RESUMEN

Arboviruses are etiological agents in an extensive group of emerging diseases with great clinical relevance in Brazil, due to the wide distribution of their vectors and the favorable environmental conditions. Among them, the Mayaro virus (MAYV) has drawn attention since its emergence as the etiologic agent of Mayaro fever, a highly debilitating disease. To study viral replication and identify new drug candidates, traditional antiviral assays based on viral antigens and/or plaque assays have been demonstrating low throughput, making it difficult to carry out larger-scale assays. Therefore, we developed and characterized two DNA-launched infectious clones reporter viruses based on the MAYV strain BeAr 20290 containing the reporter genes of firefly luciferase (FLuc) and nanoluciferase (NLuc), designated as MAYV-firefly and MAYV-nanoluc, respectively. The viruses replicated efficiently with similar properties to the parental wild-type MAYV, and luminescence expression levels reflected viral replication. Reporter genes were also preserved during passage in cell culture, remaining stably expressed for one round of passage for MAYV-firefly and three rounds for MAYV-nanoluc. Employing the infectious clone, we described the effect of Rimantadine, an FDA-approved Alzheimer's drug, as a repurposing agent for MAYV but with a broad-spectrum activity against Zika virus infection. Additionally, we validated MAYV-nanoluc as a tool for antiviral drug screening using the compound EIDD-2749 (4'-Fluorouridine), which acts as an inhibitor of alphavirus RNA-dependent RNA polymerase.

RESUMEN

Nitric oxide (NO) acts in different physiological processes, such as blood pressure control, antiparasitic activities, neurotransmission, and antitumor action. Among the exogenous NO donors, ruthenium nitrosyl/nitro complexes are potential candidates for prodrugs, due to their physicochemical properties, such as thermal and physiological pH stability. In this work, we proposed the synthesis and physical characterization of the new nitro terpyridine ruthenium (II) complexes of the type [RuII(L)(NO2)(tpy)]PF6 where tpy = 2,2':6',2″-terpyridine; L = 3,4-diaminobenzoic acid (bdq) or o-phenylenediamine (bd) and evaluation of influence of diimine bidentate ligand NH.NHq-R (R = H or COOH) in the HSA/DNA interaction as well as antiviral activity. The interactions between HSA and new nitro complexes [RuII(L)(NO2)(tpy)]+ were evaluated. The Ka values for the HSA-[RuII(bdq)(NO2)(tpy)]+ is 10 times bigger than HSA-[RuII(bd)(NO2)(tpy)]+. The sites of interaction between HSA and the complexes via synchronous fluorescence suppression indicate that the [RuII(bdq)(NO2)(tpy)]+ is found close to the Trp-241 residue, while the [RuII(bd)(NO2)(tpy)]+ complex is close to Tyr residues. The interaction with fish sperm fs-DNA using direct spectrophotometric titration (Kb) and ethidium bromide replacement (KSV and Kapp) showed weak interaction in the system fs-DNA-[RuII(bdq)(NO)(tpy)]+. Furthermore, fs-DNA-[RuII(bd)(NO2)(tpy)]+ and fs-DNA-[RuII(bd)(NO)(tpy)]3+ system showed higher intercalation constant. Circular dichroism spectra for fs-DNA-[RuII(bd)(NO2)(tpy)]+ and fs-DNA-[RuII(bd)(NO)(tpy)]3+, suggest semi-intercalative accompanied by major groove binding interaction modes. The [RuII(bd)(NO2)(tpy)]+ and [RuII(bd)(NO)(tpy)]3+ inhibit replication of Zika and Chikungunya viruses based in the nitric oxide release under S-nitrosylation reaction with cysteine viral.

Asunto(s)

Antivirales , ADN , Rutenio , Humanos , ADN/metabolismo , ADN/química , Rutenio/química , Rutenio/farmacología , Antivirales/farmacología , Antivirales/química , Antivirales/metabolismo , Ligandos , Animales , Albúmina Sérica Humana/química , Albúmina Sérica Humana/metabolismo , Piridinas/química , Piridinas/farmacología , Iminas/química , Iminas/farmacología , Complejos de Coordinación/farmacología , Complejos de Coordinación/química , Complejos de Coordinación/metabolismo

RESUMEN

Zika virus infection is associated to severe diseases such as congenital microcephaly and Zika fever causing serious harm to humans and special concern to health systems in low-income countries. Currently, there are no approved drugs against the virus, and the development of anti-Zika virus drugs is thus urgent. The present investigation describes the discovery and hit expansion of a N-acyl-2-aminobenzothiazole series of compounds against Zika virus replication. A structure-activity relationship study was obtained with the synthesis and evaluation of anti-Zika virus activity and cytotoxicity on Vero cells of nineteen derivatives. The three optimized compounds were 2.2-fold more potent than the initial hit and 20.9, 7.7 and 6.4-fold more selective. Subsequent phenotypic and biochemical assays were performed to evidence whether non-structural proteins, such as the complex NS2B-NS3pro, are related to the mechanism of action of the most active compounds.

Asunto(s)

Infección por el Virus Zika , Virus Zika , Animales , Chlorocebus aethiops , Humanos , Células Vero , Infección por el Virus Zika/tratamiento farmacológico , Relación Estructura-Actividad , Replicación Viral , Antivirales/química , Proteínas no Estructurales Virales

RESUMEN

Zika virus is the etiologic agent of Zika fever, and has been previously associated with cases of microcephaly, drawing the attention of the health authorities worldwide. However, no vaccine or antiviral are currently available. Phospholipases A2 (PLA2) isolated from snake venoms have demonstrated antiviral activity against several viruses. Here we demonstrated the anti-ZIKV activity of bothropstoxins-I and II (BthTX-I and II) isolated from Bothrops jararacussu venom. Vero E6 cells were infected with ZIKVPE243 in the presence of compounds for 72 h, when virus titers were evaluated. BthTX-I and II presented strong dose-dependent inhibition of ZIKV, with a SI of 149.1 and 1.44 × 105, respectively. These toxins mainly inhibited the early stages of the replicative cycle, such as during the entry of ZIKV into host cells, as shown by the potent virucidal effect, suggesting the action of these toxins on the virus particles. Moreover, BthTX-I and II presented significant activity towards post-entry stages of the ZIKV replicative cycle. Molecular docking analyses showed that BthTX-I and II potentially interact with DII and DIII domains from ZIKV Envelope protein. Our findings show that these PLA2s could be used as useful templates for the development of future antiviral candidate drugs against Zika fever.

Asunto(s)

Bothrops , Venenos de Crotálidos , Infección por el Virus Zika , Virus Zika , Animales , Humanos , Antivirales/farmacología , Bothrops/metabolismo , Infección por el Virus Zika/tratamiento farmacológico , Simulación del Acoplamiento Molecular , Venenos de Crotálidos/metabolismo , Anticuerpos

RESUMEN

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.