Synthesis and Anti-Trypanosoma cruzi Activity of 3-Cyanopyridine Derivatives.

Chagas disease (CD) is a parasitic neglected tropical disease (NTD) caused by the protozoan Trypanosoma cruzi that affects 6 million people worldwide, often resulting in financial burden, morbidity, and mortality in endemic regions. Given a lack of highly efficient and safe treatments, new, affordable, and fit-for-purpose drugs for CD are urgently needed. In this work, we present a hit-to-lead campaign for novel cyanopyridine analogues as antichagasic agents. In a phenotypic screening against intracellular T. cruzi, hits 1 and 2 were identified and displayed promising potency combined with balanced physicochemical properties. As part of the Lead Optimization Latin America consortium, a set of 40 compounds was designed, synthesized, and tested against T. cruzi intracellular amastigotes and relevant human cell lines. The structural modifications were focused on three positions: cyanopyridine core, linker, and right-hand side. The ADME properties of selected compounds, lipophilicity, kinetic solubility, permeability, and liver microsomal stability, were evaluated. Compounds 1-9 displayed good potency (EC50T. cruzi amastigote <1 µM), and most compounds did not present significant cytotoxicity (CC50 MRC-5 = 32-64 µM). Despite the good balance between potency and selectivity, the antiparasitic activity of the series appeared to be driven by lipophilicity, making the progression of the series unfeasible due to poor ADME properties and potential promiscuity issues.

Phytochemical characterization and antifungal potential of leaf extracts of Mosiera bullata.

The investigation of natural alternatives to conventional fungicides is of imminent need. Mosiera bullata (Britton & P. Wilson) Bisse is a Cuban endemic plant species belonging to the Myrtaceae family. The objective of the present study was to perform a bioassay-guided fractionation to explore the potential of extracts and fractions from M. bullata leaves against a panel of fungal plant pathogens. The M. bullata total extract was confirmed to have good antifungal activity against R. oryzae (IC50 = 4.86 µg/mL) and moderate activity against F. oxysporum (IC50 = 352.40 µg/mL) and F. solani (IC50 = 427.38 µg/mL) and fungicidal effect against R. oryzae. Five compounds belonging to the class of phloroglucinol dimers were tentatively characterized by UHPLC-HRMS and reported for the first time in M. bullata and the genus Mosiera. These results suggest the potential of M. bullata total extract as a natural antifungal product for the control of diseases in agriculture.

Hit-to-lead optimization of a 2-aminobenzimidazole series as new candidates for chagas disease.

Chagas disease is a neglected tropical disease caused by Trypanosoma cruzi. Because current treatments present several limitations, including long duration, variable efficacy and serious side effects, there is an urgent need to explore new antitrypanosomal drugs. The present study describes the hit-to-lead optimization of a 2-aminobenzimidazole hit 1 identified through in vitro phenotypic screening of a chemical library against intracellular Trypanosoma cruzi amastigotes, which focused on optimizing potency, selectivity, microsomal stability and lipophilicity. Multiparametric Structure-Activity Relationships were investigated using a set of 277 derivatives. Although the physicochemical and biological properties of the initial hits were improved, a combination of low kinetic solubility and in vitro cytotoxicity against mammalian cells prevented progression of the best compounds to an efficacy study using a mouse model of Chagas disease.

Semisynthetic triterpenes led to the generation of selective antitrypanosomal lead compounds.

Triterpenes α,ß-amyrin are naturally occurring molecules that can serve as building blocks for synthesizing new chemical entities. This study synthesized acyl, carboxyesther, NSAID, and nitrogenous derivatives and evaluated their antimicrobial activity. A cyclodextrin complexation method was developed to improve the solubility of the derivatives. Of the 17 derivatives tested, five exhibited activity against Trypanosoma cruzi, T. brucei, Leishmania infantum, Candida albicans, Staphylococcus aureus, and Escherichia coli. The 9a/9b mixture showed weak activity against the parasites (IC50 24.45-40.32 µM). However, it showed no activity for the other microorganisms. Derivatives 14a/14b exhibited potent activity against T. cruzi (IC50 2.0 nM) in this tested concentration did not show activity to the other microorganisms and were not cytotoxic. Derivatives 15a/15b and 16a/16b demonstrated relevant activity against the parasites (IC50 2.24-5.44 µM), but were also cytotoxic. Derivatives 17a/17b showed low activity against the tested parasites (IC50 21.70-22.79 µM), but they were selective since they did not show activity against other microorganisms. In docking studies, in general, all derivatives showed complementarity with the CYP51 binding site of the trypanosomatid mainly by hydrophobic interactions; thus, it is not conclusive that the molecules act by inhibiting this enzyme. Our results showed that triterpenes derivatives with antitrypanosomal activity could be synthesized by an inexpensive and rapid method.

Antifungal Activity of Extracts, Fractions, and Constituents from Coccoloba cowellii Leaves.

Coccoloba cowellii Britton (Polygonaceae, order Caryophyllales) is an endemic and critically endangered plant species that only grows in the municipality of Camagüey, a province of Cuba. A preliminary investigation of its total methanolic extract led to the discovery of promising antifungal activity. In this study, a bioassay-guided fractionation allowed the isolation of quercetin and four methoxyflavonoids: 3-O-methylquercetin, myricetin 3,3',4'-trimethyl ether, 6-methoxymyricetin 3,4'-dimethyl ether, and 6-methoxymyricetin 3,3',4'-trimethyl ether. The leaf extract, fractions, and compounds were tested against various fungi and showed strong in vitro antifungal activity against Cryptococcus neoformans and various Candida spp. with no cytotoxicity (CC50 > 64.0 µg/mL) on MRC-5 SV2 cells, determined by a resazurin assay. A Candida albicans SC5314 antibiofilm assay indicated that the antifungal activity of C. cowellii extracts and constituents is mainly targeted to planktonic cells. The total methanolic extract showed higher and broader activity compared with the fractions and mixture of compounds.

Development of Novel Isoindolone-Based Compounds against Trypanosoma brucei rhodesiense.

This study identified the isoindolone ring as a scaffold for novel agents against Trypanosoma brucei rhodesiense and explored the structure-activity relationships of various aromatic ring substitutions. The compounds were evaluated in an integrated in vitro screen. Eight compounds exhibited selective activity against T. b. rhodesiense (IC50 <2.2 µm) with no detectable side activity against T. cruzi and Leishmania infantum. Compound 20 showed low nanomolar potency against T. b. rhodesiense (IC50 =40 nm) and no toxicity against MRC-5 and PMM cell lines and may be regarded as a new lead template for agents against T. b. rhodesiense. The isoindolone-based compounds have the potential to progress into lead optimization in view of their highly selective in vitro potency, absence of cytotoxicity and acceptable metabolic stability. However, the solubility of the compounds represents a limiting factor that should be addressed to improve the physicochemical properties that are required to proceed further in the development of in vivo-active derivatives.

Chemical and Pharmacological Potential of Coccoloba cowellii, an Endemic Endangered Plant from Cuba.

Coccoloba cowellii Britton (Polygonaceae) is an endemic and critically endangered plant that only grows in Camagüey, a province of Cuba. In this study, a total of 13 compounds were identified in a methanolic leaf extract, employing a dereplication of the UHPLC-HRMS data by means of feature-based molecular networking (FBMN) analysis in the Global Natural Products Social Molecular Network (GNPS), together with the interpretation of the MS/MS data and comparison with the literature. The major constituents were glucuronides and glycosides of myricetin and quercetin, as well as epichatechin-3-O-gallate, catechin, epicatechin and gallic acid, all of them being reported for the first time in C. cowellii leaves. The leaf extract was also tested against various microorganisms, and it showed a strong antifungal effect against Candida albicans ATCC B59630 (azole-resistant) (IC50 2.1 µg/mL) and Cryptococcus neoformans ATCC B66663 (IC50 4.1 µg/mL) with no cytotoxicity (CC50 > 64.0 µg/mL) on MRC-5 SV2 cells, determined by the resazurin assay. Additionally, the extract strongly inhibited COX-1 and COX-2 enzyme activity using a cell-free experiment in a dose-dependent manner, being significantly more active on COX-1 (IC50 4.9 µg/mL) than on COX-2 (IC50 10.4 µg/mL). The constituents identified as well as the pharmacological activities measured highlight the potential of C. cowellii leaves, increasing the interest in the implementation of conservation strategies for this species.

2-aminobenzimidazoles for leishmaniasis: From initial hit discovery to in vivo profiling.

Leishmaniasis is a major infectious disease with hundreds of thousands of new cases and over 20,000 deaths each year. The current drugs to treat this life-threatening infection have several drawbacks such as toxicity and long treatment regimens. A library of 1.8 million compounds, from which the hits reported here are publicly available, was screened against Leishmania infantum as part of an optimization program; a compound was found with a 2-aminobenzimidazole functionality presenting moderate potency, low metabolic stability and high lipophilicity. Several rounds of synthesis were performed to incorporate chemical groups capable of reducing lipophilicity and clearance, leading to the identification of compounds that are active against different parasite strains and have improved in vitro properties. As a result of this optimization program, a group of compounds was further tested in anticipation of in vivo evaluation. In vivo tests were carried out with compounds 29 (L. infantum IC50: 4.1 µM) and 39 (L. infantum IC50: 0.5 µM) in an acute L. infantum VL mouse model, which showed problems of poor exposure and lack of efficacy, despite the good in vitro potency.

Synthesis and Structure-Activity Relationships of Imidazopyridine/Pyrimidine- and Furopyridine-Based Anti-infective Agents against Trypanosomiases.

Neglected tropical diseases remain among the most critical public health concerns in Africa and South America. The drug treatments for these diseases are limited, which invariably leads to fatal cases. Hence, there is an urgent need for new antitrypanosomal drugs. To address this issue, a large number of diverse heterocyclic compounds were prepared. Straightforward synthetic approaches tolerated pre-functionalized structures, giving rise to a structurally diverse set of analogs. We report on a set of 57 heterocyclic compounds with selective activity potential against kinetoplastid parasites. In general, 29 and 19 compounds of the total set could be defined as active against Trypanosoma cruzi and T. brucei brucei, respectively (antitrypanosomal activities <10 µM). The present work discusses the structure-activity relationships of new fused-ring scaffolds based on imidazopyridine/pyrimidine and furopyridine cores. This library of compounds shows significant potential for anti-trypanosomiases drug discovery.

Evaluation of phthalazinone phosphodiesterase inhibitors with improved activity and selectivity against Trypanosoma cruzi.

BACKGROUND: Chagas' disease, caused by the protozoan parasite Trypanosoma cruzi, needs urgent alternative therapeutic options as the treatments currently available display severe limitations, mainly related to efficacy and toxicity. OBJECTIVES: As phosphodiesterases (PDEs) have been claimed as novel targets against T. cruzi, our aim was to evaluate the biological aspects of 12 new phthalazinone PDE inhibitors against different T. cruzi strains and parasite forms relevant for human infection. METHODS: In vitro trypanocidal activity of the inhibitors was assessed alone and in combination with benznidazole. Their effects on parasite ultrastructural and cAMP levels were determined. PDE mRNA levels from the different T. cruzi forms were measured by quantitative reverse transcription PCR. RESULTS: Five TcrPDEs were found to be expressed in all parasite stages. Four compounds displayed strong effects against intracellular amastigotes. Against bloodstream trypomastigotes (BTs), three were at least as potent as benznidazole. In vitro combination therapy with one of the most active inhibitors on both parasite forms (NPD-040) plus benznidazole demonstrated a quite synergistic profile (xΣ FICI = 0.58) against intracellular amastigotes but no interaction (xΣ FICI = 1.27) when BTs were assayed. BTs treated with NPD-040 presented disrupted Golgi apparatus, a swollen flagellar pocket and signs of autophagy. cAMP measurements of untreated parasites showed that amastigotes have higher ability to efflux this second messenger than BTs. NPD-001 and NPD-040 increase the intracellular cAMP content in both BTs and amastigotes, which is also released into the extracellular milieu. CONCLUSIONS: The findings demonstrate the potential of PDE inhibitors as anti-T. cruzi drug candidates.

Structure-activity relationship of 4-azaindole-2-piperidine derivatives as agents against Trypanosoma cruzi.

The structure-activity relationship of a 4-Azaindole-2-piperidine compound selected from GlaxoSmithKline's recently disclosed open-resource "Chagas box" and possessing moderate activity against Trypanosoma cruzi, the parasite responsible for Chagas disease, is presented. Despite considerable medicinal chemistry efforts, a suitably potent and metabolically stable compound could not be identified to advance the series into in vivo studies. This research should be of interest to those in the area of neglected diseases and in particular anti-kinetoplastid drug discovery.