Neuraminidase is a host-directed approach to regulate neutrophil responses in sepsis and COVID-19.

BACKGROUND AND PURPOSE: Neutrophil overstimulation plays a crucial role in tissue damage during severe infections. Because pathogen-derived neuraminidase (NEU) stimulates neutrophils, we investigated whether host NEU can be targeted to regulate the neutrophil dysregulation observed in severe infections. EXPERIMENTAL APPROACH: The effects of NEU inhibitors on lipopolysaccharide (LPS)-stimulated neutrophils from healthy donors or COVID-19 patients were determined by evaluating the shedding of surface sialic acids, cell activation, and reactive oxygen species (ROS) production. Re-analysis of single-cell RNA sequencing of respiratory tract samples from COVID-19 patients also was carried out. The effects of oseltamivir on sepsis and betacoronavirus-induced acute lung injury were evaluated in murine models. KEY RESULTS: Oseltamivir and zanamivir constrained host NEU activity, surface sialic acid release, cell activation, and ROS production by LPS-activated human neutrophils. Mechanistically, LPS increased the interaction of NEU1 with matrix metalloproteinase 9 (MMP-9). Inhibition of MMP-9 prevented LPS-induced NEU activity and neutrophil response. In vivo, treatment with oseltamivir fine-tuned neutrophil migration and improved infection control as well as host survival in peritonitis and pneumonia sepsis. NEU1 also is highly expressed in neutrophils from COVID-19 patients, and treatment of whole-blood samples from these patients with either oseltamivir or zanamivir reduced neutrophil overactivation. Oseltamivir treatment of intranasally infected mice with the mouse hepatitis coronavirus 3 (MHV-3) decreased lung neutrophil infiltration, viral load, and tissue damage. CONCLUSION AND IMPLICATIONS: These findings suggest that interplay of NEU1-MMP-9 induces neutrophil overactivation. In vivo, NEU may serve as a host-directed target to dampen neutrophil dysfunction during severe infections.

Anti-Mayaro virus activity of a hydroethanolic extract from Fridericia chica (Bonpl.) L. G. Lohmann leaves.

ETHNOPHARMACOLOGICAL RELEVANCE: Mayaro fever is a neglected tropical disease. The region of the most significant circulation of the Mayaro virus (MAYV) is the Amazon rainforest, situated in remote areas that are difficult to access and where medicine is scarce. Thus, the regional population uses plants as an alternative for the treatment of various diseases. Fridericia chica is an endemic plant of tropical regions used in traditional medicine to treat fever, malaise, inflammation, and infectious diseases such as hepatitis B. However, its antiviral activity is poorly understood. AIM OF THE STUDY: This study aimed to investigate the anti-MAYV activity of the hydroethanolic extract of the leaves of Fridericia chica (HEFc) in mammalian cells and its possible mechanism of action. MATERIALS AND METHODS: The antiviral activity of HEFc was studied using Vero cell lines against MAYV. The cytotoxicity and antiviral activity of the extract were evaluated by the 3-(4, 5- dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay. The overall antiviral activity was confirmed by the plaque forming units (PFU) method. Then, the effects of HEFc on MAYV multiplication kinetics, virus adsorption, penetration, and post-penetration, and its virucidal activity were determined in Vero cells using standard experimental procedures. RESULTS: HEFc exerted a effect against viral infection in Vero cells at a non-cytotoxic concentration, and no virion was detected in the supernatant in a dose-dependent and selective manner. HEFc inhibited MAYV in the early and late stages of the viral multiplication cycle. The extract showed significant virucidal activity at low concentrations and did not affect adsorption or viral internalization stages. In addition, HEFc reduced virions at all post-infection times investigated. CONCLUSIONS: HEFc has good antiviral activity against MAYV, acting directly on the viral particles. This plant extract possesses an excellent and promising potential for developing effective herbal antiviral drugs.

Neuraminidase inhibitors rewire neutrophil function in vivo in murine sepsis and ex vivo in COVID-19.

Neutrophil overstimulation plays a crucial role in tissue damage during severe infections. Neuraminidase (NEU)-mediated cleavage of surface sialic acid has been demonstrated to regulate leukocyte responses. Here, we report that antiviral NEU inhibitors constrain host NEU activity, surface sialic acid release, ROS production, and NETs released by microbial-activated human neutrophils. In vivo, treatment with Oseltamivir results in infection control and host survival in peritonitis and pneumonia models of sepsis. Single-cell RNA sequencing re-analysis of publicly data sets of respiratory tract samples from critical COVID-19 patients revealed an overexpression of NEU1 in infiltrated neutrophils. Moreover, Oseltamivir or Zanamivir treatment of whole blood cells from severe COVID-19 patients reduces host NEU-mediated shedding of cell surface sialic acid and neutrophil overactivation. These findings suggest that neuraminidase inhibitors can serve as host-directed interventions to dampen neutrophil dysfunction in severe infections.

Identification of a Potential Zika Virus Inhibitor Targeting NS5 Methyltransferase Using Virtual Screening and Molecular Dynamics Simulations.

The NS5 methyltransferase (MTase) has been reported as an attractive molecular target for antivirals discovery against the Zika virus (ZIKV). Here, we report structure-based virtual screening of 42 390 structures from the Development Therapeutics Program (DTP) AIDS Antiviral Screen Database. Among the docked compounds, ZINC1652386 stood out due to its high affinity for MTase in comparison to the cocrystallized ligand MS2042, which interacts with the Asp146 residue in the MTase binding site by hydrogen bonding. Subsequent molecular dynamics simulations predicted that this compound forms a stable complex with MTase within 50 ns. Thus, ZINC1652386 may represent a promising ZIKV methyltransferase inhibitor.

Identification of Zika Virus NS2B-NS3 Protease Inhibitors by Structure-Based Virtual Screening and Drug Repurposing Approaches.

The NS2B-NS3 protease has been identified as an attractive target for drug development against Zika virus (ZIKV) and combined drug repurposing and structure-based virtual screening has improved the development of antiviral drugs. In this study, we performed a structure-based virtual screening of 1861 Food and Administration (FDA) approved drugs available in DrugBank by the selection and docking validation of crystal structure of ZIKV NS2B-NS3 protease (PDB ID 5H4I ) using Glide and DOCK 6 software. The antihistaminic chlorcyclizine (Grid score -24.8 kcal/mol) exhibited the most promising interaction with NS2B-NS3 protease in comparison to crystallography ligand (Grid score -15.6 kcal/mol) by interaction to Tyr161 by hydrophobic interactions in the binding site of NS2B-NS3 which is recognized as an important amino acid in substrate molecular recognition. Cytotoxicity and global antiviral activity assay in Vero cells by MTT method showed that chlorcyclizine reduced the ZIKV induced cytopathic effect (EC50 of 69.0 ± 7.3 µM and SI = 1.9), and explicit molecular dynamics simulations implemented on a NAMD program indicated great stability of chlorcyclizine in protease binding site, suggesting the repurposing of chlorcyclizine as a promising finding in anti-ZIKV drug development.

Synthesis and Evaluation of the in vitro Antimicrobial Activity of Triazoles, Morpholines and Thiosemicarbazones.

BACKGROUND: Microbial infections is a global public health problem. The aim of this work was to synthesize and evaluate the antimicrobial activity of novel triazoles, morpholines and thiosemicarbazones. METHODS: Compounds were synthesized using 2,4-Dihydroxyacetophenone and 4-hydroxybenzaldehyde as starting materials. The antimicrobial activity of these compounds against bacteria and yeast was evaluated by the broth microdilution method. RESULTS: The proposed route for synthesis gave high to moderate yields, moreover these compounds were successfully characterized by 1H NMR, 13C NMR and LC-MS. Antimicrobial testing indicated that the thiosemicarbazone and morphine derivatives had the best antimicrobial activity against the microorganisms tested with minimum inhibitory concentrations (MIC) between 0.29 and 5.30 µM. Thiosemicarbazone derivative (12) was able to inhibit the growth of C. tropicalis, with minimum fungicidal concentration (MFC) of 0.55 µM. In addition, this compound was active against E. coli, S. aureus and S. epidermidis, with MIC values ranging from 0.29 to 1.11 µM. Moreover, the morpholine derivative (15) had an MIC value of 0.83 µM against C. albicans and E. coli. CONCLUSION: We have efficiently synthesized a series of eleven novel triazoles, thiosemicarbazones and morpholine derivatives using 2,4-Dihydroxyacetophenone and 4-hydroxybenzaldehyde as starting materials. Thiosemicarbazone derivative (12) showed promising antifungal and antibacterial activity and these findings suggest that this compound can be used as scaffolds to design new antimicrobial drugs.

Design, synthesis, biological activity and structure-activity relationship studies of chalcone derivatives as potential anti-Candida agents.

Vulvovaginal candidiasis (VVC) affects millions of women around the world every year. Candida albicans is the most frequently isolated pathogen in women and its rapid ability to develop resistance to first and second line therapies has boosted the search for new and effective antifungal agents. In this study, we show the in vitro anti-Candida activity of fifteen synthetic chalcone analogs and their antifungal potential in an in vivo model of VVC. Chalcone 12 showed potent antifungal effects, being able to inhibit the growth of Candida spp. at a concentration of 15.6 µg mL-1. In addition, mechanism of action studies have indicated the ergosterol fungal membrane as the target of this compound. Despite a considerable antifungal activity, the chalcone 12 showed high cytotoxicity in kidney cells lineages. Moreover, this compound was able to reduce Candida-associated virulence, impairing yeast-hyphal transition in C. albicans. An in vivo model of VVC showed that chalcone 12 significantly reduces the fungal load. Taken together, these findings showed that the chalcone 12 is a potent anti-Candida agent in vitro beyond of contribute to improve the fungal infection in a model of CVV. However, it showed low selectivity and high toxicity, suggesting molecular modifications to minimize these proprieties.

γ-Benzylidene digoxin derivatives synthesis and molecular modeling: Evaluation of anticancer and the Na,K-ATPase activity effect.

Cardiotonic steroids (CS), natural compounds with traditional use in cardiology, have been recently suggested to exert potent anticancer effects. However, the repertoire of molecules with Na,K-ATPase activity and anticancer properties is limited. This paper describes the synthesis of 6 new digoxin derivatives substituted (on the C17-butenolide) with γ-benzylidene group and their cytotoxic effect on human fibroblast (WI-26 VA4) and cancer (HeLa and RKO) cell lines as well as their effect on Na,K-ATPase activity and expression. As digoxin, compound BD-4 was almost 100-fold more potent than the other derivatives for cytotoxicity with the three types of cells used and was also the only one able to fully inhibit the Na,K-ATPase of HeLa cells after 24h treatment. No change in the Na,K-ATPase α1 isoform protein expression was detected. On the other hand it was 30-40 fold less potent for direct Na,K-ATPase inhibition, when compared to the most potent derivatives, BD-1 and BD-3, and digoxin. The data presented here demonstrated that the anticancer effect of digoxin derivatives substituted with γ-benzylidene were not related with their inhibition of Na,K-ATPase activity or alteration of its expression, suggesting that this classical molecular mechanism of CS is not involved in the cytotoxic effect of our derivatives.