scholarly journals The Combined Use of Alphavirus Replicons and Pseudoinfectious Particles for the Discovery of Antivirals Derived from Natural Products

2014 ◽  
Vol 20 (5) ◽  
pp. 673-680 ◽  
Author(s):  
Phillip C. Delekta ◽  
Avi Raveh ◽  
Martha J. Larsen ◽  
Pamela J. Schultz ◽  
Giselle Tamayo-Castillo ◽  
...  
Keyword(s):  
Life Cycle ◽  
Antiviral Activity ◽  
Viral Entry ◽  
Particle System ◽  
Antiviral Agents ◽  
Viral Life Cycle ◽  
Genome Replication ◽  
Bacterial Strains ◽  
Antiviral Compounds ◽  
Western Equine Encephalitis

Alphaviruses are a prominent class of reemergent pathogens due to their globally expanding ranges, potential for lethality, and possible use as bioweapons. The absence of effective treatments for alphaviruses highlights the need for innovative strategies to identify antiviral agents. Primary screens that use noninfectious self-replicating RNAs, termed replicons, have been used to identify potential antiviral compounds for alphaviruses. Only inhibitors of viral genome replication, however, will be identified using replicons, which excludes many other druggable steps in the viral life cycle. To address this limitation, we developed a western equine encephalitis virus pseudoinfectious particle system that reproduces several crucial viral life cycle steps in addition to genome replication. We used this system to screen a library containing ~26,000 extracts derived from marine microbes, and we identified multiple bacterial strains that produce compounds with potential antiviral activity. We subsequently used pseudoinfectious particle and replicon assays in parallel to counterscreen candidate extracts, and followed antiviral activity during biochemical fractionation and purification to differentiate between inhibitors of viral entry and genome replication. This novel process led to the isolation of a known alphavirus entry inhibitor, bafilomycin, thereby validating the approach for the screening and identification of potential antiviral compounds.

scholarly journals Biological Evaluation of Molecules of the azaBINOL Class as Antiviral Agents: Specific Inhibition of HIV-1 RNase H Activity by 7-Isopropoxy-8-(naphth-1-yl)quinoline

2019 ◽  
Author(s):  
Ross D. Overacker ◽  
Somdev Banerjee ◽  
George F. Neuhaus ◽  
Selena Milicevic Sephton ◽  
Alexander Herrmann ◽  
...  
Keyword(s):  
Viral Entry ◽  
Antiviral Agents ◽  
Biological Evaluation ◽  
Rnase H ◽  
Synthetic Compounds ◽  
Infectivity Assay ◽  
Antiviral Compounds ◽  
Specific Manner ◽  
Drug Leads ◽  
Hiv 1

AbstractInspired by bioactive biaryl-containing natural products found in plants and the marine environment, a series of synthetic compounds belonging to the azaBINOL chiral ligand family was evaluated for antiviral activity against HIV-1. Testing of 39 unique azaBINOLs in a singleround infectivity assay resulted in the identification of three promising antiviral compounds, including 7-isopropoxy-8-(naphth-1-yl)quinoline (azaBINOLB#24), which exhibited low-micromolar activity. The active compounds and several close structural analogues were further tested against three different HIV-1 envelope pseudotyped viruses as well as in a full-virus replication system (EASY-HIT). Mode-of-action studies using a time-of-addition assay indicated that azaBINOLB#24acts after viral entry but before viral assembly and budding. HIV-1 reverse transcriptase (RT) assays that individually test for polymerase and RNase H activity were used to demonstrate thatB#24inhibits RNase H activity, most likely allosterically. Further binding analysis using bio-layer interferometry (BLI) showed thatB#24interacts with HIV-1 RT in a highly specific manner. These results indicate that azaBINOLB#24is a potentially viable, novel lead for the development of new HIV-1 RNase H inhibitors. Furthermore, this study demonstrates that the survey of libraries of synthetic compounds, designed purely with the goal of facilitating chemical synthesis in mind, may yield unexpected and selective drug leads for the development of new antiviral agents.

scholarly journals A Map of the Arenavirus Nucleoprotein-Host Protein Interactome Reveals that Junín Virus Selectively Impairs the Antiviral Activity of Double-Stranded RNA-Activated Protein Kinase (PKR)

2017 ◽  
Vol 91 (15) ◽  
Author(s):  
Benjamin R. King ◽  
Dylan Hershkowitz ◽  
Philip L. Eisenhauer ◽  
Marion E. Weir ◽  
Christopher M. Ziegler ◽  
...  
Keyword(s):  
Life Cycle ◽  
Protein Kinase ◽  
Antiviral Activity ◽  
Viral Life Cycle ◽  
Host Protein ◽  
Data Set ◽  
Double Stranded Rna ◽  
Junin Virus ◽  
Protein Interactome ◽  
Junín Virus

ABSTRACT Arenaviruses are enveloped negative-strand RNA viruses that cause significant human disease. These viruses encode only four proteins to accomplish the viral life cycle, so each arenavirus protein likely plays unappreciated accessory roles during infection. Here we used immunoprecipitation and mass spectrometry to identify human proteins that interact with the nucleoproteins (NPs) of the Old World arenavirus lymphocytic choriomeningitis virus (LCMV) and the New World arenavirus Junín virus (JUNV) strain Candid #1. Bioinformatic analysis of the identified protein partners of NP revealed that host translation appears to be a key biological process engaged during infection. In particular, NP associates with the double-stranded RNA (dsRNA)-activated protein kinase (PKR), a well-characterized antiviral protein that inhibits cap-dependent protein translation initiation via phosphorylation of eIF2α. JUNV infection leads to increased expression of PKR as well as its redistribution to viral replication and transcription factories. Further, phosphorylation of PKR, which is a prerequisite for its ability to phosphorylate eIF2α, is readily induced by JUNV. However, JUNV prevents this pool of activated PKR from phosphorylating eIF2α, even following exposure to the synthetic dsRNA poly(I·C), a potent PKR agonist. This blockade of PKR function is highly specific, as LCMV is unable to similarly inhibit eIF2α phosphorylation. JUNV's ability to antagonize the antiviral activity of PKR appears to be complete, as silencing of PKR expression has no impact on viral propagation. In summary, we provide a detailed map of the host machinery engaged by arenavirus NPs and identify an antiviral pathway that is subverted by JUNV. IMPORTANCE Arenaviruses are important human pathogens for which FDA-approved vaccines do not exist and effective antiviral therapeutics are needed. Design of antiviral treatment options and elucidation of the mechanistic basis of disease pathogenesis will depend on an increased basic understanding of these viruses and, in particular, their interactions with the host cell machinery. Identifying host proteins critical for the viral life cycle and/or pathogenesis represents a useful strategy to uncover new drug targets. This study reveals, for the first time, the extensive human protein interactome of arenavirus nucleoproteins and uncovers a potent antiviral host protein that is neutralized during Junín virus infection. In so doing, it shows further insight into the interplay between the virus and the host innate immune response and provides an important data set for the field.

Discovery of Bioactive Natural Products for the Treatment of Acute Respiratory Infections – An Integrated Approach

Planta Medica ◽  
2018 ◽  
Vol 84 (09/10) ◽  
pp. 684-695 ◽  
Author(s):  
Ulrike Grienke ◽  
Christina Mair ◽  
Johannes Kirchmair ◽  
Michaela Schmidtke ◽  
Judith Rollinger
Keyword(s):  
Antiviral Activity ◽  
Respiratory Infections ◽  
Antiviral Agents ◽  
Integrated Approach ◽  
Coxsackie Virus ◽  
Acute Respiratory Infections ◽  
Bioactive Natural Products ◽  
Antiviral Compounds ◽  
Antiviral Activities ◽  
Selection Of

AbstractIn this work, an integrated approach for the identification of new antiviral agents from natural sources for the treatment of acute respiratory infections is presented. The approach comprises (i) the selection of starting material based on traditional knowledge, (ii) phenotypic screening of extracts for antiviral activity, and (iii) the implementation of in silico predictions to identify antiviral compounds and derive the molecular mechanism underlying their biological activity. A variety of starting materials from plants and fungi was selected for the production of 162 extracts. These extracts were tested in cytopathic effect inhibition assays against influenza virus A/Hong Kong/68 (HK/68), rhinovirus A2 (RV-A2), and coxsackie virus B3 (CV-B3). All extracts were also evaluated regarding their cytotoxicity. At an IC50 threshold of 50 µg/mL, 20, 11, and 14% of all tested extracts showed antiviral activity against HK/68, CV-B3, and RV-A2, respectively. Among all active extracts (n = 47), 68% showed antiviral activity against one of the investigated viruses, whereas 31% inhibited at least two viruses. Herein, we present a comprehensive dataset of probed extracts along with their antiviral activities and cytotoxicity. Application examples presented in this work illustrate the phytochemical workflow for the identification of antiviral natural compounds. We also discuss the challenges, pitfalls, and advantages of the integrated approach.

scholarly journals Ceramide and Related Molecules in Viral Infections

2021 ◽  
Vol 22 (11) ◽  
pp. 5676
Author(s):  
Nadine Beckmann ◽  
Katrin Anne Becker
Keyword(s):  
Life Cycle ◽  
Viral Entry ◽  
Immune Modulation ◽  
Viral Infections ◽  
Therapeutic Benefit ◽  
Viral Life Cycle ◽  
Endosomal Escape ◽  
Sphingolipid Metabolism ◽  
Cell Surface Expression ◽  
Membrane Microdomains

Ceramide is a lipid messenger at the heart of sphingolipid metabolism. In concert with its metabolizing enzymes, particularly sphingomyelinases, it has key roles in regulating the physical properties of biological membranes, including the formation of membrane microdomains. Thus, ceramide and its related molecules have been attributed significant roles in nearly all steps of the viral life cycle: they may serve directly as receptors or co-receptors for viral entry, form microdomains that cluster entry receptors and/or enable them to adopt the required conformation or regulate their cell surface expression. Sphingolipids can regulate all forms of viral uptake, often through sphingomyelinase activation, and mediate endosomal escape and intracellular trafficking. Ceramide can be key for the formation of viral replication sites. Sphingomyelinases often mediate the release of new virions from infected cells. Moreover, sphingolipids can contribute to viral-induced apoptosis and morbidity in viral diseases, as well as virus immune evasion. Alpha-galactosylceramide, in particular, also plays a significant role in immune modulation in response to viral infections. This review will discuss the roles of ceramide and its related molecules in the different steps of the viral life cycle. We will also discuss how novel strategies could exploit these for therapeutic benefit.

scholarly journals Antiviral activity of silymarin and baicalein against dengue virus

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Zhao Xuan Low ◽  
Brian Ming OuYong ◽  
Pouya Hassandarvish ◽  
Chit Laa Poh ◽  
Babu Ramanathan
Keyword(s):  
Antiviral Activity ◽  
Dengue Virus ◽  
Viral Entry ◽  
Dengue Infection ◽  
Antiviral Agents ◽  
Antiviral Drug ◽  
Vero Cells ◽  
Viral Envelope ◽  
E Protein

AbstractDengue is an arthropod-borne viral disease that has become endemic and a global threat in many countries with no effective antiviral drug available currently. This study showed that flavonoids: silymarin and baicalein could inhibit the dengue virus in vitro and were well tolerated in Vero cells with a half-maximum cytotoxic concentration (CC50) of 749.70 µg/mL and 271.03 µg/mL, respectively. Silymarin and baicalein exerted virucidal effects against DENV-3, with a selective index (SI) of 10.87 and 21.34, respectively. Baicalein showed a better inhibition of intracellular DENV-3 progeny with a SI of 7.82 compared to silymarin. Baicalein effectively blocked DENV-3 attachment (95.59%) to the Vero cells, while silymarin prevented the viral entry (72.46%) into the cells, thus reducing viral infectivity. Both flavonoids showed promising antiviral activity against all four dengue serotypes. The in silico molecular docking showed that silymarin could bind to the viral envelope (E) protein with a binding affinity of − 8.5 kcal/mol and form hydrogen bonds with the amino acids GLN120, TRP229, ASN89, and THR223 of the E protein. Overall, this study showed that silymarin and baicalein exhibited potential anti-DENV activity and could serve as promising antiviral agents for further development against dengue infection.

scholarly journals Rapid Prescreening for Antiviral Agents against HIV-1 Based on Their Inhibitory Activity in Site-Directed Immunoassays. II. Porphyrins Reacting with the V3 Loop of gp120

1992 ◽  
Vol 3 (1) ◽  
pp. 55-63 ◽  
Author(s):  
A. R. Neurath ◽  
N. Strick ◽  
P. Haberfield ◽  
S. Jiang
Keyword(s):  
Antiviral Activity ◽  
Inhibitory Activity ◽  
Antiviral Agents ◽  
V3 Loop ◽  
Antiviral Compounds ◽  
Porphyrin Derivatives ◽  
Immunodeficiency Virus ◽  
Glycoprotein Gp120 ◽  
The Relationship ◽  
Hiv 1

Recent observations that haernin inhibited the replication of the human immunodeficiency virus (HIV-1) and the reaction between the HIV-1 envelope glycoprotein gp120 and antibodies specific for the V3 hypervariable loop of this glycoprotein were an enticement to determine whether or not additional porphyrins had similar activities. Several porphyrin derivatives, particularly meso-tetra (4-carboxyphenyl) porphine, were more potent inhibitors of HIV-1 replication than haernin. They blocked the binding of homologous antibodies to synthetic peptides corresponding to V3 hypervariable loops of 21 distinct HIV-1 isolates, and inhibited the replication in lymphocytic (MT-2) and promonocyte (U937) cell lines of several HIV-1 isolates, tested (IIIB, RF, SF-2, and MN). Compounds with inhibitory activity had a tetrapyrrole ring and, carboxyl or sulphonate groups. However, antiviral activity depended on minor structural difference's between distinct derivatives endowed with these two features. Metalloporphyrins had a drastically reduced antiviral activity in comparison with the corresponding porphyrins. An understanding of the relationship between the structure of porphyrins and their antiviral effects, perceptible from the results presented, is expected to lead to the design of additional derivatives with more potent antiviral activity and to unravelling of molecular details involved in the association between the V3 loop of gp120 and antiviral compounds targeted to this loop.

scholarly journals Investigation of monotherapy and combined anticoronaviral therapies against feline coronavirus serotype II in vitro

2021 ◽  
pp. 1098612X2110486
Author(s):  
Sarah E Cook ◽  
Helena Vogel ◽  
Diego Castillo ◽  
Mark Olsen ◽  
Niels Pedersen ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Antiviral Agents ◽  
Clinical Signs ◽  
Treatment Strategies ◽  
Antiviral Effect ◽  
Ocular Involvement ◽  
Virus Infections ◽  
Viral Inhibition ◽  
Antiviral Compounds

Objectives Feline infectious peritonitis (FIP), caused by genetic mutants of feline enteric coronavirus known as FIPV, is a highly fatal disease of cats with no currently available vaccine or US Food and Drug Administration-approved cure. Dissemination of FIPV in affected cats results in a range of clinical signs, including cavitary effusions, anorexia, fever and lesions of pyogranulomatous vasculitis and perivasculitis, with or without central nervous system or ocular involvement. The objectives of this study were to screen an array of antiviral compounds for anti-FIPV (serotype II) activity, determine cytotoxicity safety profiles of identified compounds with anti-FIPV activity and strategically combine identified monotherapies to assess compound synergy against FIPV in vitro. Based upon clinically successful combination treatment strategies for human patients with HIV and hepatitis C virus infections, we hypothesized that a combined anticoronaviral therapy approach featuring concurrent multiple mechanisms of drug action would result in an additive or synergistic antiviral effect. Methods This study screened 90 putative antiviral compounds for efficacy and cytotoxicity using a multimodal in vitro strategy, including plaque bioassays, real-time RT-PCR viral inhibition and cytotoxicity assays. Results Through this process, we identified 26 compounds with effective antiviral activity against FIPV, representing a variety of drug classes and mechanisms of antiviral action. The most effective compounds include GC376, GS-441524, EIDD2081 and EIDD2931. We documented antiviral efficacy for combinations of antiviral agents, with a few examined drug combinations demonstrating evidence of limited synergistic antiviral activity. Conclusions and relevance Although evidence of compound synergy was identified for several combinations of antiviral agents, monotherapies were ultimately determined to be the most effective in the inhibition of viral transcription.

Recent Advances in Our Understanding of Receptor Binding, Viral Fusion and Cell Entry of Hepatitis C Virus: New Targets for the Design of Antiviral Agents

2007 ◽  
Vol 18 (4) ◽  
pp. 169-189 ◽  
Author(s):  
Pantelis Poumbourios ◽  
Heidi E Drummer
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Receptor Binding ◽  
Viral Entry ◽  
Antiviral Agents ◽  
Genome Replication ◽  
Type I ◽  
Viral Fusion ◽  
Multiple Drugs

Improvements to antiviral therapies for the treatment of hepatitis C virus (HCV) infections will require the use of multiple drugs that target viral proteins essential for replication. The discovery of anti-HCV compounds has been severely hampered by the lack of cell culture replication systems. Since the late 1990s, the advent of sub-genomic replicons that model the intracellular events leading to HCV genome replication have enabled the discovery of HCV protease and polymerase inhibitors, but did not allow the study of HCV entry or entry inhibitors. More recently, retroviral pseudotyping of the viral glycoproteins and the development of a cell culture-based system that recapitulates the entire HCV replication cycle were achieved. These new experimental systems have enabled a rapid advance in our knowledge of how HCV glycoproteins, E1 and E2, mediate receptor binding and viral entry. These systems have facilitated the discovery of a range of viral receptors. Evidence is emerging that CD81, scavenger receptor class B type I, claudin-1 and the low-density lipoprotein receptor are involved in viral entry. In addition, DC-SIGN and L-SIGN may function to internalize virus into dendritic or endothelial cells, facilitating the transport of virions to sites of infection such as the liver. This review focuses on the interaction between the HCV glycoproteins and cellular receptors, and our current understanding of the viral entry pathway. In addition, key questions on the role that these receptors play in viral entry are raised and potential avenues for the discovery of new antiviral agents are highlighted.

scholarly journals Analysis of the Replication Mechanisms of the Human Papillomavirus Genomes

2021 ◽  
Vol 12 ◽  
Author(s):  
Lisett Liblekas ◽  
Alla Piirsoo ◽  
Annika Laanemets ◽  
Eva-Maria Tombak ◽  
Airiin Laaneväli ◽  
...  
Keyword(s):  
Life Cycle ◽  
Viral Genome ◽  
Maintenance Phase ◽  
Viral Life Cycle ◽  
Human Papillomaviruses ◽  
Genome Replication ◽  
Viral Genomes ◽  
Viral Genome Replication ◽  
Infected Cells ◽  
Cell Genome

The life-cycle of human papillomaviruses (HPVs) includes three distinct phases of the viral genome replication. First, the viral genome is amplified in the infected cells, and this amplification is often accompanied by the oligomerization of the viral genomes. Second stage includes the replication of viral genomes in concert with the host cell genome. The viral genome is further amplified during the third stage of the viral-life cycle, which takes place only in the differentiated keratinocytes. We have previously shown that the HPV18 genomes utilize at least two distinct replication mechanisms during the initial amplification. One of these mechanisms is a well-described bidirectional replication via theta type of replication intermediates. The nature of another replication mechanism utilized by HPV18 involves most likely recombination-dependent replication. In this paper, we show that the usage of different replication mechanisms is a property shared also by other HPV types, namely HPV11 and HPV5. We further show that the emergence of the recombination dependent replication coincides with the oligomerization of the viral genomes and is dependent on the replicative DNA polymerases. We also show that the oligomeric genomes of HPV18 replicate almost exclusively using recombination dependent mechanism, whereas monomeric HPV31 genomes replicate bi-directionally during the maintenance phase of the viral life-cycle.

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