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.

scholarly journals Eupafolin and Ethyl Acetate Fraction ofKalanchoe gracilisStem Extract Show Potent Antiviral Activities against Enterovirus 71 and Coxsackievirus A16

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Ching-Ying Wang ◽  
Shun-Chueh Huang ◽  
Zhen-Rung Lai ◽  
Yu-Ling Ho ◽  
Yu-Jen Jou ◽  
...  
Keyword(s):  
Ethyl Acetate ◽  
Folk Medicine ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Ethyl Acetate Fraction ◽  
Coxsackievirus A16 ◽  
Aqueous Fraction ◽  
Antiviral Activities ◽  
Clinical Potential

Enterovirus 71 (EV71) and coxsackievirus A16 (CoxA16) are main pathogens of hand-foot-and-mouth disease, occasionally causing aseptic meningitis and encephalitis in tropical and subtropical regions.Kalanchoe gracilis,Da-Huan-Hun, is a Chinese folk medicine for treating pain and inflammation, exhibiting antioxidant and anti-inflammatory activities. Our prior report (2012) citedK. gracilisleaf extract as moderately active against EV71 and CoxA16. This study further rates antienteroviral potential ofK. gracilisstem (KGS) extract to identify potent antiviral fractions and components. The extract moderately inhibits viral cytopathicity and virus yield, as well asin vitroreplication of EV71 (IC50= 75.18 μg/mL) and CoxA16 (IC50= 81.41 μg/mL). Ethyl acetate (EA) fraction of KGS extract showed greater antiviral activity than that ofn-butanol or aqueous fraction: IC50values of 4.21 μg/mL against EV71 and 9.08 μg/mL against CoxA16. HPLC analysis, UV-Vis absorption spectroscopy, and plaque reduction assay indicate that eupafolin is a vital component of EA fraction showing potent activity against EV71 (IC50= 1.39 μM) and CoxA16 (IC50= 5.24 μM). Eupafolin specifically lessened virus-induced upregulation of IL-6 and RANTES by inhibiting virus-induced ERK1/2, AP-1, and STAT3 signals. Anti-enteroviral potency of KGS EA fraction and eupafolin shows the clinical potential against EV71 and CoxA16 infection.

scholarly journals Luteoloside Acts as 3C Protease Inhibitor of Enterovirus 71 In Vitro

PLoS ONE ◽  
2016 ◽  
Vol 11 (2) ◽  
pp. e0148693 ◽  
Author(s):  
Zeyu Cao ◽  
Yue Ding ◽  
Zhipeng Ke ◽  
Liang Cao ◽  
Na Li ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Enterovirus 71 ◽  
3C Protease

scholarly journals Synthesis and Broad Antiviral Activity of Novel 2-aryl-isoindolin-1-ones towards Diverse Enterovirus A71 Clinical Isolates

Molecules ◽  
2019 ◽  
Vol 24 (5) ◽  
pp. 985 ◽  
Author(s):  
Yixuan Wang ◽  
Huiqiang Wang ◽  
Xinbei Jiang ◽  
Zhi Jiang ◽  
Tingting Guo ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Viral Disease ◽  
Clinical Isolates ◽  
Dependent Manner ◽  
Potent Antiviral Activity ◽  
Sar Studies ◽  
Enterovirus A71

Enterovirus 71 (EV-A71) is the main causative pathogen of childhood hand, foot and mouth disease. Effective medicine is currently unavailable for the treatment of this viral disease. Using the fragment-hopping strategy, a series of 2-aryl-isoindolin-1-one compounds were designed, synthesized and investigated for their in vitro antiviral activity towards multiple EV-A71 clinical isolates (H, BrCr, Shenzhen98, Jiangsu52) in Vero cell culture in this study. The structure–activity relationship (SAR) studies identified 2-phenyl-isoindolin-1-ones as a new potent chemotype with potent antiviral activity against EV-A71. Ten out of the 24 tested compounds showed significant antiviral activity (EC50 < 10 µM) towards four EV-A71 strains. Compounds A3 and A4 exhibited broad and potent antiviral activity with the 50% effective concentration (EC50) values in the range of 1.23–1.76 μM. Moreover, the selectivity indices of A3 and A4 were significantly higher than those of the reference compound, pirodavir. The western blotting experiment indicated that the viral VP1 was significantly decreased at both the protein and RNA level in a dose-dependent manner following treatment with compound A3. Moreover, compound A3 inhibited the viral replication by acting on the virus entry stage. In summary, this study led to the discovery of 2-aryl-isoindolin-1-ones as a promising scaffold with potent anti-EV-A71 activities, which deserves further in-depth studies.

scholarly journals Temperature-sensitive mutants of enterovirus 71 show attenuation in cynomolgus monkeys

2005 ◽  
Vol 86 (5) ◽  
pp. 1391-1401 ◽  
Author(s):  
Minetaro Arita ◽  
Hiroyuki Shimizu ◽  
Noriyo Nagata ◽  
Yasushi Ami ◽  
Yuriko Suzaki ◽  
...  
Keyword(s):  
Cultured Cells ◽  
Virulent Strain ◽  
Infectious Clone ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Infection Model ◽  
Temperature Sensitive ◽  
Cynomolgus Monkeys

Enterovirus 71 (EV71) is one of the major causative agents of hand, foot and mouth disease and is sometimes associated with serious neurological disorders. In this study, an attempt was made to identify molecular determinants of EV71 attenuation of neurovirulence in a monkey infection model. An infectious cDNA clone of the virulent strain of EV71 prototype BrCr was constructed; temperature-sensitive (ts) mutations of an attenuated strain of EV71 or of poliovirus (PV) Sabin vaccine strains were then introduced into the infectious clone. In vitro and in vivo phenotypes of the parental and mutant viruses were analysed in cultured cells and in cynomolgus monkeys, respectively. Mutations in 3D polymerase (3Dpol) and in the 3′ non-translated region (NTR), corresponding to ts determinants of Sabin 1, conferred distinct temperature sensitivity to EV71. An EV71 mutant [EV71(S1-3′)] carrying mutations in the 5′ NTR, 3Dpol and in the 3′ NTR showed attenuated neurovirulence, resulting in limited spread of virus in the central nervous system of monkeys. These results indicate that EV71 and PV1 share common genetic determinants of neurovirulence in monkeys, despite the distinct properties in their original pathogenesis.

scholarly journals Antiviral Efficacy of Flavonoids against Enterovirus 71 Infection in Vitro and in Newborn Mice

Viruses ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 625 ◽  
Author(s):  
Wenwen Dai ◽  
Jinpeng Bi ◽  
Fang Li ◽  
Shuai Wang ◽  
Xinyu Huang ◽  
...  
Keyword(s):  
Protective Efficacy ◽  
Lethal Dose ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Neurological Complications ◽  
Ev71 Infection ◽  
Newborn Mice ◽  
Reported Data

Enterovirus 71 (EV71) infection is known to cause hand, foot, and mouth disease (HFMD), which is associated with neurological complications; however, there is currently no effective treatment for this infection. Flavonoids are a large group of naturally occurring compounds with multiple bioactivities, and the inhibitory effects of several flavonoids against EV71 have been studied in cell cultures; however, to date, there are no reported data on their effects in animal models. In this study, we confirmed the in vitro activities of eight flavonoids against EV71 infection, based on the inhibition of cytopathic effects. Moreover, these flavonoids were found to reduce viral genomic RNA replication and protein synthesis. We further demonstrated the protective efficacy of these flavonoids in newborn mice challenged with a lethal dose of EV71. Apigenin, luteolin, kaempferol, formononetin, and penduletin conferred survival protection of 88.89%, 91.67%, 88.89%, 75%, and 66.67%, respectively, from the lethal EV71 challenge. In addition, isorhamnetin provided the highest mice survival protection of 100% at a dose of 10 mg/kg. This study, to the best of our knowledge, is the first to evaluate the in vivo anti-EV7l activities of multiple flavonoids, and we accordingly identified flavonoids as potential leading compounds for anti-EV71 drug development.

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 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 Natural Phytochemicals, Luteolin and Isoginkgetin, Inhibit 3C Protease and Infection of FMDV, In Silico and In Vitro

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2118
Author(s):  
Sirin Theerawatanasirikul ◽  
Nattarat Thangthamniyom ◽  
Chih-Jung Kuo ◽  
Ploypailin Semkum ◽  
Nantawan Phecharat ◽  
...  
Keyword(s):  
Viral Entry ◽  
Antiviral Agents ◽  
Resonance Energy ◽  
Foot And Mouth Disease ◽  
Binding Pocket ◽  
Economic Losses ◽  
3C Protease ◽  
Compound Libraries ◽  
Mouth Disease Virus

Foot-and-mouth-disease virus (FMDV) is a picornavirus that causes a highly contagious disease of cloven-hoofed animals resulting in economic losses worldwide. The 3C protease (3Cpro) is the main protease essential in the picornavirus life cycle, which is an attractive antiviral target. Here, we used computer-aided virtual screening to filter potential anti-FMDV agents from the natural phytochemical compound libraries. The top 23 filtered compounds were examined for anti-FMDV activities by a cell-based assay, two of which possessed antiviral effects. In the viral and post-viral entry experiments, luteolin and isoginkgetin could significantly block FMDV growth with low 50% effective concentrations (EC50). Moreover, these flavonoids could reduce the viral load as determined by RT-qPCR. However, their prophylactic activities were less effective. Both the cell-based and the fluorescence resonance energy transfer (FRET)-based protease assays confirmed that isoginkgetin was a potent FMDV 3Cpro inhibitor with a 50% inhibition concentration (IC50) of 39.03 ± 0.05 and 65.3 ± 1.7 μM, respectively, whereas luteolin was less effective. Analyses of the protein–ligand interactions revealed that both compounds fit in the substrate-binding pocket and reacted to the key enzymatic residues of the 3Cpro. Our findings suggested that luteolin and isoginkgetin are promising antiviral agents for FMDV and other picornaviruses.

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.

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