scholarly journals Peptidyl Aldehyde NK-1.8k Suppresses Enterovirus 71 and Enterovirus 68 Infection by Targeting Protease 3C

2015 ◽  
Vol 59 (5) ◽  
pp. 2636-2646 ◽  
Author(s):  
Yaxin Wang ◽  
Ben Yang ◽  
Yangyang Zhai ◽  
Zheng Yin ◽  
Yuna Sun ◽  
...  
Keyword(s):  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Binding Pocket ◽  
The United States ◽  
Single Amino Acid ◽  
Global Public Health ◽  
Causative Agents ◽  
Low Cytotoxicity ◽  
Neurological Infections

ABSTRACTEnterovirus (EV) is one of the major causative agents of hand, foot, and mouth disease in the Pacific-Asia region. In particular, EV71 causes severe central nervous system infections, and the fatality rates from EV71 infection are high. Moreover, an outbreak of respiratory illnesses caused by an emerging EV, EV68, recently occurred among over 1,000 young children in the United States and was also associated with neurological infections. Although enterovirus has emerged as a considerable global public health threat, no antiviral drug for clinical use is available. In the present work, we screened our compound library for agents targeting viral protease and identified a peptidyl aldehyde, NK-1.8k, that inhibits the proliferation of different EV71 strains and one EV68 strain and that had a 50% effective concentration of 90 nM. Low cytotoxicity (50% cytotoxic concentration, >200 μM) indicated a high selective index of over 2,000. We further characterized a single amino acid substitution inside protease 3C (3Cpro), N69S, which conferred EV71 resistance to NK-1.8k, possibly by increasing the flexibility of the substrate binding pocket of 3Cpro. The combination of NK-1.8k and an EV71 RNA-dependent RNA polymerase inhibitor or entry inhibitor exhibited a strong synergistic anti-EV71 effect. Our findings suggest that NK-1.8k could potentially be developed for anti-EV therapy.

scholarly journals Inhibition of Enterovirus 71 by Picochlorum sp. 122 via AKT and ATM/ATR Signaling Pathways

2021 ◽  
Author(s):  
Min Guo ◽  
Ruilin Zheng ◽  
Hua-lian Wu ◽  
Danyang Chen ◽  
Jingyao Su ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Signaling Pathways ◽  
Antiviral Agents ◽  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Vero Cells ◽  
Host Cells ◽  
Global Public Health ◽  
Protein Levels

Abstract Enterovirus 71 (EV71) pose a critical threat in global public health and may lead to severe and even lethal cases of hand-foot-and-mouth disease (HFMD). No effective antiviral agents are available to the masses for treatment of HFMD caused by EV71. Polysaccharide provides a good clinical application for antivirus. Polysaccharides extracted from Picochlorum sp. 122 (PPE) is a kind of seaweed Polysaccharides, the reports on its antiviral activity are limited. In this study, the antiviral activity was verified in Vero cells. Briefly, PPE has been demonstrated to restrain EV71 infection through MTT assay and cellular cytopathic effect. In addition, the decrease of the nucleic acid and protein levels of VP1 indicated PPE effectively inhibited the proliferation of EV71 in Vero cells. Furthermore, the annexinV-affinity assay suggested that PPE protected host cells from apoptosis. The mechanistic investigations revealed that PPE restrained EV71-induced host-cells apoptosis by AKT and ATM/ATR signaling pathways. In conclusion, these results demonstrate PPE is a hopeful antiviral drug for the infection of EV71.

scholarly journals Antidepressant Sertraline is a Broad-Spectrum Inhibitor of Enteroviruses Targeting Viral Entry Through Neutralization of Endolysosomal Acidification

2021 ◽  
Author(s):  
Kuan-Chi Tseng ◽  
Bang-Yan Hsu ◽  
Pin Ling ◽  
Wen-Wen Lu ◽  
Cheng-Wen Lin ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Viral Entry ◽  
Broad Spectrum ◽  
Culture Media ◽  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Antiviral Effect ◽  
Neurological Complications ◽  
Low Ph

Enterovirus 71 (EV71) is an etiological agent of hand foot and mouth disease and can also cause neurological complications in young children. However, there are no approved drugs to treat EV71 infections. In this study, we conducted an antiviral drug screening by using a Food and Drug Administration (FDA)-approved drug library. We identified five drugs that showed dose-dependent inhibition of viral replication. Sertraline was further characterized because it exhibited the most potent antiviral activity with the highest selectivity index among the five hits. The antiviral activity of sertraline was noted for other EV serotypes. The drug’s antiviral effect is not likely associated with its approved indications as an antidepressant and its mode-of-action as a selective serotonin reuptake inhibitor. The time-of-addition assay revealed that sertraline inhibited an EV71 infection at the entry stage. We also showed that sertraline partitioned into acidic compartments, such as endolysosomes, to neutralize the low pH levels. In agreement with the findings, the antiviral effect of sertraline could be relieved greatly by exposing virus-infected cells to extracellular low-pH culture media. Together, we have identified an FDA-approved antidepressant with the new indication for the broad-spectrum EV inhibition by blocking viral entry through the alkalization of the endolysosomal route.

scholarly journals A nucleobase-binding pocket in a viral RNA-dependent RNA polymerase contributes to elongation complex stability

2019 ◽  
Vol 48 (3) ◽  
pp. 1392-1405 ◽  
Author(s):  
Wei Shi ◽  
Han-Qing Ye ◽  
Cheng-Lin Deng ◽  
Rui Li ◽  
Bo Zhang ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Binding ◽  
Vaccine Development ◽  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Binding Pocket ◽  
Genome Replication ◽  
Elongation Complex ◽  
Rna Dependent Rna Polymerase

Abstract The enterovirus 71 (EV71) 3Dpol is an RNA-dependent RNA polymerase (RdRP) that plays the central role in the viral genome replication, and is an important target in antiviral studies. Here, we report a crystal structure of EV71 3Dpol elongation complex (EC) at 1.8 Å resolution. The structure reveals that the 5′-end guanosine of the downstream RNA template interacts with a fingers domain pocket, with the base sandwiched by H44 and R277 side chains through hydrophobic stacking interactions, and these interactions are still maintained after one in-crystal translocation event induced by nucleotide incorporation, implying that the pocket could regulate the functional properties of the polymerase by interacting with RNA. When mutated, residue R277 showed an impact on virus proliferation in virological studies with residue H44 having a synergistic effect. In vitro biochemical data further suggest that mutations at these two sites affect RNA binding, EC stability, but not polymerase catalytic rate (kcat) and apparent NTP affinity (KM,NTP). We propose that, although rarely captured by crystallography, similar surface pocket interaction with nucleobase may commonly exist in nucleic acid motor enzymes to facilitate their processivity. Potential applications in antiviral drug and vaccine development are also discussed.

scholarly journals A Conserved Inhibitory Mechanism of a Lycorine Derivative against Enterovirus and Hepatitis C Virus

2015 ◽  
Vol 60 (2) ◽  
pp. 913-924 ◽  
Author(s):  
Yu Guo ◽  
Yaxin Wang ◽  
Lin Cao ◽  
Peng Wang ◽  
Jie Qing ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Asia Pacific ◽  
Human Hand ◽  
Content Type ◽  
Viral Proteases ◽  
Causative Agents ◽  
Multiple Strains

ABSTRACTEnterovirus 71 (EV71) (Picornaviridaefamily) and hepatitis C virus (HCV) (Flaviviridaefamily) are the causative agents of human hand, foot, and mouth disease (HFMD) and hepatitis C, resulting in a severe pandemic involving millions of infections in the Asia-Pacific region and worldwide. The great impact of EV71 and HCV on public health highlights the need to further our understanding of the biology of these two viruses and develop effective therapeutic antivirals. Here, we evaluated a total of 32 lycorine derivatives and demonstrated that 1-acetyllycorine suppressed the proliferation of multiple strains of EV71 in various cells. The results of the drug resistance analysis revealed that 1-acetyllycorine targeted a phenylalanine (F76) in EV71 2A protease (2Apro) to stabilize the conformation of a unique zinc finger. Most interestingly, the zinc binding site in EV71 2Aprois the exclusive homolog of HCV NS3 among all viruses. Further analysis revealed that 1-acetyllycorine also inhibits HCV with high efficacy, and the mutation on R118 in HCV NS3, which corresponds to F76 in EV71 2Apro, confers the resistance of HCV to 1-acetyllycorine. These results revealed a conserved mechanism of 1-acetyllycorine against EV71 and HCV through targeting viral proteases. We also documented the significant synergistic anti-EV71 and anti-HCV effects of 1-acetyllycorine with reported inhibitors, supporting potential combination therapy for the treatment of EV71 and HCV infections.

scholarly journals Isolating Compounds that Inhibit EV 71 Virus

2020 ◽  
Author(s):  
Giridhar Murali ◽  
Rishabh Kejriwal ◽  
David Olson ◽  
Lauren Alexandrescu ◽  
Simon White
Keyword(s):  
Protein Purification ◽  
Genetic Information ◽  
Drug Testing ◽  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
E Coli ◽  
Atpase Assay ◽  
Viral Proliferation

Enterovirus 71, or EV 71, is responsible for causing Hand, Foot, and Mouth disease in humans. In particular, it is especially deadly when children and small infants are exposed. The objective of this research paper is to address the possibility of a novel antiviral drug that can be used once infection of EV 71 has occurred. The methods for this research include transformation of E. coli with the genetic information from Enterovirus 71, growth of the E. coli colonies in the lab setting, 2C protein purification, and ATPase assays with drug testing. Of the 364 drugs tested in the ATPase assay, a combination of two of them (Mitrofudil and N6- Benzyladenosine) indicated a stoppage in activity of ATPase, signaling no further activity of the enzyme and viral proliferation.

scholarly journals Design of stable Enterovirus 71 virus like particles (VLPs) as a potential approach to vaccine development

2020 ◽  
Vol 2 (7A) ◽  
Author(s):  
Agnieszka Martyna ◽  
Helen Fox ◽  
Andrew Macadam
Keyword(s):  
Vaccine Development ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Systemic Complications ◽  
Polio Virus ◽  
Virus Like Particles ◽  
Alternative Approach ◽  
Causative Agents ◽  
Inactivated Vaccines ◽  
Single Construct

We have previously described the design of stable and immunogenic polio virus-like particles (VLPs) (Fox, et al. 2017) as an alternative approach to vaccine production. Unlike current polio vaccines, recombinantly-expressed VLP vaccines are non-infectious so would pose no risk of accidental escape from production plants, threatening eradication. To do this we devised a pipeline for the identification of stabilising mutations which could then be combined in a single construct to produce suitable particles; this strategy may have applications for other enterovirus vaccines. Enterovirus 71 (EV71) is one of causative agents of hand, foot and mouth disease which is usually mild but in some cases neurological and systemic complications may occur. Recently there have been several outbreaks with significant mortality in South East Asia as well as increasing numbers of reports of outbreaks in Europe. VLP vaccines might be a useful alternative to inactivated vaccines currently in use or development. EV71, like poliovirus, produces empty particles that are antigenically different from the virion. If, like poliovirus, these empty particles are less immunogenic than the virion, it would be necessary to stabilise them in the native conformation. We are attempting to do this (1) by incorporating modifications that proved successful in the context of poliovirus and (2) by identifying new candidate mutations using an analogous pipeline. Here we will report the characterisation of a range of different modifications that have stabilising and de-stabilising effects on EV71 particles as well as unexpected effects on morphogenesis.

scholarly journals Antidepressant Sertraline Is a Broad-Spectrum Inhibitor of Enteroviruses Targeting Viral Entry through Neutralization of Endolysosomal Acidification

Viruses ◽  
2022 ◽  
Vol 14 (1) ◽  
pp. 109
Author(s):  
Kuan-Chi Tseng ◽  
Bang-Yan Hsu ◽  
Pin Ling ◽  
Wen-Wen Lu ◽  
Cheng-Wen Lin ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Viral Entry ◽  
Broad Spectrum ◽  
Culture Media ◽  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Antiviral Effect ◽  
Neurological Complications ◽  
Low Ph

Enterovirus 71 (EV71) is an etiological agent of hand foot and mouth disease and can also cause neurological complications in young children. However, there are no approved drugs as of yet to treat EV71 infections. In this study, we conducted antiviral drug screening by using a Food and Drug Administration (FDA)-approved drug library. We identified five drugs that showed dose-dependent inhibition of viral replication. Sertraline was further characterized because it exhibited the most potent antiviral activity with the highest selectivity index among the five hits. The antiviral activity of sertraline was noted for other EV serotypes. The drug’s antiviral effect is not likely associated with its approved indications as an antidepressant and its mode-of-action as a selective serotonin reuptake inhibitor. The time-of-addition assay revealed that sertraline inhibited an EV71 infection at the entry stage. We also showed that sertraline partitioned into acidic compartments, such as endolysosomes, to neutralize the low pH levels. In agreement with the findings, the antiviral effect of sertraline could be greatly relieved by exposing virus-infected cells to extracellular low-pH culture media. Ultimately, we have identified a use for an FDA-approved antidepressant in broad-spectrum EV inhibition by blocking viral entry through the alkalization of the endolysosomal route.

scholarly journals Single Neutralizing Monoclonal Antibodies Targeting the VP1 GH Loop of Enterovirus 71 Inhibit both Virus Attachment and Internalization during Viral Entry

2015 ◽  
Vol 89 (23) ◽  
pp. 12084-12095 ◽  
Author(s):  
Zhiqiang Ku ◽  
Xiaohua Ye ◽  
Jinping Shi ◽  
Xiaoli Wang ◽  
Qingwei Liu ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Heparan Sulfate ◽  
Viral Entry ◽  
Neutralizing Antibodies ◽  
Critical Role ◽  
Antiviral Drug ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Independent Manner ◽  
Virus Attachment

ABSTRACTAntibodies play a critical role in immunity against enterovirus 71 (EV71). However, how EV71-specific antibodies neutralize infections remains poorly understood. Here we report the working mechanism for a group of three monoclonal antibodies (MAbs) that potently neutralize EV71. We found that these three MAbs (termed D5, H7, and C4, respectively) recognize the same conserved neutralizing epitope within the VP1 GH loop of EV71. Single MAbs in this group, exemplified by D5, could inhibit EV71 infection in cell cultures at both the pre- and postattachment stages in a cell type-independent manner. Specifically, MAb treatment resulted in the blockade of multiple steps of EV71 entry, including virus attachment, internalization, and subsequent uncoating and RNA release. Furthermore, we show that the D5 and C4 antibodies can interfere with EV71 binding to its key receptors, including heparan sulfate, SCARB2, and PSGL-1, thus providing a possible explanation for the observed multi-inhibitory function of the MAbs. Collectively, our study unravels the mechanism of neutralization by a unique group of anti-EV71 MAbs targeting the conserved VP1 GH loop. The findings should enhance our understanding of MAb-mediated immunity against enterovirus infections and accelerate the development of MAb-based anti-EV71 therapeutic drugs.IMPORTANCEEnterovirus 71 (EV71) is a major causative agent of hand, foot, and mouth disease (HFMD), which has caused significant morbidities and mortalities in young children. Neither a vaccine nor an antiviral drug is available. Neutralizing antibodies are major protective components in EV71 immunity. Here, we unraveled an unusual mechanism of EV71 neutralization by a group of three neutralizing monoclonal antibodies (MAbs). All of these MAbs bound the same conserved epitope located at the VP1 GH loop of EV71. Interestingly, mechanistic studies showed that single antibodies in this MAb group could block EV71 attachment and internalization during the viral entry process and interfere with EV71 binding to heparan sulfate, SCARB2, and PSGL-1 molecules, which are key receptors involved in different steps of EV71 entry. Our findings greatly enhance the understanding of the interplays among EV71, neutralizing antibodies, and host receptors, which in turn should facilitate the development of an MAb-based anti-EV71 therapy.

scholarly journals Structure of the Enterovirus 71 3C Protease in Complex with NK-1.8k and Indications for the Development of Antienterovirus Protease Inhibitor

2017 ◽  
Vol 61 (7) ◽  
Author(s):  
Yaxin Wang ◽  
Lin Cao ◽  
Yangyang Zhai ◽  
Zheng Yin ◽  
Yuna Sun ◽  
...  
Keyword(s):  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Serine Residue ◽  
3C Protease ◽  
The Pacific ◽  
Causative Agents ◽  
Pacific Asia ◽  
Protease Assay ◽  
Shed Light

ABSTRACT Hand-foot-and-mouth disease (HFMD), caused by enterovirus, is a threat to public health worldwide. To date, enterovirus 71 (EV71) has been one of the major causative agents of HFMD in the Pacific-Asia region, and outbreaks with EV71 cause millions of infections. However, no drug is currently available for clinical therapeutics. In our previous works, we developed a set of protease inhibitors (PIs) targeting the EV71 3C protease (3Cpro). Among these are NK-1.8k and NK-1.9k, which have various active groups and high potencies and selectivities. In the study described here, we determined the structures of the PI NK-1.8k in complex with wild-type (WT) and drug-resistant EV71 3Cpro. Comparison of these structures with the structure of unliganded EV71 3Cpro and its complex with AG7088 indicated that the mutation of N69 to a serine residue destabilized the S2 pocket. Thus, the mutation influenced the cleavage activity of EV71 3Cpro and the inhibitory activity of NK-1.8k in an in vitro protease assay and highlighted that site 69 is an additional key site for PI design. More information for the optimization of the P1′ to P4 groups of PIs was also obtained from these structures. Together with the results of our previous works, these in-depth results elucidate the inhibitory mechanism of PIs and shed light to develop PIs for the clinical treatment of infections caused by EV71 and other enteroviruses.

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