Marine alkaloids as bioactive agents against protozoal neglected tropical diseases and malaria.

Covering: 2000 up to 2021Natural products are an important resource in drug discovery, directly or indirectly delivering numerous small molecules for potential development as human medicines. Among the many classes of natural products, alkaloids have a rich history of therapeutic applications. The extensive chemodiversity of alkaloids found in the marine environment has attracted considerable attention for such uses, while the scarcity of these natural materials has stimulated efforts towards their total synthesis. This review focuses on the biological activity of marine alkaloids (covering 2000 to up to 2021) towards Neglected Tropical Diseases (NTDs) caused by protozoan parasites, and malaria. Chemotherapy represents the only form of treatment for Chagas disease, human African trypanosomiasis, leishmaniasis and malaria, but there is currently a restricted arsenal of drugs, which often elicit severe adverse effects, show variable efficacy or resistance, or are costly. Natural product scaffolds have re-emerged as a focus of academic drug discovery programmes, offering a different resource to discover new chemical entities with new modes of action. In this review, the potential of a range of marine alkaloids is analyzed, accompanied by coverage of synthetic efforts that enable further studies of key antiprotozoal natural product scaffolds.

Activity of Chitosan and Its Derivatives against Leishmania major and Leishmania mexicana In Vitro.

There is an urgent need for safe, efficacious, affordable, and field-adapted drugs for the treatment of cutaneous leishmaniasis, which newly affects around 1.5 million people worldwide annually. Chitosan, a biodegradable cationic polysaccharide, has previously been reported to have antimicrobial, antileishmanial, and immunostimulatory activities. We investigated the in vitro activity of chitosan and several of its derivatives and showed that the pH of the culture medium plays a critical role in antileishmanial activity of chitosan against both extracellular promastigotes and intracellular amastigotes of Leishmania major and Leishmania mexicana Chitosan and its derivatives were approximately 7 to 20 times more active at pH 6.5 than at pH 7.5, with high-molecular-weight chitosan being the most potent. High-molecular-weight chitosan stimulated the production of nitric oxide and reactive oxygen species by uninfected and Leishmania-infected macrophages in a time- and dose-dependent manner at pH 6.5. Despite the in vitro activation of bone marrow macrophages by chitosan to produce nitric oxide and reactive oxygen species, we showed that the antileishmanial activity of chitosan was not mediated by these metabolites. Finally, we showed that rhodamine-labeled chitosan is taken up by pinocytosis and accumulates in the parasitophorous vacuole of Leishmania-infected macrophages.

Linking global drivers of agricultural trade to on-the-ground impacts on biodiversity.

Consumption of globally traded agricultural commodities like soy and palm oil is one of the primary causes of deforestation and biodiversity loss in some of the world's most species-rich ecosystems. However, the complexity of global supply chains has confounded efforts to reduce impacts. Companies and governments with sustainability commitments struggle to understand their own sourcing patterns, while the activities of more unscrupulous actors are conveniently masked by the opacity of global trade. We combine state-of-the-art material flow, economic trade, and biodiversity impact models to produce an innovative approach for understanding the impacts of trade on biodiversity loss and the roles of remote markets and actors. We do this for the production of soy in the Brazilian Cerrado, home to more than 5% of the world´s species. Distinct sourcing patterns of consumer countries and trading companies result in substantially different impacts on endemic species. Connections between individual buyers and specific hot spots explain the disproportionate impacts of some actors on endemic species and individual threatened species, such as the particular impact of European Union consumers on the recent habitat losses for the iconic giant anteater (Myrmecophaga tridactyla). In making these linkages explicit, our approach enables commodity buyers and investors to target their efforts much more closely to improve the sustainability of their supply chains in their sourcing regions while also transforming our ability to monitor the impact of such commitments over time.

Synthesis of a Microcystis aeruginosa predicted metabolite with antimalarial activity.

The synthesis of a Microcystis aeruginosa predicted metabolite analog of aerucyclamide B was performed. This hexacyclopeptide was obtained from three heterocyclic building blocks by a convergent macrocycle-assembly methodology. The compound exhibited good in vitro antiplasmodial activity (IC(50): 0.18 µM, K1, cholorquine resistant strain).

In vitro susceptibility of Trypanosoma cruzi strains from Santander, Colombia, to hexadecylphosphocholine (miltefosine), nifurtimox and benznidazole

Introduction. The current chemotherapy for Chagas disease is unsatisfactory with only two drugs available for treatment. Research to discover new drugs for Chagas disease is urgent. Hexadecyl-phosphocholine (HPC, miltefosine) has been demonstrated to have in vitro activity against Trypanosoma cruzi parasites, but its activity on different Colombian T. cruzi strains is not known. Objective. To evaluate the in vitro susceptibility of T. cruzi strains isolated from humans and vectors in Santander, Colombia, to miltefosine, nifurtimox and benznidazole. Materials and methods. Eight T. cruzi Colombian strains and three reference strains (Esmeraldo, SilvioX10 and Y) were studied. Drug activities against extracellular epimastigotes and intracellular amastigotes were determined by microscopic counting. The results were expressed as the concentrations that inhibited 50% and 90% growth (IC50 and IC90). Results. For miltefosine a similar range of drug activity was observed against all the Colombian strains, all parasites being more susceptible to miltefosine than to the reference drugs. The intracellular amastigotes were more susceptible to miltefosine (IC50 0.08 to 0.63 ìM and IC90 0.21 to 2.21 ìM) than extracellular forms (IC50 <0.92 to 2.29 ìM and IC90 1.38 to 4.76 ìM). For reference drugs, parasites were more susceptible to nifurtimox than to benznidazole and some differences in activity of benznidazole between T. cruzi strains was observed. Conclusions. The results showed the significant in vitro activity of miltefosine against T. cruzi stages, and the expected results for the reference drugs. Further in vivo studies with miltefosine are planned.

Introducción. Los tratamientos actuales para la enfermedad de Chagas son insatisfactorios y sólo existen dos medicamentos disponibles. La búsqueda de alternativas terapéuticas es prioritaria. La hexadecilfosfocolina (miltefosina) ha mostrado actividad in vitro contra Trypanosoma cruzi. Sin embargo, su actividad en aislamientos de T. cruzi obtenidos en Colombia aún no ha sido reportada. Objetivo. Evaluar la susceptibilidad in vitro a miltefosina, nifurtimox y benznidazole de cepas de T. cruzi aisladas de humanos y vectores en Santander, Colombia. Materiales y métodos. Se evaluó la susceptibilidad de los tres medicamentos en ocho cepas colombianas de T. cruzi y tres cepas de referencia: Esmeraldo, Silvio X10 y Y. La actividad de los compuestos fue determinada en epimastigotes extracelulares y amastigotes intracelulares, por conteo microscópico. Los resultados se expresaron en concentraciones inhibitorias 50 y 90 (CI50 y CI90). Resultados. Para la miltefosina, se observaron rangos similares en la actividad del medicamento entre las cepas colombianas; todos los parásitos fueron más susceptibles a la miltefosina que a los medicamentos de referencia. Los amastigotes intracelulares fueron más sensibles a la miltefosina (CI50, 0,08 a 0,63 µM y CI90, 0,21 a 2,21 µM) que las formas extracelulares (CI50, <0,92 a 2,29 µM y CI90, 1,38 a 4,76 µM). En los medicamentos de referencia, los parásitos fueron más susceptibles al nifurtimox que al benznidazole. Se observaron algunas diferencias en la actividad del benznidazole en las cepas estudiadas de T. cruzi. Conclusiones. Los resultados obtenidos de la actividad in vitro de miltefosina y de los medicamentos de referencia contra aislamientos de T. cruzi son satisfactorios y serán considerados en estudios posteriores in vivo.

Anti-African trypanocidal and antimalarial activity of natural flavonoids, dibenzoylmethanes and synthetic analogues.

OBJECTIVES: The known anti-protozoal activity of flavonoids has stimulated the testing of other derivatives from natural and synthetic sources. METHODS: As part of our efforts to find potential lead compounds, a number of flavonoids isolated from Neoraputia paraensis, N. magnifica, Murraya paniculata, (Rutaceae), Lonchocarpus montanus, L. latifolius, L. subglaucescens, L. atropurpureus, L. campestris, Deguelia hatschbachii (Leguminosae), dibenzoylmethanes from L. subglaucescens and synthetic analogues were tested for in-vitro activity against chloroquine-sensitive Plasmodium falciparum and Trypanosoma brucei rhodesiense bloodstream form trypomastigotes. An assay with KB cells has been developed in order to compare in-vitro cytotoxicity of flavonoids with a selective action on the parasites. KEY FINDINGS: Thirteen of the compounds tested had IC50 values ranging from 4.6 to 9.9 microm against T. brucei rhodesiense. In contrast, a small number of compounds showed significant activity against P. falciparum; seven of those tested had IC50 values ranging from 2.7 to 9.5 microm. Among the flavones only one had IC50 < 10 microm (7.6 microm), whereas against T. brucei rhodesiense seven had IC50 < 10 microm. Synthetic dibenzoylmethanes were the most active in terms of number (five) of compounds and the IC50 values (2.7-9.5 microm) against P. falciparum. CONCLUSIONS: Dibenzoylmethanes represent a novel class of compounds tested for the first time as antimalarial and trypanocidal agents.

In vitro susceptibility of Trypanosoma cruzi strains from Santander, Colombia, to hexadecylphosphocholine (miltefosine), nifurtimox and benznidazole.

INTRODUCTION: The current chemotherapy for Chagas disease is unsatisfactory with only two drugs available for treatment. Research to discover new drugs for Chagas disease is urgent. Hexadecyl-phosphocholine (HPC, miltefosine) has been demonstrated to have in vitro activity against Trypanosoma cruzi parasites, but its activity on different Colombian T. cruzi strains is not known. OBJECTIVE: To evaluate the in vitro susceptibility of T. cruzi strains isolated from humans and vectors in Santander, Colombia. to miltefosine, nifurtimox and benznidazole. MATERIALS AND METHODS: Eight T. cruzi Colombian strains and three reference strains (Esmeraldo, SilvioX10 and Y) were studied. Drug activities against extracellular epimastigotes and intracellular amastigotes were determined by microscopic counting. The results were expressed as the concentrations that inhibited 50% and 90% growth (IC50 and IC90). RESULTS: For miltefosine a similar range of drug activity was observed against all the Colombian strains, all parasites being more susceptible to miltefosine than to the reference drugs. The intracellular amastigotes were more susceptible to miltefosine (IC50 0.08 to 0.63 microM and IC90 0.21 to 2.21 microM) than extracellular forms (IC50 <0.92 to 2.29 microM and IC90 1.38 to 4.76 microM). For reference drugs, parasites were more susceptible to nifurtimox than to benznidazole and some differences in activity of benznidazole between T. cruzi strains was observed. CONCLUSIONS: The results showed the significant in vitro activity of miltefosine against T. cruzi stages, and the expected results for the reference drugs. Further in vivo studies with miltefosine are planned.

American tegumentary leishmaniasis: Is antimonial treatment outcome related to parasite drug susceptibility?

BACKGROUND: Antimonials are the first drug of choice for the treatment of American tegumentary leishmaniasis (ATL); however, their efficacy is not predictable, and this may be linked to parasite drug resistance. We aimed to characterize the in vitro antimony susceptibility of clinical isolates of Peruvian patients with ATL who were treated with sodium stibogluconate and to correlate this in vitro phenotype with different treatment outcomes. METHODS: Thirty-seven clinical isolates were obtained from patients with known disease and treatment histories. These isolates were typed, and the susceptibility of intracellular amastigotes to pentavalent (SbV) and trivalent (SbIII) antimonials was determined. RESULTS: We observed 29 SbV-resistant isolates among 4 species of subgenus Viannia, most of which exhibited primary resistance; isolates resistant only to SbIII; and 3 combinations of in vitro phenotypes: (1) parasites sensitive to both drugs, (2) parasites resistant to both drugs, and (3) parasites resistant to SbV only (the majority of isolates fell into this category). There was no correlation between in vitro susceptibility to both antimonials and the clinical outcome of therapy. CONCLUSION: Antimony insensitivity might occur in a stepwise fashion (first to SbV and then to SbIII). Our data question the definition of true parasite resistance to antimonials. Further studies of treatment efficacy should apply standardized protocols and definitions and should also consider host factors.

The sensitivity of clinical isolates of Leishmania from Peru and Nepal to miltefosine.

Clinical isolates of Leishmania, from visceral leishmaniasis (VL) cases in Nepal and from cutaneous leishmaniasis (CL) cases in Peru, were cultured using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to type species and strain. Promastigotes from 38 isolates, within eight passages from isolation, were used to infect mouse peritoneal macrophage cultures in vitro, and the amastigote sensitivity to miltefosine was determined. The concentration required to kill 50% of intracellular amastigotes from Nepalese VL isolates, all typed as Leishmania (L.) donovani (N = 24) from both Sbv responders and nonresponders, ranged from 8.7 to 0.04 microg/mL. In contrast, the concentration required to kill 50% intracellular amastigotes from isolates from Peru, typed as L.(V.) braziliensis (N = 8), was > 30 to 8.4 microg/mL, L.(V.) guyanensis (N = 2) > 30 to 1.9 microg/mL, L.(L.) mexicana (N = 1) > 30 microg/mL, and L. (V.) lainsoni (N = 4) was 3.4 to 1.9 microg/mL. This demonstrates a notable difference in the intrinsic sensitivity of Leishmania species to miltefosine in vitro. If this model can be correlated to therapeutic outcome, it may have implications for the interpretation of clinical trials.

Inhibitors of Leishmania mexicana CRK3 cyclin-dependent kinase: chemical library screen and antileishmanial activity.

The CRK3 cyclin-dependent kinase of Leishmania has been shown by genetic manipulation of the parasite to be essential for proliferation. We present data which demonstrate that chemical inhibition of CRK3 impairs the parasite's viability within macrophages, thus further validating CRK3 as a potential drug target. A microtiter plate-based histone H1 kinase assay was developed to screen CRK3 against a chemical library enriched for protein kinase inhibitors. Twenty-seven potent CRK3 inhibitors were discovered and screened against Leishmania donovani amastigotes in vitro. Sixteen of the CRK3 inhibitors displayed antileishmanial activity, with a 50% effective dose (ED50) of less than 10 microM. These compounds fell into four chemical classes: the 2,6,9-trisubstituted purines, including the C-2-alkynylated purines; the indirubins; the paullones; and derivatives of the nonspecific kinase inhibitor staurosporine. The paullones and staurosporine derivatives were toxic to macrophages. The 2,6,9-trisubstituted purines inhibited CRK3 in vitro, with 50% inhibitory concentrations ranging from high nanomolar to low micromolar concentrations. The most potent inhibitors of CRK3 (compounds 98/516 and 97/344) belonged to the indirubin class; the 50% inhibitory concentrations for these inhibitors were 16 and 47 nM, respectively, and the ED50s for these inhibitors were 5.8 and 7.6 microM, respectively. In culture, the indirubins caused growth arrest, a change in DNA content, and aberrant cell types, all consistent with the intracellular inhibition of a cyclin-dependent kinase and disruption of cell cycle control. Thus, use of chemical inhibitors supports genetic studies to confirm CRK3 as a validated drug target in Leishmania and provides pharmacophores for further drug development.

Cissampeloflavone, a chalcone-flavone dimer from Cissampelos pareira.

From the aerial parts of Cissampelos pareira L. (Menispermaceae), a chalcone-flavone dimer has been isolated which, mainly from NMR spectroscopic and MS data, was proved to be 2-(4-hydroxy-3-methoxyphenyl)-7-(4-methoxyphenyl)-6-(2-hydroxy-4,6-dimethoxybenzoyl)-furano[3,2-g]benzopyran-4-one. This has been assigned the trivial name cissampeloflavone. The compound has good activity against Trypanosoma cruzi and T. brucei rhodesiense and has a low toxicity to the human KB cell line.

Visceral leishmaniasis: current status of control, diagnosis, and treatment, and a proposed research and development agenda.

Visceral leishmaniasis is common in less developed countries, with an estimated 500000 new cases each year. Because of the diversity of epidemiological situations, no single diagnosis, treatment, or control will be suitable for all. Control measures through case finding, treatment, and vector control are seldom used, even where they could be useful. There is a place for a vaccine, and new imaginative approaches are needed. HIV co-infection is changing the epidemiology and presents problems for diagnosis and case management. Field diagnosis is difficult; simpler, less invasive tests are needed. Current treatments require long courses and parenteral administration, and most are expensive. Resistance is making the mainstay of treatment, agents based on pentavalent antimony, useless in northeastern India, where disease incidence is highest. Second-line drugs (pentamidine and amphotericin B) are limited by toxicity and availability, and newer formulations of amphotericin B are not affordable. The first effective oral drug, miltefosine, has been licensed in India, but the development of other drugs in clinical phases (paromomycin and sitamaquine) is slow. No novel compound is in the pipeline. Drug combinations must be developed to prevent drug resistance. Despite these urgent needs, research and development has been neglected, because a disease that mainly affects the poor ranks as a low priority in the private sector, and the public sector currently struggles to undertake the development of drugs and diagnostics in the absence of adequate funds and infrastructure. This article reviews the current situation and perspectives for diagnosis, treatment, and control of visceral leishmaniasis, and lists some priorities for research and development.