Discovery of new Schistosoma mansoni aspartyl protease inhibitors by structure-based virtual screening.

BACKGROUND: Schistosomiasis is a neglected tropical disease caused by trematodes of the genus Schistosoma, with a limited treatment, mainly based on the use of praziquantel (PZQ). Currently, several aspartic proteases genes have already been identified within the genome of Schistosoma species. At least one enzyme encoded from this gene family (SmAP), named SmCD1, has been validated for the development of schistosomicidal drugs, since it has a key role in haemoglobin digestion by worms. OBJECTIVE: In this work, we integrated a structure-based virtual screening campaign, enzymatic assays and adult worms ex vivo experiments aiming to discover the first classes of SmCD1 inhibitors. METHODS: Initially, the 3D-structures of SmCD1, SmCD2 and SmCD3 were generated using homology modelling approach. Using these models, we prioritised 50 compounds from 20,000 compounds from ChemBridge database for further testing in adult worm aqueous extract (AWAE) and recombinant SmCD1 using enzymatic assays. FINDINGS: Seven compounds were confirmed as hits and among them, two compounds representing new chemical scaffolds, named 5 and 19, had IC50 values against SmCD1 close to 100 µM while presenting binding efficiency indexes comparable to or even higher than pepstatin, a classical tight-binding peptide inhibitor of aspartyl proteases. Upon activity comparison against mammalian enzymes, compound 50 was selective and the most potent against the AWAE aspartic protease activity (IC50 = 77.7 µM). Combination of computational and experimental results indicate that compound 50 is a selective inhibitor of SmCD2. Compounds 5, 19 and 50 tested at low concentrations (10 uM) were neither cytotoxic against WSS-1 cells (48 h) nor could kill adult worms ex-vivo, although compounds 5 and 50 presented a slight decrease on female worms motility on late incubations times (48 or 72 h). MAIN CONCLUSION: Overall, the inhibitors identified in this work represent promising hits for further hit-to-lead optimisation.

Repurposing miconazole and tamoxifen for the treatment of Mycobacterium abscessus complex infections through in silico chemogenomics approach.

Drug repositioning is an alternative to overcome the complexity of the drug discovery and approval procedures for the treatment of Mycobacterium abscessus Complex (MABSC) infections that are increasing globally due to the emergency of antimicrobial resistance mechanisms. Here, an in silico chemogenomics approach was performed to compare the sequences from 4942 M. abscessus subsp. abscessus (M. abscessus) proteins with 5258 or 3473 therapeutic targets registered in the DrugBank or Therapeutic Target Database, respectively. This comparison identified 446 drugs or drug candidates whose targets were homologous to M. abscessus proteins. These identified drugs were considered potential inhibitors of MABSC (anti-MABSC activity). Further screening and inspection resulted in the selection of ezetimibe, furosemide, itraconazole, miconazole (MCZ), tamoxifen (TAM), and thiabendazole (THI) for experimental validation. Among them, MCZ and TAM showed minimum inhibitory concentrations (MIC) of 32 and 24 µg mL-1 against M. abscessus, respectively. For M. bolletii and M. massiliense strains, MCZ and TAM showed MICs of 16 and 24 µg mL-1, in this order. Subsequently, the antibacterial activity of MCZ was confirmed in vivo, indicating its potential to reduce the bacterial load in the lungs of infected mice. These results show that MCZ and TAM can serve as molecular scaffolds for the prospective hit-2-lead optimization of new analogs with greater potency, selectivity, and permeability.

Discovery of New Zika Protease and Polymerase Inhibitors through the Open Science Collaboration Project OpenZika.

The Zika virus (ZIKV) is a neurotropic arbovirus considered a global threat to public health. Although there have been several efforts in drug discovery projects for ZIKV in recent years, there are still no antiviral drugs approved to date. Here, we describe the results of a global collaborative crowdsourced open science project, the OpenZika project, from IBM's World Community Grid (WCG), which integrates different computational and experimental strategies for advancing a drug candidate for ZIKV. Initially, molecular docking protocols were developed to identify potential inhibitors of ZIKV NS5 RNA-dependent RNA polymerase (NS5 RdRp), NS3 protease (NS2B-NS3pro), and NS3 helicase (NS3hel). Then, a machine learning (ML) model was built to distinguish active vs inactive compounds for the cytoprotective effect against ZIKV infection. We performed three independent target-based virtual screening campaigns (NS5 RdRp, NS2B-NS3pro, and NS3hel), followed by predictions by the ML model and other filters, and prioritized a total of 61 compounds for further testing in enzymatic and phenotypic assays. This yielded five non-nucleoside compounds which showed inhibitory activity against ZIKV NS5 RdRp in enzymatic assays (IC50 range from 0.61 to 17 µM). Two compounds thermally destabilized NS3hel and showed binding affinity in the micromolar range (Kd range from 9 to 35 µM). Moreover, the compounds LabMol-301 inhibited both NS5 RdRp and NS2B-NS3pro (IC50 of 0.8 and 7.4 µM, respectively) and LabMol-212 thermally destabilized the ZIKV NS3hel (Kd of 35 µM). Both also protected cells from death induced by ZIKV infection in in vitro cell-based assays. However, while eight compounds (including LabMol-301 and LabMol-212) showed a cytoprotective effect and prevented ZIKV-induced cell death, agreeing with our ML model for prediction of this cytoprotective effect, no compound showed a direct antiviral effect against ZIKV. Thus, the new scaffolds discovered here are promising hits for future structural optimization and for advancing the discovery of further drug candidates for ZIKV. Furthermore, this work has demonstrated the importance of the integration of computational and experimental approaches, as well as the potential of large-scale collaborative networks to advance drug discovery projects for neglected diseases and emerging viruses, despite the lack of available direct antiviral activity and cytoprotective effect data, that reflects on the assertiveness of the computational predictions. The importance of these efforts rests with the need to be prepared for future viral epidemic and pandemic outbreaks.

DNA-Based Electrodes and Computational Approaches on the Intercalation Study of Antitumoral Drugs.

The binding between anticancer drugs and double-stranded DNA (dsDNA) is a key issue to understand their mechanism of action, and many chemical methods have been explored on this task. Molecular docking techniques successfully predict the affinity of small molecules into the DNA binding sites. In turn, various DNA-targeted drugs are electroactive; in this regard, their electrochemical behavior may change according to the nature and strength of interaction with DNA. A carbon paste electrode (CPE) modified with calf thymus ds-DNA (CPDE) and computational methods were used to evaluate the drug-DNA intercalation of doxorubicin (DOX), daunorubicin (DAU), idarubicin (IDA), dacarbazine (DAR), mitoxantrone (MIT), and methotrexate (MTX), aiming to evaluate eventual correlations. CPE and CPDE were immersed in pH 7 0.1 mM solutions of each drug with different incubation times. As expected, the CPDE response for all DNA-targeted drugs was higher than that of CPE, evidencing the drug-DNA interaction. A peak current increase of up to 10-fold was observed; the lowest increase was seen for MTX, and the highest increase for MIT. Although this increase in the sensitivity is certainly tied to preconcentration effects of DNA, the data did not agree entirely with docking studies, evidencing the participation of other factors, such as viscosity, interfacial electrostatic interactions, and coefficient of diffusion.

Computational Chemogenomics Drug Repositioning Strategy Enables the Discovery of Epirubicin as a New Repurposed Hit for Plasmodium falciparum and P. vivax.

Widespread resistance against antimalarial drugs thwarts current efforts for controlling the disease and urges the discovery of new effective treatments. Drug repositioning is increasingly becoming an attractive strategy since it can reduce costs, risks, and time-to-market. Herein, we have used this strategy to identify novel antimalarial hits. We used a comparative in silico chemogenomics approach to select Plasmodium falciparum and Plasmodium vivax proteins as potential drug targets and analyzed them using a computer-assisted drug repositioning pipeline to identify approved drugs with potential antimalarial activity. Among the seven drugs identified as promising antimalarial candidates, the anthracycline epirubicin was selected for further experimental validation. Epirubicin was shown to be potent in vitro against sensitive and multidrug-resistant P. falciparum strains and P. vivax field isolates in the nanomolar range, as well as being effective against an in vivo murine model of Plasmodium yoelii Transmission-blocking activity was observed for epirubicin in vitro and in vivo Finally, using yeast-based haploinsufficiency chemical genomic profiling, we aimed to get insights into the mechanism of action of epirubicin. Beyond the target predicted in silico (a DNA gyrase in the apicoplast), functional assays suggested a GlcNac-1-P-transferase (GPT) enzyme as a potential target. Docking calculations predicted the binding mode of epirubicin with DNA gyrase and GPT proteins. Epirubicin is originally an antitumoral agent and presents associated toxicity. However, its antiplasmodial activity against not only P. falciparum but also P. vivax in different stages of the parasite life cycle supports the use of this drug as a scaffold for hit-to-lead optimization in malaria drug discovery.

Nitrogen Catabolite Repression in members of Paracoccidioides complex.

Paracoccidioides complex is a genus that comprises pathogenic fungi which are responsible by systemic disease Paracoccidioidomycosis. In host tissues, pathogenic fungi need to acquire nutrients in order to survive, making the uptake of nitrogen essential for their establishment and dissemination. Nitrogen utilization is employed by the alleviation of Nitrogen Catabolite Repression (NCR) which ensures the use of non-preferential or alternative nitrogen sources when preferential sources are not available. NCR is controlled by GATA transcription factors which act through GATA binding sites on promoter regions in NCR-sensitive genes. This process is responsible for encoding proteins involved with the scavenge, uptake and catabolism of a wide variety of non-preferential nitrogen sources. In this work, we predict the existence of AreA GATA transcription factor and feature the zinc finger domain by three-dimensional structure in Paracoccidioides. Furthermore, we demonstrate the putative genes involved with NCR response by means of in silico analysis. The gene expression profile under NCR conditions was evaluated. Demonstrating that P. lutzii supported transcriptional regulation and alleviated NCR in non-preferential nitrogen-dependent medium. The elucidation of NCR in members of Paracoccidioides complex will provide new knowledge about survival, dissemination and virulence for these pathogens with regard to nitrogen-scavenging strategies in the interactions of host-pathogens.

Flavonoids from Brazilian Cerrado: Biosynthesis, Chemical and Biological Profile.

Flavonoids are highly bioactive compounds with very low toxicity, which makes them attractive starting points in drug discovery. This study aims to provide information on plant species containing flavonoids, which are found in the Brazilian Cerrado. First, we present the characterization and plant diversity with emphasis on the families of flavonoid-producing plants, and then we describe the phenylpropanoid pathway which represents the flavonoids' main route biosynthesis-generally conserved in all species. Chemical structures and biological activities of flavonoids isolated from the Cerrado's plant species are also described based on examples from the relevant literature studies. Finally, research on the biodiversity of the Cerrado biome should be encouraged, due to the discovery of new sources of flavonoids which can provide several benefits to human health and the possibility of developing new drugs by the pharmaceutical industry.

Synthesis and molecular modelling studies of pyrimidinones and pyrrolo[3,4-d]-pyrimidinodiones as new antiplasmodial compounds.

BACKGROUND Malaria is responsible for 429,000 deaths per year worldwide, and more than 200 million cases were reported in 2015. Increasing parasite resistance has imposed restrictions to the currently available antimalarial drugs. Thus, the search for new, effective and safe antimalarial drugs is crucial. Heterocyclic compounds, such as dihydropyrimidinones (DHPM), synthesised via the Biginelli multicomponent reaction, as well as bicyclic compounds synthesised from DHPMs, have emerged as potential antimalarial candidates in the last few years. METHODS Thirty compounds were synthesised employing the Biginelli multicomponent reaction and subsequent one-pot substitution/cyclisation protocol; the compounds were then evaluated in vitro against chloroquine-resistant Plasmodium falciparum parasites (W2 strain). Drug cytotoxicity in baseline kidney African Green Monkey cells (BGM) was also evaluated. The most active in vitro compounds were evaluated against P. berghei parasites in mice. Additionally, we performed an in silico target fishing approach with the most active compounds, aiming to shed some light into the mechanism at a molecular level. RESULTS The synthetic route chosen was effective, leading to products with high purity and yields ranging from 10-84%. Three out of the 30 compounds tested were identified as active against the parasite and presented low toxicity. The in silico study suggested that among all the molecular targets identified by our target fishing approach, Protein Kinase 3 (PK5) and Glycogen Synthase Kinase 3ß (GSK-3ß) are the most likely molecular targets for the synthesised compounds. CONCLUSIONS We were able to easily obtain a collection of heterocyclic compounds with in vitro anti-P. falciparum activity that can be used as scaffolds for the design and development of new antiplasmodial drugs.

Whole genome sequencing identifies novel mutations in malaria parasites resistant to artesunate (ATN) and to ATN + mefloquine combination.

Introduction: The global evolution of resistance to Artemisinin-based Combination Therapies (ACTs) by malaria parasites, will severely undermine our ability to control this devastating disease. Methods: Here, we have used whole genome sequencing to characterize the genetic variation in the experimentally evolved Plasmodium chabaudi parasite clone AS-ATNMF1, which is resistant to artesunate + mefloquine. Results and discussion: Five novel single nucleotide polymorphisms (SNPs) were identified, one of which was a previously undescribed E738K mutation in a 26S proteasome subunit that was selected for under artesunate pressure (in AS-ATN) and retained in AS-ATNMF1. The wild type and mutated three-dimensional (3D) structure models and molecular dynamics simulations of the P. falciparum 26S proteasome subunit Rpn2 suggested that the E738K mutation could change the toroidal proteasome/cyclosome domain organization and change the recognition of ubiquitinated proteins. The mutation in the 26S proteasome subunit may therefore contribute to altering oxidation-dependent ubiquitination of the MDR-1 and/or K13 proteins and/or other targets, resulting in changes in protein turnover. In light of the alarming increase in resistance to artemisin derivatives and ACT partner drugs in natural parasite populations, our results shed new light on the biology of resistance and provide information on novel molecular markers of resistance that may be tested (and potentially validated) in the field.

Drug to genome to drug: a computational large-scale chemogenomics screening for novel drug candidates against sporotrichosis.

Sporotrichosis is recognized as the predominant subcutaneous mycosis in South America, attributed to pathogenic species within the Sporothrix genus. Notably, in Brazil, Sporothrix brasiliensis emerges as the principal species, exhibiting significant sapronotic, zoonotic and enzootic epidemic potential. Consequently, the discovery of novel therapeutic agents for the treatment of sporotrichosis is imperative. The present study is dedicated to the repositioning of pharmaceuticals for sporotrichosis therapy. To achieve this goal, we designed a pipeline with the following steps: (a) compilation and preparation of Sporothrix genome data; (b) identification of orthologous proteins among the species; (c) identification of homologous proteins in publicly available drug-target databases; (d) selection of Sporothrix essential targets using validated genes from Saccharomyces cerevisiae; (e) molecular modeling studies; and (f) experimental validation of selected candidates. Based on this approach, we were able to prioritize eight drugs for in vitro experimental validation. Among the evaluated compounds, everolimus and bifonazole demonstrated minimum inhibitory concentration (MIC) values of 0.5 µg/mL and 4.0 µg/mL, respectively. Subsequently, molecular docking studies suggest that bifonazole and everolimus may target specific proteins within S. brasiliensis- namely, sterol 14-α-demethylase and serine/threonine-protein kinase TOR, respectively. These findings shed light on the potential binding affinities and binding modes of bifonazole and everolimus with their probable targets, providing a preliminary understanding of the antifungal mechanism of action of these compounds. In conclusion, our research advances the understanding of the therapeutic potential of bifonazole and everolimus, supporting their further investigation as antifungal agents for sporotrichosis in prospective hit-to-lead and preclinical investigations.

Fragment library screening by X-ray crystallography and binding site analysis on thioredoxin glutathione reductase of Schistosoma mansoni.

Schistosomiasis is caused by parasites of the genus Schistosoma, which infect more than 200 million people. Praziquantel (PZQ) has been the main drug for controlling schistosomiasis for over four decades, but despite that it is ineffective against juvenile worms and size and taste issues with its pharmaceutical forms impose challenges for treating school-aged children. It is also important to note that PZQ resistant strains can be generated in laboratory conditions and observed in the field, hence its extensive use in mass drug administration programs raises concerns about resistance, highlighting the need to search for new schistosomicidal drugs. Schistosomes survival relies on the redox enzyme thioredoxin glutathione reductase (TGR), a validated target for the development of new anti-schistosomal drugs. Here we report a high-throughput fragment screening campaign of 768 compounds against S. mansoni TGR (SmTGR) using X-ray crystallography. We observed 49 binding events involving 35 distinct molecular fragments which were found to be distributed across 16 binding sites. Most sites are described for the first time within SmTGR, a noteworthy exception being the "doorstop pocket" near the NADPH binding site. We have compared results from hotspots and pocket druggability analysis of SmTGR with the experimental binding sites found in this work, with our results indicating only limited coincidence between experimental and computational results. Finally, we discuss that binding sites at the doorstop/NADPH binding site and in the SmTGR dimer interface, should be prioritized for developing SmTGR inhibitors as new antischistosomal drugs.

Artificial intelligence-guided approach for efficient virtual screening of hits against Schistosoma mansoni.

Background: The impact of schistosomiasis, which affects over 230 million people, emphasizes the urgency of developing new antischistosomal drugs. Artificial intelligence is vital in accelerating the drug discovery process. Methodology & results: We developed classification and regression machine learning models to predict the schistosomicidal activity of compounds not experimentally tested. The prioritized compounds were tested on schistosomula and adult stages of Schistosoma mansoni. Four compounds demonstrated significant activity against schistosomula, with 50% effective concentration values ranging from 9.8 to 32.5 µM, while exhibiting no toxicity in animal and human cell lines. Conclusion: These findings represent a significant step forward in the discovery of antischistosomal drugs. Further optimization of these active compounds can pave the way for their progression into preclinical studies.

The CDR3 region as the major driver of TREM-1 interaction with its ligands, an in silico characterization.

The triggering receptor expressed on myeloid cells-1 (TREM-1) is a pattern recognition receptor heavily investigated in infectious and non-infectious diseases. Because of its role in amplifying inflammation, TREM-1 has been explored as a diagnostic/prognostic biomarker. Further, as the receptor has been implicated in the pathophysiology of several diseases, therapies aiming at modulating its activity represent a promising strategy to constrain uncontrolled inflammatory or infectious diseases. Despite this, several aspects concerning its interaction with ligands and activation process, remain unclear. Although many molecules have been suggested as TREM-1 ligands, only five have been confirmed to interact with the receptor: actin, eCIRP, HMGB1, Hsp70 and PGLYRP1. However, the domains involved in the interaction between the receptor and these proteins are not clarified yet. Therefore, here we used in silico approaches to investigate the putative binding domains in the receptor, using hot spots analysis, molecular docking and molecular dynamics simulations between TREM-1 and its five known ligands. Our results indicated the complementarity-determining regions (CDRs) of the receptor as the main mediators of antigen recognition, especially the CDR3 loop. We believe that our study could be used as structural basis for the elucidation of TREM-1's recognition process, and may be useful for prospective in silico and biological investigations exploring the receptor in different contexts.

In silico-chemogenomic repurposing of new chemical scaffolds for histoplasmosis treatment.

BACKGROUND: Histoplasmosis is a systemic form of endemic mycosis to the American continent and may be lethal to people living with HIV/AIDS. The drugs available for treating histoplasmosis are limited, costly, and highly toxic. New drug development is time-consuming and costly; hence, drug repositioning is an advantageous strategy for discovering new therapeutic options. OBJECTIVE: This study was conducted to identify drugs that can be repositioned for treating histoplasmosis in immunocompromised patients. METHODS: Homologous proteins among Histoplasma capsulatum strains were selected and used to search for homologous targets in the DrugBank and Therapeutic Target Database. Essential genes were selected using Saccharomyces cerevisiae as a model, and functional regions of the therapeutic targets were analyzed. The antifungal activity of the selected drugs was verified, and homology modeling and molecular docking were performed to verify the interactions between the drugs with low inhibitory concentration values and their corresponding targets. RESULTS: We selected 149 approved drugs with potential activity against histoplasmosis, among which eight were selected for evaluating their in vitro activity. For drugs with low minimum inhibitory concentration values, such as mebendazole, everolimus, butenafine, and bifonazole, molecular docking studies were performed. A chemogenomic framework revealed lanosterol 14-α-demethylase, squalene monooxygenase, serine/threonine-protein kinase mTOR, and the ß-4B tubulin chain of H. capsulatum, respectively, as the protein targets of the drugs. CONCLUSIONS: Our strategy can be used to identify promising antifungal targets, and drugs with repositioning potential for treating H. capsulatum.

Cheminformatics-driven discovery of hit compounds against Paracoccidioides spp.

Aims: The development of safe and effective therapies for treating paracoccidioidomycosis using computational strategies were employed to discover anti-Paracoccidioides compounds. Materials & methods: We 1) collected, curated and integrated the largest library of compounds tested against Paracoccidioides spp.; 2) employed a similarity search to virtually screen the ChemBridge database and select nine compounds for experimental evaluation; 3) performed an experimental evaluation to determine the minimum inhibitory concentration and minimum fungicidal concentration as well as cytotoxicity; and 4) employed computational tools to identify potential targets for the most active compounds. Seven compounds presented activity against Paracoccidioides spp. Conclusion: These compounds are new hits with a predicted mechanisms of action, making them potentially attractive to develop new compounds.

Hit-to-lead optimization of a pyrazinylpiperazine series against Leishmania infantum and Leishmania braziliensis.

An early hit-to-lead optimization of a novel pyrazinylpiperazine series against L. infantum and L. braziliensis has been performed after an extensive SAR focusing on the benzoyl fragment of hit (4). Deletion of the meta-Cl of (4) led to the obtention of the para-hydroxyl derivative (12), on which the design of most monosubstituted derivatives of the SAR was based. Further optimization of the series, involving disubstituted benzoyl fragments and the hydroxyl substituent of (12), allowed the obtention of a total of 15 compounds with increased antileishmanial potency (IC50 < 10 µM), nine of which displayed activity in the low micromolar range (IC50 < 5 µM). This optimization ultimately identified the ortho, meta-dihydroxyl derivative (46) as an early lead for this series (IC50 (L. infantum) = 2.8 µM, IC50 (L. braziliensis) = 0.2 µM). Additional assessment of some selected compounds against other trypanosomatid parasites revealed that this series is selective towards Leishmania parasites, and in silico ADMET predictions revealed satisfactory profiles for these compounds, allowing further lead optimization of the pyrazinylpiperazine class against Leishmania.

Multitask learning-driven identification of novel antitrypanosomal compounds.

Background: Chagas disease and human African trypanosomiasis cause substantial death and morbidity, particularly in low- and middle-income countries, making the need for novel drugs urgent. Methodology & results: Therefore, an explainable multitask pipeline to profile the activity of compounds against three trypanosomes (Trypanosoma brucei brucei, Trypanosoma brucei rhodesiense and Trypanosoma cruzi) were created. These models successfully discovered four new experimental hits (LC-3, LC-4, LC-6 and LC-15). Among them, LC-6 showed promising results, with IC50 values ranging 0.01-0.072 µM and selectivity indices >10,000. Conclusion: These results demonstrate that the multitask protocol offers predictivity and interpretability in the virtual screening of new antitrypanosomal compounds and has the potential to improve hit rates in Chagas and human African trypanosomiasis projects.

A structure-based approach for the discovery of inhibitors against methylcitrate synthase of Paracoccidioides lutzii.

Paracoccidioidomycosis (PCM) is a systemic mycosis, endemic in Latin America, caused by fungi of the genus Paracoccidioides. The treatment of PCM is complex, requiring a long treatment period, which often results in serious side effects. The aim of this study was to screen for inhibitors of a specific target of the fungus that is absent in humans. Methylcitrate synthase (MCS) is a unique enzyme of microorganisms and is responsible for the synthesis of methylcitrate at the beginning of the propionate degradation pathway. This pathway is essential for several microorganisms, since the accumulation of propionyl-CoA can impair virulence and prevent the development of the pathogen. We performed the modeling and molecular dynamics of the structure of Paracoccidioides lutzii MCS (PlMCS) and performed a virtual screening on 89,415 compounds against the active site of the enzyme. The compounds were selected according to the affinity and efficiency criteria of in vitro tests. Six compounds were able to inhibit the enzymatic activity of recombinant PlMCS but only the compound ZINC08964784 showed fungistatic and fungicidal activity against Paracoccidioides spp. cells. The analysis of the interaction profile of this compound with PlMCS showed its effectiveness in terms of specificity and stability when compared to the substrate (propionyl-CoA) of the enzyme. In addition, this compound did not show cytotoxicity in mammalian cells, with an excellent selectivity index. Our results suggest that the compound ZINC08964784 may become a promising alternative antifungal against Paracoccidioides spp. Communicated by Ramaswamy H. Sarma.

Natural Compounds as Non-Nucleoside Inhibitors of Zika Virus Polymerase through Integration of In Silico and In Vitro Approaches.

Although the past epidemic of Zika virus (ZIKV) resulted in severe neurological consequences for infected infants and adults, there are still no approved drugs to treat ZIKV infection. In this study, we applied computational approaches to screen an in-house database of 77 natural and semi-synthetic compounds against ZIKV NS5 RNA-dependent RNA-polymerase (NS5 RdRp), an essential protein for viral RNA elongation during the replication process. For this purpose, we integrated computational approaches such as binding-site conservation, chemical space analysis and molecular docking. As a result, we prioritized nine virtual hits for experimental evaluation. Enzymatic assays confirmed that pedalitin and quercetin inhibited ZIKV NS5 RdRp with IC50 values of 4.1 and 0.5 µM, respectively. Moreover, pedalitin also displayed antiviral activity on ZIKV infection with an EC50 of 19.28 µM cell-based assays, with low toxicity in Vero cells (CC50 = 83.66 µM) and selectivity index of 4.34. These results demonstrate the potential of the natural compounds pedalitin and quercetin as candidates for structural optimization studies towards the discovery of new anti-ZIKV drug candidates.