agents

We report herein the synthesis and trypanocidal profile of new (E)-cinnamic N-acylhydrazones (NAHs) designed by exploiting molecular hybridization between the potent cruzain inhibitors (E)-1-(benzo[d][1,3]dioxol-5-yl)-3-(4-bromophenyl)prop-2-en-1-one and (E)-3-hydroxy-N′-((2-hydroxynaphthalen-1-yl)methylene)-7-methoxy-2-naphthohydrazide. These derivatives were evaluated against both amastigote and trypomastigote forms of Trypanosoma cruzi and lead us to identify two compounds that were approximately two times more active than the reference drug, benznidazole, and with good cytotoxic index. Although designed as cruzain inhibitors, the weak potency displayed by the best cinnamyl NAH derivatives indicated that another mechanism of action was likely responsible for their trypanocide action.

Novel 3-Nitro-1H-1,2,4-triazole-Based Amides and Sulfonamides as Potential Antitrypanosomal Agents

A series of novel 3-nitro-1H-1,2,4-triazole-based (and in some cases 2-nitro-1H-imidazole-based) amides and sulfonamides were characterized for their in vitro antitrypanosomal and antileishmanial activities as well as mammalian toxicity. Out of 36 compounds tested, 29 (mostly 3-nitro-1H-1,2,4-triazoles) displayed significant activity against Trypanosoma cruzi intracellular amastigotes (IC(50) ranging from 28 nM to 3.72 μM) without concomitant toxicity to L6 host cells (selectivity 66-2782). Twenty-three of these active compounds were more potent (up to 58-fold) than the reference drug benznidazole, tested in parallel. In addition, nine nitrotriazoles which were moderately active (0.5 μM ≤ IC(50) < 6.0 μM) against Trypanosoma brucei rhodesiense trypomastigotes were 5-31-fold more active against bloodstream-form Trypanosoma brucei brucei trypomastigotes engineered to overexpress reduced nicotinamide adenine dinucleotide dependent nitroreductase. Finally, three nitrotriazoles displayed a moderate activity against the axenic form of Leishmania donovani . Therefore, 3-nitro-1H-1,2,4-triazole-based amides and sulfonamides are potent antitrypanosomal agents.

Identification In Silico and In Vitro of Novel Trypanosomicidal Drug‐Like Compounds

Atom-based bilinear indices and linear discriminant analysis are used to discover novel trypanosomicidal compounds. The obtained linear discriminant analysis-based quantitative structure-activity relationship models, using non-stochastic and stochastic indices, provide accuracies of 89.02% (85.11%) and 89.60% (88.30%) of the chemicals in the training (test) sets, respectively. Later, both models were applied to the virtual screening of 18 in-house synthesized compounds to find new pro-lead antitrypanosomal agents. The in vitro antitrypanosomal activity of this set against epimastigote forms of Trypanosoma cruzi is assayed. Predictions agree with experimental results to a great extent (16/18) of the chemicals. Sixteen compounds show more than 70% of epimastigote inhibition at a concentration 100 μg/mL. In addition, three compounds (CRIS 112, CRIS 140 and CRIS 147) present more than 70% of epimastigote inhibition at a concentration of 10 μg/mL (79.95%, 73.97% and 78.13%, respectively) with low values of cytotoxicity (19.7%, 7.44% and 20.63%, correspondingly).Taking into account all these results, we could say that these three compounds could be optimized in forthcoming works. Even though none of them resulted more active than nifurtimox, the current results constitute a step forward in the search for efficient ways to discover new lead antitrypanosomals.

Prescreening evaluation of some plant extracts used in ethno- veterinary practices as antitrypanosomal agents

Extracts from some plants were screened for bioactive properties based on ethnopharmacolgical information on their potential as antitrypanosomal agent using brine shrimp lethality assay. After 24 h of exposure, the results of the experiment, which examines the effect of plant type, concentration effect and the interaction of the two factors were significant (P 0.05) when their mean separations of percentage mortality to PM values obtained for Khaya senegalensis, Aristolochia albida, Butryrospermum paradoxium and Prosopis africana were compared. This preliminary result suggest that these plants with promising bioactivity may possess component that may provide the chemical lead towards the discovery of new generation trypanocides that are more potent and less toxic than the currently available and marketed trypanocidal drugs. Key words: Antitrypanosomal agent, brine shrimp, lethality bioassays, bioactivity, and medicinal plant extract.

Synthesis of precursors and macrocycle analogs of aerucyclamides as anti-trypanosomal agents

Macrocycle analogs and key fragments of aerucyclamides were obtained and evaluated against Trypanosoma brucei brucei.

New organoruthenium complexes with bioactive thiosemicarbazones as co-ligands: potential anti-trypanosomal agents.

In the search for new therapeutic tools against neglected diseases produced by trypanosomatid parasites, and particularly against African Trypanosomiasis, whose etiological agent is Trypanosoma brucei, organoruthenium compounds with bioactive nitrofuran containing thiosemicarbazones (L) as co-ligands were obtained. Four ruthenium(II) complexes with the formula [Ru(2)(p-cymene)(2)(L)(2)]X(2), where X = Cl or PF(6), were synthesized and the crystal structures of two of them were solved by X-ray diffraction methods. Two of the complexes show significant in vitro growth inhibition activity against Trypanosoma brucei brucei and are highly selective towards trypanosomal cells with respect to mammalian cells (J774 murine macrophages). These promising results make the title organoruthenium compounds good lead candidates for further developments towards potential antitrypanosomal organometallic drugs.

Manadoperoxides, a new class of potent antitrypanosomal agents of marine origin

Chemical investigation of the marine sponge Plakortis cfr. lita afforded a library of endoperoxyketal polyketides, manadoperoxides B-K (3-5 and 7-13) and peroxyplakoric esters B(3) (6) and C (14). Eight of these metabolites are new compounds and some contain an unprecedented chlorine-bearing THF-type ring in the side chain. The library of endoperoxide derivatives was evaluated for in vitro activity against Trypanosoma brucei rhodesiense and Leishmania donovani. Some compounds, such as manadoperoxide B, exhibited ultrapotent trypanocidal activity (IC(50) = 3 ng mL(-1)) without cytotoxicity. Detailed examination of the antitrypanosomal activity data and comparison with those available in the literature for related dioxane derivatives enabled us to draw a series of structure-activity relationships. Interestingly, it appears that minor structural changes, such as a shift of the methyl group around the dioxane ring, can dramatically affect the antitrypanosomal activity. This information can be valuable to guide the design of optimized antitrypanosomal agents based on the dioxane scaffold.

Antitrypanosomal Activity of Extract Fractions of Cetraria Islandica

The antitrypanosomal activity of various extract fractions of C. islandica (Iceland moss) was evaluated by using the well-established Alamar blue™ 96 well microplate assay. The antitrypanosomal activity significantly increased with increasing amounts of fractions (from 10-20 µg/ml) and MIC of 12.5µg/ml were observed for certain fractions. The results obtained in the present study indicate that C. islandica is a potential source of natural antitrypanosomal agents. © 2011 IGJPS. All rights reserved.

Design, synthesis and the effect of 1,2,3-triazole sialylmimetic neoglycoconjugates on Trypanosoma cruzi and its cell surface trans-sialidase

This work describes the synthesis of a series of sialylmimetic neoglycoconjugates represented by 1,4-disubstituted 1,2,3-triazole-sialic acid derivatives containing galactose modified at either C-1 or C-6 positions, glucose or gulose at C-3 position, and by the amino acid derivative 1,2,3-triazole fused threonine-3-O-galactose as potential TcTS inhibitors and anti-trypanosomal agents. This series was obtained by Cu(I)-catalysed azide–alkyne cycloaddition reaction (‘click chemistry’) between the azido-functionalized sugars 1-N3-Gal (commercial), 6-N3-Gal, 3-N3-Glc and 3-N3-Gul with the corresponding alkyne-based 2-propynyl-sialic acid, as well as by click chemistry reaction between the amino acid N3-ThrOBn with 3-O-propynyl-GalOMe. The 1,2,3-triazole linked sialic acid-6-O-galactose and the sialic acid-galactopyranoside showed high Trypanosoma cruzitrans-sialidase (TcTS) inhibitory activity at 1.0mM (approx. 90%), whilst only the former displayed relevant trypanocidal activity (IC50 260μM). These results highlight the 1,2,3-triazole linked sialic acid-6-O-galactose as a prototype for further design of new neoglycoconjugates against Chagas’ disease.

Trypanocidal Activity of Nitroaromatic Prodrugs: Current Treatments and Future Perspectives

Chagas disease and African sleeping sickness are trypanosomal infections that represent important public health problems in Latin America and Africa, respectively. The restriction of these diseases to the poorer parts of the world has meant that they have been largely neglected and limited progress has been made in their treatment. The nitroheterocyclic prodrugs nifurtimox and benznidazole, in use against Chagas disease for >40 years, remain the only agents available for this infection. In the case of African sleeping sickness, nifurtimox has recently been added to the arsenal of medicines, with the nitroheterocycle fexinidazole currently under evaluation. For a long time, the cytotoxic mechanism of these drugs was poorly understood: nifurtimox was thought to act via production of superoxide anions and nitro radicals, while the mode of benznidazole action was more obscure. The trypanocidal activity of nitroheterocyclic drugs is now known to depend on a parasite type I nitroreductase (NTR). This enzyme is absent from mammalian cells, a difference that forms the basis for the drug selectivity. The role of this enzyme in drug activation has been genetically and biochemically validated. It catalyses the 2-electron reduction of nitroheterocyclic compounds within the parasite, producing toxic metabolites without significant generation of superoxide. Recognition that this enzyme is responsible for activation of nitroheterocyclic prodrugs has allowed screening for compounds that preferentially target the parasite. This approach has led to the identification of two new classes of anti-trypanosomal agents, nitrobenzylphosphoramide mustards and aziridinyl nitrobenzamides, and promises to yield new, safer, more effective drugs.