RESUMO
BACKGROUND: In recent years, an increase in the occurrence of illnesses caused by two clinically- important arboviruses has been reported: Zika virus (ZIKV) and Chikungunya virus (CHIKV). There is no licensed antiviral treatment for either of the two abovementioned viruses. Bearing in mind that the antiviral effect of indole alkaloids has been reported for other arboviral models, the present study proposed to evaluate the antiviral in vitro and in silico effects of four indole alkaloids on infections by these two viruses in different cell lines. METHODS: The antiviral effects of voacangine (VOAC), voacangine-7-hydroxyindolenine (VOAC-OH), rupicoline and 3-oxo voacangine (OXO-VOAC) were evaluated in Vero, U937 and A549 cells using different experimental strategies (Pre, Trans, Post and combined treatment). Viral infection was quantified by different methodologies, including infectious viral particles by plating, viral genome by RT-qPCR, and viral protein by cell ELISA. Moreover, molecular docking was used to evaluate the possible interactions between structural and nonstructural viral proteins and the compounds. The results obtained from the antiviral strategies for each experimental condition were compared in all cases with the untreated controls. Statistically significant differences were identified using a parametric Student's t-test. In all cases, p values below 0.05 (p < 0.05) were considered statistically significant. RESULTS: In the pre-treatment strategy in Vero cells, VOAC and VOAC-OH inhibited both viral models and OXO-VOAC inhibited only ZIKV; in U937 cells infected with CHIKV/Col, only VOAC-OH inhibited infection, but none of the compounds had activity in A549 cells; in U937 cells and A549 cells infected with ZIKV/Col, the three compounds that were effective in Vero cells also had antiviral activity. In the trans-treatment strategy, only VOAC-OH was virucidal against ZIKV/Col. In the post-treatment strategy, only rupicoline was effective in the CHIKV/Col model in Vero and A549 cells, whereas VOAC and VOAC-OH inhibited ZIKV infection in all three cell lines. In the combined strategy, VOAC, VOAC-OH and rupicoline inhibited CHIKV/Col and ZIKV/Col, but only rupicoline improved the antiviral effect of ZIKV/Col-infected cultures with respect to the individual strategies. Molecular docking showed that all the compounds had favorable binding energies with the structural proteins E2 and NSP2 (CHIKV) and E and NS5 (ZIKV). CONCLUSIONS: The present study demonstrates that indole alkaloids are promising antiviral drugs in the process of ZIKV and CHIKV infection; however, the mechanisms of action evaluated in this study would indicate that the effect is different in each viral model and, in turn, dependent on the cell line.
Assuntos
Antivirais/farmacologia , Febre de Chikungunya/tratamento farmacológico , Alcaloides Indólicos/farmacologia , Células Vero/efeitos dos fármacos , Infecção por Zika virus/tratamento farmacológico , Zika virus/efeitos dos fármacos , Animais , Chlorocebus aethiops/metabolismo , HumanosRESUMO
The growing spread of dengue, chikungunya and Zika viruses demand the development of new and environmentally safe control methods for their vector, the mosquito Aedes aegypti. This study aims to find novel larvicidal agents from mutualistic (endophytic and rhizospheric) or edaphic bacteria that have no action against non-target organisms. Eleven out of the 254 bacterial strains tested were able to kill Ae. aegypti larvae. Larvicidal activity did not depend on presence of cells, since culture supernatants or crude lipopeptide extracts (CLEs) killed the larvae. Bacillus safensis BacI67 and Bacillus paranthracis C21 supernatants were the best performing supernatants, displaying the lowest lethal concentrations (LC50 = 31.11 µL/mL and 45.84 µL/mL, respectively). Bacillus velezensis B64a and Bacillus velezensis B15 produced the best performing CLEs (LC50 = 0.11 mg/mL and 0.12 mg/mL, respectively). Mass spectrometry analysis of CLEs detected a mixture of surfactins, iturins, and fengycins. The samples tested were weakly- or non-toxic to mammalian cells (RAW 264.7 macrophages and VERO cells) and non-target organisms (Caenorhabditis elegans, Galleria mellonella, Scenedesmus obliquus, and Tetrahymena pyriformis) - especially B. velezensis B15 CLE. The biosynthetic gene clusters related to secondary metabolism identified by whole genome sequencing of the four best performing bacteria strains revealed clusters for bacteriocin, beta-lactone, lanthipeptide, non-ribosomal peptide synthetases, polyketide synthases (PKS), siderophores, T3PKS, type 1 PKS-like, terpenes, thiopeptides, and trans-AT-PKS. Purification of lipopeptides may clarify the mechanisms by which these extracts kill Ae. aegypti larvae.
Assuntos
Aedes/fisiologia , Bacillus/metabolismo , Controle de Mosquitos , Aedes/crescimento & desenvolvimento , Aedes/microbiologia , Animais , Caenorhabditis elegans/efeitos dos fármacos , Chlorocebus aethiops , Larva/crescimento & desenvolvimento , Larva/microbiologia , Larva/fisiologia , Camundongos , Mariposas/efeitos dos fármacos , Células RAW 264.7/efeitos dos fármacos , Scenedesmus/efeitos dos fármacos , Tetrahymena pyriformis/efeitos dos fármacos , Testes de Toxicidade , Células Vero/efeitos dos fármacosRESUMO
Phenolic compounds have been related to multiple biological activities, and the antiviral effect of these compounds has been demonstrated in several viral models of public health concern. In this review, we show the antiviral role of phenolic compounds against dengue virus (DENV), the most widespread arbovirus globally that, after its re-emergence, has caused multiple epidemic outbreaks, especially in the last two years. Twenty phenolic compounds with anti-DENV activity are discussed, including the multiple mechanisms of action, such as those directed against viral particles or viral proteins, host proteins or pathways related to the productive replication viral cycle and the spread of the infection.
Assuntos
Antivirais/uso terapêutico , Dengue/tratamento farmacológico , Fenóis/uso terapêutico , Replicação Viral/efeitos dos fármacos , Animais , Chlorocebus aethiops , Dengue/genética , Dengue/patologia , Dengue/virologia , Vírus da Dengue/efeitos dos fármacos , Vírus da Dengue/genética , Vírus da Dengue/patogenicidade , Humanos , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Células Vero/efeitos dos fármacos , Proteínas Virais/antagonistas & inibidores , Proteínas Virais/genéticaRESUMO
Aim: To evaluate the activity, cytotoxicity and efflux pumps inhibition of a series of 12 novels (-)-camphene-based 1,3,4-thiadiazoles (TDZs) against Mycobacterium tuberculosis (Mtb). Materials & methods: The minimum inhibitory concentration (MIC), cytotoxicity for three cell lines, ethidium bromide accumulation and checkerboard methods were carried out. Results: Compounds (6a, 6b, 6c, 6g, 6h and 6j) showed significant anti-Mtb activity (MIC 3.9-7.8 µg/ml) and no antagonism with anti-TB drugs already used in the TB treatment. Selectivity index (SI) was also determined, with values reaching 42.9 for H37Rv strain and 97.1 for clinical isolate. Five compounds also showed bacterial efflux pumps inhibition and one showed modulator effect with three drugs. Conclusion: These six TDZs should be considered as new scaffolds to develop anti-TB drugs.
Assuntos
Antituberculosos/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Tiadiazóis/farmacologia , Animais , Proteínas da Membrana Bacteriana Externa/efeitos dos fármacos , Células Sanguíneas/efeitos dos fármacos , Chlorocebus aethiops , Descoberta de Drogas , Sinergismo Farmacológico , Humanos , Macrófagos/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Ovinos/sangue , Terpenos/farmacologia , Tiadiazóis/síntese química , Tiadiazóis/toxicidade , Tuberculose/tratamento farmacológico , Células Vero/efeitos dos fármacosRESUMO
Combination therapy has been proposed as an ideal strategy to reduce drug toxicity and improve treatment efficacy in Chagas disease. Previously, we demonstrated potent in vivo anti-Trypanosoma cruzi activity of voriconazole. In this work, we aimed to study the synergistic effect of voriconazole (VCZ) and benznidazole (BZ) both in vitro and in vivo models of T. cruzi infection using the Tulahuen strain. Combining VCZ and BZ at fixed concentrations, the inhibitory concentration 50% (IC50) on amastigotes was lower than the obtained IC50 for BZ alone and the Fractional Inhibitory Concentration Index (∑FIC) suggested an in vitro additive effect on T. cruzi amastigotes inhibition at concentrations devoid of cytotoxic effects. Treatment response in the in vivo model was evaluated by comparing behavior and physical aspects, parasitemia and mortality of mice infected with Tulahuen strain. VCZ and BZ treatments alone or in combination were well tolerated. All treated animals displayed significantly lower mean peak parasitemia and mortality compared to infected non-treated controls (p< 0.05). However, VCZ + BZ combination elicited no additional benefits over BZ monotherapy. VCZ efficacy was not enhanced by combination therapy with BZ at the doses studied, requiring further and astringent non-clinical studies to establish the VCZ efficacy and eventually moving forward to clinical trials.
Assuntos
Doença de Chagas/tratamento farmacológico , Nitroimidazóis/farmacologia , Nitroimidazóis/uso terapêutico , Parasitemia/tratamento farmacológico , Tripanossomicidas/farmacologia , Tripanossomicidas/uso terapêutico , Voriconazol/uso terapêutico , Animais , Chlorocebus aethiops , Sinergismo Farmacológico , Técnicas In Vitro , Camundongos , Trypanosoma cruzi/efeitos dos fármacos , Células Vero/efeitos dos fármacos , Voriconazol/farmacologiaRESUMO
Accidents with venomous snakes are a major health hazard in tropical countries. Bothrops genus is responsible for almost 80% of snakebites in Brazil. Immunotherapy is the only approved specific treatment against snake toxins and the production of therapeutic antivenoms requires quality control tests to determine their neutralizing potency. Currently, these controls are performed by in vivo lethality neutralization, however, the inhibition of particular events produced by bothropic venoms such as coagulopathy, hemorrhage, edema or cytotoxic effects are also required. The aim of this work is to develop an in vitro alternative assay for antivenom pre-clinical evaluation. In this sense, we designed a cell viability assay using different amounts (0.2-10 µL/well) of low and high potency anti-bothropic sera, previously classified by the traditional in vivo test, for assessing the antivenom capacity to protect the cells against B. jararaca venom cytotoxicity (5xEC50â¯=â¯58.95⯵g/mL). We found that high potency sera are more effective in neutralizing B. jararaca venom cytotoxicity when compared to low potency sera, which is in accordance to their pre-determined in vivo potency. Considering sera in vitro inhibitory concentration able to prevent 50% cell death (IC50) and their known in vivo potency, a cut-off point was determined to discriminate low and high potency sera. Our data provide insights for the development of an in vitro method which can determine the anti-bothropic antivenom potency during its production.
Assuntos
Antivenenos/análise , Bothrops , Sobrevivência Celular/efeitos dos fármacos , Venenos de Crotalídeos/imunologia , Animais , Bioensaio , Chlorocebus aethiops , Ensaio de Imunoadsorção Enzimática , Feminino , Cavalos/sangue , Cavalos/imunologia , Técnicas In Vitro/métodos , Masculino , Células Vero/efeitos dos fármacosRESUMO
AIM: Novel 4-methoxy-naphthalene derivatives were synthesized based on hits structures in order to evaluate the antifungal activity against Paracoccidioides spp. METHODS: Antifungal activity of compounds was evaluated against P. brasiliensis and most promising compounds 2 and 3 were tested against eight clinically important fungal species. RESULTS: Compound 3 was the more active compound with MIC 8 to 32 µg.ml-1 for Paracoccidioides spp without toxicity monkey kidney and murine macrophagecells. Carbohydrazide 3 showed good synergistic antifungal activity with amphotericin B against P. brasiliensis specie. Titration assay of carbohydrazide 3 with PbHSD enzyme demonstrates the binding ligand-protein. Molecular dynamics simulations show that ligand 3 let the PbHSD protein more stable. CONCLUSION: New carbohydrazide 3 is an attractive lead for drug development to treat paracoccidioidomycoses.
Assuntos
Antifúngicos/farmacologia , Naftalenos/farmacologia , Paracoccidioides/efeitos dos fármacos , Paracoccidioidomicose/tratamento farmacológico , Anfotericina B/farmacologia , Animais , Antifúngicos/uso terapêutico , Chlorocebus aethiops , Combinação de Medicamentos , Sinergismo Farmacológico , Homosserina Desidrogenase/metabolismo , Hidrazinas/farmacologia , Macrófagos/efeitos dos fármacos , Camundongos , Testes de Sensibilidade Microbiana , Simulação de Dinâmica Molecular , Naftalenos/síntese química , Naftalenos/uso terapêutico , Paracoccidioides/patogenicidade , Estabilidade Proteica , Células Vero/efeitos dos fármacosRESUMO
The development of alternatives for the use of chemical pesticides for plant disease control is the present-day and ongoing challenge for achieving sustainable agriculture. Pseudomonas fluorescens SF4c, native strain from wheat, produces tailocins (phage-tail-like bacteriocins) with antimicrobial activity against several phytopathogenic strains. We thus investigated the efficacy of foliar application of these bacteriocins to control the bacterial-spot disease in tomato caused by Xanthomonas vesicatoria Xcv Bv5-4a. The disease severity and incidence index were reduced by 44 and 36%, respectively; while the number of viable cells of X. vesicatoria Xcv Bv5-4a decreased after bacteriocin treatment. Furthermore, bacteriocin was effective in reducing bacterial-spot-disease symptoms on tomato fruits even when applied 12â¯h after infection. Tailocin activity was not affected by abiotic influences such as adjuvant, light and temperature and, biotic factors such as apoplastic-fluids. In contrast, no antibacterial activity of these tailocins was observed when the bacteriocin was exposed to extremely dry conditions. Finally, that no cytotoxic effects on mammalian cells were observed with this representative tailocins is highly significant and demonstrates the safety of such compounds in humans. All these findings indicate that the SF4c tailocins represent an attractive alternative to copper-containing bactericides for use in the control of bacterial spot.
Assuntos
Infecções Bacterianas/tratamento farmacológico , Bacteriocinas/farmacologia , Doenças das Plantas/microbiologia , Doenças das Plantas/terapia , Pseudomonas fluorescens/metabolismo , Solanum lycopersicum/microbiologia , Xanthomonas vesicatoria/efeitos dos fármacos , Animais , Antibacterianos/biossíntese , Antibacterianos/farmacologia , Antibiose , Bacteriocinas/química , Bacteriocinas/isolamento & purificação , Bacteriocinas/metabolismo , Agentes de Controle Biológico/metabolismo , Agentes de Controle Biológico/farmacologia , Chlorocebus aethiops , Eritrócitos/efeitos dos fármacos , Luz , Solanum lycopersicum/crescimento & desenvolvimento , Folhas de Planta/citologia , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/microbiologia , Ovinos , Temperatura , Células Vero/efeitos dos fármacos , Xanthomonas vesicatoria/patogenicidadeRESUMO
In this study we evaluated the antioxidant capacity, antimicrobial activity, and cytotoxicity of an aqueous extract of the Pleurotus ostreatoroseus mushroom, which was cooked. Fresh basidiocarps were heated and steamed at 100°C and the resulting aqueous extract was assessed before and after in vitro digestion. Cooking reduced the amounts of phenolic compounds in the extract. The antioxidant activity of the extract was evaluated through the use of 4 methods. The lowest half-maximal effective concentration (EC50) against ABTS radicals was 0.057 ± 0.002 mg/mL for the uncooked basidiocarp extract. Cooking and the digestive process led to decreased activity (P > 0.05) against ABTS and DPPH radicals. A significant increase in chelating activity (P > 0.05) occurred after the basidiocarps were cooked (EC50 = 0.279 ± 0.007 mg/mL). The reducing power did not significantly change among the different extracts. The uncooked basidiocarp extract was cytotoxic to Vero cells. After cooking and subsequent in vitro digestion, the cytotoxicity of the extracts decreased (P < 0.05). Bacillus subtilis, Staphylococcus aureus, and Candida albicans were sensitive to the fresh mushroom extract. The data showed that after being cooked and digested, the P. ostreatoroseus mushroom maintains antioxidant activity and has a low cytotoxic effect.
Assuntos
Anti-Infecciosos/análise , Antioxidantes/análise , Misturas Complexas/química , Culinária , Digestão , Pleurotus/química , Animais , Bacillus subtilis/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Misturas Complexas/toxicidade , Temperatura Alta , Pleurotus/metabolismo , Pleurotus/efeitos da radiação , Staphylococcus aureus/efeitos dos fármacos , Células Vero/efeitos dos fármacosRESUMO
In this study, Moringa oleifera flower extract and a trypsin inhibitor (MoFTI) isolated from it were evaluated for anti-protozoal activity against Trypanosoma cruzi and cytotoxicity to mammalian cells. The presence of flavonoids was remarkable in the HPLC fingerprints of the extract at 254 and 360 nm. Amino acid sequences of peptides derived from in-gel digestion of MoFTI were determined. Both the extract and MoFTI caused lysis of T. cruzi trypomastigotes with LC50/24 h of 54.18 ± 6.62 and 41.20 ± 4.28 µg/mL, respectively. High selectivity indices (7.9 to >12) for T. cruzi cells over murine peritoneal macrophages and Vero cells were found for the extract and MoFTI. The results show that MoFTI is a trypanocidal principle of the flower extract.