Insights to Design New Drugs against Human African Trypanosomiasis Targeting Rhodesain using Covalent Docking, Molecular Dynamics Simulations, and MM-PBSA Calculations.

BACKGROUND: Neglected tropical diseases (NTDs) are parasitic and bacterial diseases that affect approximately 149 countries, mainly the poor population without basic sanitation. Among these, African Human Trypanosomiasis (HAT), known as sleeping sickness, shows alarming data, with treatment based on suramin and pentamidine in the initial phase and melarsoprol and eflornithine in the chronic phase. Thus, to discover new drugs, several studies point to rhodesain as a promising drug target due to the function of protein degradation and intracellular transport of proteins between the insect and host cells and is present in all cycle phases of the parasite. METHODOLOGY: Here, based on the previous studies by Nascimento et al. (2021) that show the main rhodesain inhibitors development in the last decade, molecular docking and dynamics were applied in these inhibitors datasets to reveal crucial information that can be into drug design. Thus, conventional and covalent docking was employed and highlighted the presence of Michael acceptors in the ligands in a peptidomimetics scaffold, and interaction with Gly19, Gly23, Gly65, Asp161, and Trp184 is essential to the inhibiting activity. RESULTS: Also, our findings using MD simulations and MM-PBSA calculations confirmed Gly19, Gly23, Gly65, Asp161, and Trp184, showing high binding energy (ΔGbind between -72.782 to -124.477 kJ.mol-1). In addition, Van der Waals interactions have a better contribution (-140,930 to -96,988 kJ.mol-1) than electrostatic forces (-43,270 to -6,854 kJ.mol-1), indicating Van der Waals interactions are the leading forces in forming and maintaining ligand-rhodesain complexes. CONCLUSION: Furthermore, the Dynamic Cross-Correlation Maps (DCCM) show more correlated movements for all complexes than the free rhodesain and strong interactions in the regions of the aforementioned residues. Principal Component Analysis (PCA) demonstrates complex stability corroborating with RMSF and RMSD. This study can provide valuable insights that can guide researchers worldwide to discover a new promising drug against HAT.

Current Prospects of Saponins as Promising Anti-Trypanosoma brucei Compounds: Insight into the Mechanisms of Action.

BACKGROUND: Human African trypanosomiasis (HAT) is a parasitic infection that may lead to death if left untreated. This disease is caused by a protozoan parasite of the genus Trypanosoma and is transmitted to humans through tsetse fly bites. The disease is widespread across Sub-Saharan Africa, with 70% of cases in recent reports in the Democratic Republic of the Congo and an average of less than 1000 cases are declared annually. Since there is no appropriate treatment for HAT, steroidal and triterpenoid saponins have been reported to be effective in in vitro studies and might serve as scaffolds for the discovery of new treatments against this disease. AIM OF THE STUDY: The present study aimed to summarize up-to-date information on the anti-Trypanosoma brucei activity of steroidal and triterpenoid saponins. The mechanisms of action of in vitro bioactive compounds were also discussed. METHODS: Information on the anti-Trypanosoma brucei activity of plant saponins was obtained from published articles, dissertations, theses, and textbooks through a variety of libraries and electronic databases. RESULTS: There has been incredible progress in the identification of steroidal and triterpenoid saponins with pronounced in vitro activity against Trypanosoma brucei. Indeed, more than forty saponins were identified as having anti-T. brucei effect with activity ranging from moderate to highly active. The mechanisms of action of most of these saponins included DNA damage, cell cycle arrest, induction of apoptosis through downregulation of bcl-2 and MDM2, and upregulation of Bax and Bak, among others. CONCLUSION: Referring to in vitro studies, plant saponins have shown anti-Trypanosoma brucei activity; however, more cytotoxic and in vivo studies and detailed mechanisms of action of the bioactive saponins should be further considered.

Human African Trypanosomiasis (Sleeping Sickness)-Epidemiology, Clinical Manifestations, Diagnosis, Treatment, and Prevention.

Purpose of Review: Human African Trypanosomiasis (HAT), also known as sleeping sickness, is a vector-borne parasitic neglected tropical disease (NTD) endemic in sub-Saharan Africa. This review aims to enhance our understanding of HAT and provide valuable insights to combat this significant public health issue by synthesizing the latest research and evidence. Recent Findings: HAT has reached a historical < 1000 cases in 2018. In patients without neurologic symptoms and signs, the likelihood of a severe meningoencephalitic stage is deemed low, obviating the need for a lumbar puncture to guide treatment decisions using fexinidazole. Summary: Both forms of the disease, gambiense HAT (gHAT) and rhodesiense HAT (rHAT), have specific epidemiology, risk factors, diagnosis, and treatment. Disease management still requires a high index of suspicion, infectious disease expertise, and specialized medical care. Essential stakeholders in health policy are critical to accomplishing the elimination goals of the NTD roadmap for 2021-2030.

Hybrid-Compounds Against Trypanosomiases.

Neglected tropical diseases (NTDs) are a global public health problem associated with approximately 20 conditions. Among these, Chagas disease (CD), caused by Trypanosoma cruzi, and human African trypanosomiasis (HAT), caused by T. brucei gambiense or T. brucei rhodesiense, affect mainly the populations of the countries from the American continent and sub- Saharan Africa. Pharmacological therapies used for such illnesses are not yet fully effective. In this context, the search for new therapeutic alternatives against these diseases becomes necessary. A drug design tool, recently recognized for its effectiveness in obtaining ligands capable of modulating multiple targets for complex diseases, concerns molecular hybridization. Therefore, this review aims to demonstrate the importance of applying molecular hybridization in facing the challenges of developing prototypes as candidates for the treatment of parasitic diseases. Therefore, studies involving different chemical classes that investigated and used hybrid compounds in recent years were compiled in this work, such as thiazolidinones, naphthoquinones, quinolines, and others. Finally, this review covers several applications of the exploration of molecular hybridization as a potent strategy in the development of molecules potentially active against trypanosomiases, in order to provide information that can help in designing new drugs with trypanocidal activity.

Approaches to advance drug discovery for neglected tropical diseases.

Neglected tropical diseases (NTDs), which include leishmaniasis, Chagas disease, human African trypanosomiasis (HAT), and schistosomiasis, remain public health problems in developing countries, as highlighted in the 2021-2030 WHO Roadmap on NTDs. This agenda sets the challenges for the control and elimination of NTDs by 2030. Fortunately, NTD drug discovery has shifted from traditional to modern strategies combining medicinal chemistry, phenotypic and molecular assays, multiparameter optimization, structural biology, and omics approaches. Structure- and ligand-based drug design have fostered NTD drug discovery by enabling data-driven molecular optimization, expansion to previously inaccessible chemical spaces, and knowledge building from biological data. These efforts have integrated parasite biology and medicinal chemistry to advance drug discovery in this key area of global health.

A simple, robust, and affordable bioluminescent assay for drug discovery against infective African trypanosomes.

African trypanosomiasis is a major problem for human and animal health in endemic countries, where it threatens millions of people and affects economic development. New drugs are needed to overcome the toxicity, administration, low efficacy, and resistance issues of the current chemotherapy. Robust, simple, and economical high-throughput, whole-cell-based assays are required to accelerate the identification of novel chemical entities. With this aim, we generated a bioluminescent cell line of the bloodstream stage of Trypanosoma brucei brucei and established a screening assay. Trypanosomes were stably transfected to constitutively express a thermostable red-shifted luciferase. The growth phenotype and drug sensitivity of the reporter cell line were essentially identical to that of the parental cell line. The endogenous luciferase activity, measured by a simple bioluminescence assay, proved to be proportional to parasite number and metabolic status. The assay, optimized to detect highly potent compounds in a 96-well-plate format, was validated by screening a small compound library (inter-assay values for Z' factor and coefficient variation were 0.77 and 5.8%, respectively). With a hit-confirmation ratio of ~97%, the assay was potent enough to identify several hits with EC50 ≤ 10 µM. Preliminary tests indicated that the assay can be scaled up to a 384-well-plate format without compromising its robustness. In summary, we have generated reporter trypanosomes and a simple, robust, and affordable bioluminescence screening assay with great potential to speed up the early-phase drug discovery against African trypanosomes.

Cruzain and Rhodesain Inhibitors: Last Decade of Advances in Seeking for New Compounds Against American and African Trypanosomiases.

Neglected tropical diseases (NTDs) are a group of approximately 20 diseases that affect part of the population in Sub- and Tropical countries. In the past, pharmaceutical industries and governmental agencies have invested in the control, elimination and eradication of such diseases. Among these diseases, Chagas disease (CD) and Human African trypanosomiasis (HAT) are a public health problem, mainly in the countries from the American continent and sub-Saharan African. In this context, the search for new therapeutic alternatives against such diseases has been growing in recent years, presenting cysteine proteases as the main strategy to discover new anti-trypanosomal drugs. Thus, cruzain and rhodesain enzymes are targets widely studied, since the cruzain is present in all stages of the parasite's life, related to the stages of proliferation and differentiation and infection of macrophages; while the rhodesain is related to the immune defense process. In addition, knowledge about the amino acid sequences and availability of X-ray complexes have stimulated the drug searching against these targets, mainly through molecular modeling studies. Thus, this review manuscript will be addressed to cruzain and rhodesain inhibitors developed in the last 10 years, which could provide basis for new lead compounds in the discovery of new trypanocidal drugs. We found 117 studies involving inhibitors of cruzain and rhodesain, being thiosemicarbazones, semicarbazones, N-acylhydrazones, thiazoles-hydrazone, thiazolidinones-hydrazones, oxadiazoles, triazoles, triazines, imidazoles, peptidomimetic, and others. All references were obtained using "cruzain" or "rhodesain" and "inhibitor" as keywords in Science Direct, Bentham Science, PubMed, Espacenet, Springer, ACS Publisher, Wiley, Taylor and Francis, and MDPI (Multidisciplinary Digital Publishing Institute) databases. Finally, we highlighted all these chemical classes of molecules to provide valuable information that could be used to design new inhibitors against Chagas disease and sleeping sickness in the future.

VLP-Based Vaccines as a Suitable Technology to Target Trypanosomatid Diseases.

Research on vaccines against trypanosomatids, a family of protozoa that cause neglected tropical diseases, such as Chagas disease, leishmaniasis, and sleeping sickness, is a current need. Today, according to modern vaccinology, virus-like particle (VLP) technology is involved in many vaccines, including those undergoing studies related to COVID-19. The potential use of VLPs as vaccine adjuvants opens an opportunity for the use of protozoan antigens for the development of vaccines against diseases caused by Trypanosoma cruzi, Leishmania spp., and Trypanosoma brucei. In this context, it is important to consider the evasion mechanisms of these protozoa in the host and the antigens involved in the mechanisms of the parasite-host interaction. Thus, the immunostimulatory properties of VLPs can be part of an important strategy for the development and evaluation of new vaccines. This work aims to highlight the potential of VLPs as vaccine adjuvants for the development of immunity in complex diseases, specifically in the context of tropical diseases caused by trypanosomatids.

Metallodrugs for the Treatment of Trypanosomatid Diseases: Recent Advances and New Insights.

Trypanosomatid parasites are responsible for many Neglected Tropical Diseases (NTDs). NTDs are a group of illnesses that prevail in low-income populations, such as in tropical and subtropical areas of Africa, Asia, and the Americas. The three major human diseases caused by trypanosomatids are African trypanosomiasis, Chagas disease and leishmaniasis. There are known drugs for the treatment of these diseases that are used extensively and are affordable; however, the use of these medicines is limited by several drawbacks such as the development of chemo-resistance, side effects such as cardiotoxicity, low selectivity, and others. Therefore, there is a need to develop new chemotherapeutic against these tropical parasitic diseases. Metal-based drugs against NTDs have been discussed over the years as alternative ways to overcome the difficulties presented by approved antiparasitic agents. The study of late transition metal-based drugs as chemotherapeutics is an exciting research field in chemistry, biology, and medicine due to the ability to develop multitarget antiparasitic agents. The evaluation of the late transition metal complexes for the treatment of trypanosomatid diseases is provided here, as well as some insights about their mechanism of action.

Trypanosomatid-Caused Conditions: State of the Art of Therapeutics and Potential Applications of Lipid-Based Nanocarriers.

Trypanosomatid-caused conditions (African trypanosomiasis, Chagas disease, and leishmaniasis) are neglected tropical infectious diseases that mainly affect socioeconomically vulnerable populations. The available therapeutics display substantial limitations, among them limited efficacy, safety issues, drug resistance, and, in some cases, inconvenient routes of administration, which made the scenarios with insufficient health infrastructure settings inconvenient. Pharmaceutical nanocarriers may provide solutions to some of these obstacles, improving the efficacy-safety balance and tolerability to therapeutic interventions. Here, we overview the state of the art of therapeutics for trypanosomatid-caused diseases (including approved drugs and drugs undergoing clinical trials) and the literature on nanolipid pharmaceutical carriers encapsulating approved and non-approved drugs for these diseases. Numerous studies have focused on the obtention and preclinical assessment of lipid nanocarriers, particularly those addressing the two currently most challenging trypanosomatid-caused diseases, Chagas disease, and leishmaniasis. In general, in vitro and in vivo studies suggest that delivering the drugs using such type of nanocarriers could improve the efficacy-safety balance, diminishing cytotoxicity and organ toxicity, especially in leishmaniasis. This constitutes a very relevant outcome, as it opens the possibility to extended treatment regimens and improved compliance. Despite these advances, last-generation nanosystems, such as targeted nanocarriers and hybrid systems, have still not been extensively explored in the field of trypanosomatid-caused conditions and represent promising opportunities for future developments. The potential use of nanotechnology in extended, well-tolerated drug regimens is particularly interesting in the light of recent descriptions of quiescent/dormant stages of Leishmania and Trypanosoma cruzi, which have been linked to therapeutic failure.

Trypanocidal Essential Oils: A Review.

Trypanosomiases are diseases caused by parasitic protozoan trypanosomes of the genus Trypanosoma. In humans, this includes Chagas disease and African trypanosomiasis. There are few therapeutic options, and there is low efficacy to clinical treatment. Therefore, the search for new drugs for the trypanosomiasis is urgent. This review describes studies of the trypanocidal properties of essential oils, an important group of natural products widely found in several tropical countries. Seventy-seven plants were selected from literature for the trypanocidal activity of their essential oils. The main chemical constituents and mechanisms of action are also discussed. In vitro and in vivo experimental data show the therapeutic potential of these natural products for the treatment of infections caused by species of Trypanosoma.

Human African trypanosomiasis & heart.

INTRODUCTION: Human African Trypanosomiasis is a neglected tropical disease resulting from the infection with the parasite Trypanosoma brucei. Neurological compromise often dominates, and the impact of cardiovascular involvement has not been fully investigated. Recently, publications indicate that cardiovascular compromise is more frequent than previously thought. Early detection of cardiac complications may be of utmost importance for healthcare teams. AREA COVERED: As a part of the 'Neglected Tropical Diseases and other Infectious Diseases involving the Heart' (the NET-Heart Project), the purpose of this article is to review all the information available regarding cardiovascular implications of this disease, focusing on diagnosis and treatment, and proposing strategies for early detection of cardiac manifestations. An electronic systematic literature review of articles published in MEDLINE, PubMed and EMBASE was performed. From 50 initial studies, 18 were selected according to inclusion criteria. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement was used for conducting and reporting this review. EXPERT OPINION: Cardiovascular compromise through infiltrative and inflammatory mechanisms seems to be frequent, and includes a wide spectrum of severity. Conventional 12-lead electrocardiogram could be a useful test for screening cardiovascular manifestations and used as a guide for considering specific treatments or more sophisticated diagnostic tools.

The use of Sulfonamide Derivatives in the Treatment of Trypanosomatid Parasites including Trypanosoma cruzi, Trypanosoma brucei, and Leishmania ssp.

More than 10 million people around the world are afflicted by Neglected Tropical Diseases, such as Chagas Disease, Human African Trypanosomiasis, and Leishmania. These diseases mostly occur in undeveloped countries that suffer from a lack of economic incentive, research, and policy for new compound development. Sulfonamide moieties are effective scaffolds present in several compounds that are determinants to treat various diseases, principally neglected tropical diseases. This review article examines the contribution of these scaffolds in medicinal chemistry in the last five years, focusing on three trypanosomatid parasites: Trypanosoma cruzi, Trypanosoma brucei, and Leishmania ssp. We also present perspectives for their use in drug designs in an effort to contribute to new drug development. In addition, we consider the physicochemical parameters, whose molecules all presented according to Lipinski's rule. The correlation between the selective index and LogP was evaluated, showing that sulfonamide derivatives can act differently against each trypanosomatid parasite. Moreover, the approaches of novel drugs and technologies are very important for the eventual drug discovery against trypanosomatid diseases.

Variable Surface Glycoprotein from Trypanosoma brucei Undergoes Cleavage by Matrix Metalloproteinases: An in silico Approach.

In order to survive as extracellular parasites in the mammalian host environment, Trypanosoma brucei has developed efficient mechanisms of immune system evasion, which include the abundant expression of a variable surface glycoprotein (VSG) coat. VSGs are anchored in the parasite membrane by covalent C-terminal binding to glycosylphosphatidylinositol and may be periodically removed by a phospholipase C (PLC) and a major surface protein (TbMSP). VSG molecules show extraordinary antigenic diversity and a comparative analysis of protein sequences suggests that conserved elements may be a suitable target against African trypanosomiasis. However, the cleavage mechanisms of these molecules remain unclear. Moreover, in protozoan infections, including those caused by Trypanosoma brucei, it is possible to observe an increased expression of the matrix metalloproteinases (MMPs). To address the cleavage mechanism of VSGs, the PROSPER server was used for the identification of VSG sequence cleavage sites. After data compilation, it was observed that 64 VSG consensus sequences showed a high conservation of hydrophobic residues, such as valine (V), methionine (M), leucine (L) and isoleucine (I) in the fifth position-the exact location of the cleavage site. In addition, the PROSPER server identified conserved cleavage site portions of VSG proteins recognized by three matrix metalloproteases (gelatinases: MMP-2, MMP-3 and MMP-9). However, further biological studies are needed in order to analyze and confirm this prediction.

Natural Occurrence in Venomous Arthropods of Antimicrobial Peptides Active against Protozoan Parasites.

Arthropoda is a phylum of invertebrates that has undergone remarkable evolutionary radiation, with a wide range of venomous animals. Arthropod venom is a complex mixture of molecules and a source of new compounds, including antimicrobial peptides (AMPs). Most AMPs affect membrane integrity and produce lethal pores in microorganisms, including protozoan pathogens, whereas others act on internal targets or by modulation of the host immune system. Protozoan parasites cause some serious life-threatening diseases among millions of people worldwide, mostly affecting the poorest in developing tropical regions. Humans can be infected with protozoan parasites belonging to the genera Trypanosoma, Leishmania, Plasmodium, and Toxoplasma, responsible for Chagas disease, human African trypanosomiasis, leishmaniasis, malaria, and toxoplasmosis. There is not yet any cure or vaccine for these illnesses, and the current antiprotozoal chemotherapeutic compounds are inefficient and toxic and have been in clinical use for decades, which increases drug resistance. In this review, we will present an overview of AMPs, the diverse modes of action of AMPs on protozoan targets, and the prospection of novel AMPs isolated from venomous arthropods with the potential to become novel clinical agents to treat protozoan-borne diseases.

Application of Dithiocarbamates as Potential New Antitrypanosomatids-Drugs: Approach Chemistry, Functional and Biological.

Dithiocarbamates represent a class of compounds that were evaluated in different biomedical applications because of their chemical versatility. For this reason, several pharmacological activities have already been attributed to these compounds, such as antiparasitic, antiviral, antifungal activities, among others. Therefore, compounds that are based on dithiocarbamates have been evaluated in different in vivo and in vitro models as potential new antimicrobials. Thus, the purpose of this review is to present the possibilities of using dithiocarbamate compounds as potential new antitrypanosomatids-drugs, which could be used for the pharmacological control of Chagas disease, leishmaniasis, and African trypanosomiasis.

Flavonoid-derived Privileged Scaffolds in anti-Trypanosoma brucei Drug Discovery.

BACKGROUND: Human African Trypanosomiasis (HAT), also known as sleeping sickness is one of the 20 neglected tropical diseases listed by the World Health Organization, which lead to death if left untreated. This disease is caused by Trypanosoma brucei gambiense, which is the chronic form of the disease present in western and central Africa, and by T. brucei rhodesiense, which is the acute form of the disease located in eastern and southern Africa. Many reports have highlighted the effectiveness of flavonoid-based compounds against T. brucei. OBJECTIVE: The present review summarizes the current standings and perspectives for the use of flavonoids as lead compounds for the potential treatment of HAT. METHODS: A literature search was conducted for naturally occurring and synthetic anti-T brucei flavonoids by referencing textbooks and scientific databases (SciFinder, PubMed, Science Direct, Wiley, ACS, SciELO, Google Scholar, Springer, among others) from their inception until February 2019. RESULTS: Flavonoids isolated from different parts of plants and species were reported to exhibit moderate to high in vitro antitrypanosomal activity against T. brucei. In addition, synthetic flavonoids revealed anti-T. brucei activity. Molecular interactions of bioactive flavonoids with T. brucei protein targets showed promising results. CONCLUSION: According to in vitro anti-T brucei studies, there is evidence that flavonoids might be lead compounds for the potential treatment of HAT. However, toxicological studies, as well as the mechanism of action of the in vitro active flavonoids are needed to support their use as potential leads for the treatment of HAT.

Synthesis and structure-activity relationship studies of cruzain and rhodesain inhibitors.

Chagas disease and Human African trypanosomiasis (HAT) are important public health issues in Latin American and sub-Saharan African countries, respectively, and are responsible for a significant number of deaths. The drugs currently used to treat Chagas disease and HAT present efficacy, toxicity, and/or resistance issues; thus, there is a clear need for the discovery of novel targets and drug candidates to combat these diseases. In recent years, much effort has been made to find inhibitors of cruzain and rhodesain, which are promising targets for the design of novel trypanocidal compounds, since they are essential for parasite survival. Many reviews covering the design of novel cruzain and rhodesain inhibitors have been published; however, none have focused on the chemistry of the inhibitors. Thus, in the present work we reviewed the synthetic strategies and routes for the preparation of relevant classes of cruzain and rhodesain inhibitors. Perhaps the most important are the vinyl sulfone derivatives, and a very efficient synthetic strategy based on the Horner-Wadsworth-Emmons reaction was developed to yield these compounds. Modern approaches such as the asymmetric addition of substituted ethynyllithium to N-sulfinyl ketimines were used to produce the chiral alkynes that were employed in the preparation of important chiral triazole derivatives (potent cruzain inhibitors) and chiral HPLC resolution was used for the preparation of enantiopure 3-bromoisoxazoline derivatives (rhodesain inhibitors). Moreover, we also highlight the most important activity results and updated SAR results.

Structure-Based Virtual Screening and Biochemical Evaluation for the Identification of Novel Trypanosoma brucei Aldolase Inhibitors.

INTRODUCTION: The glycolytic enzyme fructose-1,6-bisphosphate aldolase is a validated molecular target in human African trypanosomiasis (HAT) drug discovery, a neglected tropical disease (NTD) caused by the protozoan Trypanosoma brucei. Herein, a structure-based virtual screening (SBVS) approach to the identification of novel T. brucei aldolase inhibitors is described. Distinct molecular docking algorithms were used to screen more than 500,000 compounds against the X-ray structure of the enzyme. This SBVS strategy led to the selection of a series of molecules which were evaluated for their activity on recombinant T. brucei aldolase. The effort led to the discovery of structurally new ligands able to inhibit the catalytic activity of the enzyme. RESULTS: The predicted binding conformations were additionally investigated in molecular dynamics simulations, which provided useful insights into the enzyme-inhibitor intermolecular interactions. CONCLUSION: The molecular modeling results along with the enzyme inhibition data generated practical knowledge to be explored in further structure-based drug design efforts in HAT drug discovery.

The first case report of trypanosomiasis caused by Trypanosoma evansi in Uruguay.

Trypanosoma evansi, the causal agent of Surra, is a salivarian African trypanosoma introduced in America during the 15th century. In this study we report the first case of African trypanosomiasis in Uruguay. We describe clinical signs and the evolution of the disease on an Uruguayan Cimarron dog, as well as the isolation and molecular characterization of the Trypanosoma evansi reported strain. This constitutes the first case of Surra in Uruguay, showing the expansion of the parasite to the south of the American continent. Future work is needed to identify the vector and possible reservoirs in order to avoid the spread of the disease. The epidemiological relevance of the findings are discussed.