Preclinical Studies in Anti-Trypanosomatidae Drug Development

The trypanosomatid parasites Trypanosoma brucei, Trypanosoma cruzi and Leishmania are the causative agents of human African trypanosomiasis, Chagas Disease and Leishmaniasis, respectively. These infections primarily affect poor, rural communities in the developing world, and are responsible for trapping sufferers and their families in a disease/poverty cycle. The development of new chemotherapies is a priority given that existing drug treatments are problematic. In our search for novel anti-trypanosomatid agents, we assess the growth-inhibitory properties of >450 compounds from in-house and/or “Pathogen Box” (PBox) libraries against L. infantum, L. amazonensis, L.braziliensis, T. cruzi and T. brucei and evaluate the toxicities of the most promising agents towards murine macrophages. Screens using the in-house series identified 17 structures with activity against and selective toward Leishmania: Compounds displayed 50% inhibitory concentrations between 0.09 and 25 μM and had selectivity index values >10. For the PBox library, ~20% of chemicals exhibited anti-parasitic properties including five structures whose activity against L. infantum had not been reported before. These five compounds displayed no toxicity towards murine macrophages over the range tested with three being active in an in vivo murine model of the cutaneous disease, with 100% survival of infected animals. Additionally, the oral combination of three of them in the in vivo Chagas disease murine model demonstrated full control of the parasitemia. Interestingly, phenotyping revealed that the reference strain responds differently to the five PBox-derived chemicals relative to parasites isolated from a dog. Together, our data identified one drug candidate that displays activity against Leishmania and other Trypanosomatidae in vitro and in vivo, while exhibiting low toxicity to cultured mammalian cells and low in vivo acute toxicity.

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In Vitro and In Vivo Trypanocidal Effects of the Cyclopalladated Compound 7a, a Drug Candidate for Treatment of Chagas' Disease

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00323-10 ◽

2010 ◽

Vol 54 (8) ◽

pp. 3318-3325 ◽

Cited By ~ 32

Author(s):

Alisson L. Matsuo ◽

Luis S. Silva ◽

Ana C. Torrecilhas ◽

Bruno S. Pascoalino ◽

Thiago C. Ramos ◽

...

Keyword(s):

Electron Microscopy ◽

Chagas Disease ◽

Cell Invasion ◽

Mammalian Cells ◽

New Drugs ◽

Drug Candidate ◽

Acute Form ◽

Intracellular Amastigote

ABSTRACT Chagas' disease, a neglected tropical infection, affects about 18 million people, and 100 million are at risk. The only drug available, benznidazole, is effective in the acute form and in the early chronic form, but its efficacy and tolerance are inversely related to the age of the patients. Side effects are frequent in elderly patients. The search for new drugs is thus warranted. In the present study we evaluated the in vitro and in vivo effect of a cyclopalladated compound (7a) against Trypanosoma cruzi, the agent of Chagas' disease. The 7a compound inhibits trypomastigote cell invasion, decreases intracellular amastigote proliferation, and is very effective as a trypanocidal drug in vivo, even at very low dosages. It was 340-fold more cytotoxic to parasites than to mammalian cells and was more effective than benznidazole in all in vitro and in vivo experiments. The 7a cyclopalladate complex exerts an apoptosis-like death in T. cruzi trypomastigote forms and causes mitochondrion disruption seen by electron microscopy.

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The Role of nitro (NO2-), chloro (Cl), and fluoro (F) Substitution in the Design of Antileishmanial and Antichagasic Compounds

Current Drug Targets ◽

10.2174/1389450121666201228122239 ◽

2020 ◽

Vol 21 ◽

Author(s):

Boniface Pone ◽

Ferreira Igne Elizabeth

Keyword(s):

Chagas Disease ◽

Mammalian Cells ◽

Neglected Tropical Diseases ◽

New Drugs ◽

Pharmacokinetic Studies ◽

Scientific Publications ◽

Antitrypanosomal Activity ◽

Chemical Groups

: Neglected tropical diseases (NTDs) are responsible for over 500,000 deaths annually and are characterized by multiple disabilities. Leishmaniasis and Chagas disease are among the most severe NTDs, and are caused by the Leishmania sp, and Trypanosoma cruzi, respectively. Glucantime, pentamidine and miltefosine are commonly used to treat leishmaniasis, whereas nifurtimox, benznidazole are current treatments for Chagas disease. However, these treatments are associated with drug resistance, and severe side effects. Hence, the development of synthetic products, especially those containing N02, F, or Cl, which chemical groups are known to improve the biological activity. The present work summarizes the information on the antileishmanial and antitrypanosomal activity of nitro-, chloro-, and fluoro-synthetic derivatives. Scientific publications referring to halogenated derivatives in relation to antileishmanial and antitrypanosomal activities were hand searched in databases such as SciFinder, Wiley, Science Direct, PubMed, ACS, Springer, Scielo, and so on. According to the literature information, more than 90 compounds were predicted as lead molecules with reference to their IC50/EC50 values in in vitro studies. It is worth to mention that only active compounds with known cytotoxic effects against mammalian cells were considered in the present study. The observed activity was attributed to the presence of nitro-, fluoro- and chloro-groups in the compound backbone. All in all, nitro and h0alogenated derivatives are active antileishmanial and antitrypanosomal compounds and can serve as baseline for the development of new drugs against leishmaniasis and Chagas disease. However, efforts on in vitro and in vivo toxicity studies of the active synthetic compounds is still needed. Pharmacokinetic studies, and the mechanism of action of the promising compounds need to be explored. The use of new catalysts and chemical transformation can afford unexplored halogenated compounds with improved antileishmanial and antitrypanosomal activity.

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Neurotrophin Receptor TrkC Is an Entry Receptor for Trypanosoma cruzi in Neural, Glial, and Epithelial Cells

Infection and Immunity ◽

10.1128/iai.05403-11 ◽

2011 ◽

Vol 79 (10) ◽

pp. 4081-4087 ◽

Cited By ~ 16

Author(s):

Craig Weinkauf ◽

Ryan Salvador ◽

Mercio PereiraPerrin

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Mammalian Cells ◽

Chinese Hamster ◽

Neuronal Cell ◽

Host Cells ◽

Neurotrophin Receptor ◽

Content Type

ABSTRACTTrypanosoma cruzi, the agent of Chagas' disease, infects a variety of mammalian cells in a process that includes multiple cycles of intracellular division and differentiation starting with host receptor recognition by a parasite ligand(s). Earlier work in our laboratory showed that the neurotrophin-3 (NT-3) receptor TrkC is activated byT. cruzisurfacetrans-sialidase, also known as parasite-derived neurotrophic factor (PDNF). However, it has remained unclear whether TrkC is used byT. cruzito enter host cells. Here, we show that a neuronal cell line (PC12-NNR5) relatively resistant toT. cruzibecame highly susceptible to infection when overexpressing human TrkC but not human TrkB. Furthermore,trkCtransfection conferred an ∼3.0-fold intracellular growth advantage. Sialylation-deficient Chinese hamster ovarian (CHO) epithelial cell lines Lec1 and Lec2 also became much more permissive toT. cruziafter transfection with thetrkCgene. Additionally, NT-3 specifically blockedT. cruziinfection of the TrkC-NNR5 transfectants and of naturally permissive TrkC-bearing Schwann cells and astrocytes, as did recombinant PDNF. Two specific inhibitors of Trk autophosphorylation (K252a and AG879) and inhibitors of Trk-induced MAPK/Erk (U0126) and Akt kinase (LY294002) signaling, but not an inhibitor of insulin-like growth factor 1 receptor, abrogated TrkC-mediated cell invasion. Antibody to TrkC blockedT. cruziinfection of the TrkC-NNR5 transfectants and of cells that naturally express TrkC. The TrkC antibody also significantly and specifically reduced cutaneous infection in a mouse model of acute Chagas' disease. TrkC is ubiquitously expressed in the peripheral and central nervous systems, and in nonneural cells infected byT. cruzi, including cardiac and gastrointestinal muscle cells. Thus, TrkC is implicated as a functional PDNF receptor in cell entry, independently of sialic acid recognition, mediating broadT. cruziinfection bothin vitroandin vivo.

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Enantiomers of Nifurtimox Do Not Exhibit Stereoselective Anti-Trypanosoma cruzi Activity, Toxicity, or Pharmacokinetic Properties

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05139-14 ◽

2015 ◽

Vol 59 (6) ◽

pp. 3645-3647 ◽

Cited By ~ 1

Author(s):

Carolina B. Moraes ◽

Karen L. White ◽

Stéphanie Braillard ◽

Catherine Perez ◽

Junghyun Goo ◽

...

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Murine Model ◽

Therapeutic Benefit ◽

Racemic Mixture ◽

Content Type ◽

Pharmacokinetic Properties

ABSTRACTWith the aim of improving the available drugs for the treatment of Chagas disease, individual enantiomers of nifurtimox were characterized. The results indicate that the enantiomers are equivalent in theirin vitroactivity against a panel ofTrypanosoma cruzistrains;in vivoefficacy in a murine model of Chagas disease;in vitrotoxicity and absorption, distribution, metabolism, and excretion characteristics; andin vivopharmacokinetic properties. There is unlikely to be any therapeutic benefit of an individual nifurtimox enantiomer over the racemic mixture.

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Organometallic gold(III) [Au(Hdamp)(L14)]Cl (L1 =SNS-donating thiosemicarbazone) complex protects mice against acuteT. cruziinfection

10.1101/312702 ◽

2018 ◽

Author(s):

Carla Duque Lopes ◽

Ana Paula S. Gaspari ◽

Ronaldo J. Oliveira ◽

Ulrich Abram ◽

José P. A. Almeida ◽

...

Keyword(s):

Chagas Disease ◽

Mammalian Cells ◽

Hydrophobic Interactions ◽

Covalent Bond ◽

Acute Stage ◽

Health Concern ◽

Gold Compound ◽

Docking Simulations

AbstractChagas disease remains a serious public health concern with unsatisfactory treatment outcomes due to strain-specific drug resistance and various side effects. To identify new therapeutic drugs againstTrypanosoma cruzi, we evaluated both thein vitroandin vivoactivity of the organometallic gold(III) complex [Au(Hdamp)(L14)]Cl (L1 =SNS- donating thiosemicarbazone), which was denoted 4-Cl. Our results demonstrated that 4- Cl was more effective than benznidazole (Bz) in eliminating both the extracellular trypomastigote and the intracellular amastigote forms of the parasite without cytotoxic effects on mammalian cells. In very-low-dosein vivoassays, 4-Cl reduced parasitaemia and tissue parasitism in addition to protecting the liver and heart from tissue damage. All these changes resulted in the survival of 100% of the mice treated with 4-Cl during the acute phase. We hypothesised that 4-Cl can act directly on the parasite and may participate in the modulation of IFN-γ production at the acute stage of the disease. Molecular docking simulations showed that the compound may interact with cruzain, a thiol protease considered a possible antiparasitic drug target, primarily by hydrophobic interactions. These analyses predicted that the Cys25 residue in the cruzain binding site is approximately 3.0 Å away from the S and Au atoms of the gold compound, which could suggest formation of a possible covalent bond between cruzain and the inhibitor. Overall, we confirmed the potential of 4-Cl as a new candidate for Chagas disease treatment.

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Antiproliferative Effects and Mechanism of Action of SCH 56592 against Trypanosoma (Schizotrypanum)cruzi: In Vitro and In Vivo Studies

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.42.7.1771 ◽

1998 ◽

Vol 42 (7) ◽

pp. 1771-1777 ◽

Cited By ~ 109

Author(s):

Julio A. Urbina ◽

Gilberto Payares ◽

Lellys M. Contreras ◽

Andreína Liendo ◽

Cristina Sanoja ◽

...

Keyword(s):

Chagas Disease ◽

Mechanism Of Action ◽

Murine Model ◽

Vero Cells ◽

In Vivo Studies ◽

Antiproliferative Effects ◽

Sterol Biosynthesis Inhibitor ◽

High Resolution Gas Chromatography

ABSTRACT We have investigated the antiproliferative effects of SCH 56592, a new experimental triazole, against Trypanosoma(Schizotrypanum) cruzi, the etiological agent of Chagas’ disease in Latin America. SCH 56592 blocked the proliferation of the epimastigote form of the parasite in vitro at 30 nM, a concentration 30- to 100-fold lower than that required with the reference compounds ketoconazole and itraconazole. At that concentration all the parasite’s endogenous sterols (ergosterol, 24-ethyl-cholesta-5,7,22-trien-3β-ol, and its 22-dihydro analogs), were replaced by methylated sterols (lanosterol and 24-methylene-dihydrolanosterol), as revealed by high-resolution gas chromatography coupled with mass spectrometry. This indicated that the primary mechanism of action of the drug was inhibition of the parasite’s sterol C-14α demethylase. Against the clinically relevant intracellular amastigote form, grown in cultured Vero cells at 37°C, the MIC of SCH 56592 was 0.3 nM, again 33- to 100-fold lower than that of ketoconazole or itraconazole. In a murine model of acute Chagas’ disease, SCH 56592 given at ≥ 10 mg/kg of body weight/day for a total of 43 doses allowed 85 to 100% survival and 90 to 100% cure of the surviving animals, as verified by parasitological, serological, and PCR-based tests, while ketoconazole given at 30 mg/kg day allowed 60% survival but only 20% cure. In a murine model of chronic Chagas’ disease, SCH 56592 was again more effective than ketoconazole, providing 75 to 85% protection from death, with 60 to 75% parasitological cures of the surviving animals, while no parasitological cures were observed with ketoconazole. The results indicate that SCH 56592 is the most powerful sterol biosynthesis inhibitor ever tested against T. cruzi and may be useful in the treatment of human Chagas’ disease.

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Requirement of norD for Brucella suis Virulence in a Murine Model of In Vitro and In Vivo Infection

Infection and Immunity ◽

10.1128/iai.74.3.1973-1976.2006 ◽

2006 ◽

Vol 74 (3) ◽

pp. 1973-1976 ◽

Cited By ~ 38

Author(s):

Séverine Loisel-Meyer ◽

Maria Pilar Jiménez de Bagüés ◽

Eugénie Bassères ◽

Jacques Dornand ◽

Stephan Köhler ◽

...

Keyword(s):

Nitric Oxide ◽

Murine Model ◽

Nitric Oxide Reductase ◽

Murine Macrophages ◽

Rapid Elimination ◽

Brucella Suis

ABSTRACT A mutant of Brucella suis bearing a Tn5 insertion in norD, the last gene of the operon norEFCBQD, encoding nitric oxide reductase, was unable to survive under anaerobic denitrifying conditions. The norD strain exhibited attenuated multiplication within nitric oxide-producing murine macrophages and rapid elimination in mice, hence demonstrating that norD is essential for Brucella virulence.

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Identification of a New Amide-Containing Thiazole as a Drug Candidate for Treatment of Chagas' Disease

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03814-14 ◽

2014 ◽

Vol 59 (3) ◽

pp. 1398-1404 ◽

Cited By ~ 22

Author(s):

Guzmán Álvarez ◽

Javier Varela ◽

Eugenia Cruces ◽

Marcelo Fernández ◽

Martín Gabay ◽

...

Keyword(s):

Chagas Disease ◽

Parasitic Infection ◽

Preclinical Study ◽

Drug Candidate ◽

Pharmacological Characterization ◽

Reference Drug ◽

Content Type ◽

Optimization Stage

ABSTRACTAlthough the parasitic infection Chagas' disease was described over 100 years ago, even now there are not suitable drugs. The available drugs nifurtimox and benznidazole have limited efficacies and tolerances, with proven mutagenic effects. Attempting to find appropriate drugs to deal with this problem, here we report on the development and pharmacological characterization of new amide-containing thiazoles. In the present study, we evaluated thein vitroandin vivoeffects of new candidates againstTrypanosoma cruzi, the etiological agent of Chagas' disease. The lead amide-containing thiazole derivative had potentin vitroactivity, an absence of bothin vitromutagenic andin vivoclastogenic effects, and excellentin vitroselectivity andin vivotolerance. The compound suppressed parasitemia in mice, modifying the anti-T. cruziantibodies like the reference drug, benznidazole, and displayed the lowest mortality among the tested drugs. The present evidence suggests that this compound is a promising anti-T. cruziagent surpassing the lead optimization stage in drug development and leading to a candidate for preclinical study.

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Concomitant Benznidazole and Suramin Chemotherapy in Mice Infected with a Virulent Strain of Trypanosoma cruzi

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00779-15 ◽

2015 ◽

Vol 59 (10) ◽

pp. 5999-6006 ◽

Cited By ~ 24

Author(s):

Eliziária C. Santos ◽

Rômulo D. Novaes ◽

Marli C. Cupertino ◽

Daniel S. S. Bastos ◽

Raphael C. Klein ◽

...

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Virulent Strain ◽

Harmful Effect ◽

Drug Candidate ◽

Concomitant Treatment ◽

Necrosis Factor Alpha ◽

Content Type

ABSTRACTAlthough suramin (Sur) is suggested as a potential drug candidate in the management of Chagas disease, this issue has not been objectively tested. In this study, we examined the applicability of concomitant treatment with benznidazole (Bz) and suramin in mice infected with a virulent strain ofTrypanosoma cruzi. Eighty 12-week-old male C57BL/6 mice were equally randomized in eight groups: (i) noninfected mice (negative control) and mice infected withT. cruziY strain receiving (ii) no treatment (positive control), (iii) Bz, 100 mg/kg of body weight per day, (iv) Sur, 20 mg/kg/day, and (v to viii) Sur, 20 mg/kg/day, combined with Bz, 100, 50, 25, or 5 mg/kg/day. Bz was administered by gavage, and Sur was administered intraperitoneally. Sur dramatically increased the parasitemia, cardiac content of parasite DNA, inflammation, oxidative tissue damage, and mortality. In response to high parasitic load in cardiac tissue, Sur stimulated the immune system in a manner typical of the acute phase of Chagas disease, increasing tissue levels of gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α) and inducing a preferential IgG2a anti-T. cruziserum pattern. When Sur and Bz were combined, the infection severity was attenuated, showing a dose-dependent Bz response. Sur therapy had a more harmful effect on the host than on the parasite and reduced the efficacy of Bz againstT. cruziinfection. Considering that Sur drastically reinforced the infection evolution, potentiating the inflammatory process and the severity of cardiac lesions, thein vivofindings contradicted thein vitroanti-T. cruzipotential described for this drug.

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