Antiviral Potential of a Novel Compound CW-33 against Enterovirus A71 via Inhibition of Viral 2A Protease.

Ching-Ying Wang,Su-Hua Huang,Yuan-Shiun Chang,I-Chieh Chen,Szu-Hao Kung,Chao-Hsien Chen,Mann-Jen Hour,An-Cheng Huang,Jin-Cherng Lien,Cheng-Wen Lin

doi:10.3390/v7062764

Abstract

Enterovirus A71 (EV-A71) in the Picornaviridae family causes hand-foot-and-mouth disease, aseptic meningitis, severe central nervous system disease, even death. EV-A71 2A protease cleaves Type I interferon (IFN)-α/β receptor 1 (IFNAR1) to block IFN-induced Jak/STAT signaling. This study investigated anti-EV-A7l activity and synergistic mechanism(s) of a novel furoquinoline alkaloid compound CW-33 alone and in combination with IFN-β. Anti-EV-A71 activities of CW-33 alone and in combination with IFN-β were evaluated by inhibitory assays of virus-induced apoptosis, plaque formation, and virus yield. CW-33 showed antiviral activities with an IC50 of near 200 μM in EV-A71 plaque reduction and virus yield inhibition assays. While, anti-EV-A71 activities of CW-33 combined with 100 U/mL IFN-β exhibited a synergistic potency with an IC50 of approximate 1 μM in plaque reduction and virus yield inhibition assays. Molecular docking revealed CW-33 binding to EV-A71 2A protease active sites, correlating with an inhibitory effect of CW33 on in vitro enzymatic activity of recombinant 2A protease (IC50 = 53.1 μM). Western blotting demonstrated CW-33 specifically inhibiting 2A protease-mediated cleavage of IFNAR1. CW-33 also recovered Type I IFN-induced Tyk2 and STAT1 phosphorylation as well as 2′,5′-OAS upregulation in EV-A71 infected cells. The results demonstrated CW-33 inhibiting viral 2A protease activity to reduce Type I IFN antagonism of EV-A71. Therefore, CW-33 combined with a low-dose of Type I IFN could be applied in developing alternative approaches to treat EV-A71 infection.

Highlights

Enterovirus A71 (EV-A71) belongs to the “Enterovirus A” species, genus Enterovirus in the Picornaviridae family, comprising an icosahedral capsid and single positive-strand RNA genome of approximately 7400 nucleotides [1]
This study showed that IFN-β was less effective against EV-A71 (Figure 3C,D), consistent with
This study showed that IFN-β was less effective against EV-A71 (Figs. 3C and 3D), consistent with prior reports in which EV-A71 antagonized antiviral actions of Type I IFN [10,27,29]

Summary

Introduction

Enterovirus A71 (EV-A71) belongs to the “Enterovirus A” species, genus Enterovirus in the Picornaviridae family, comprising an icosahedral capsid and single positive-strand RNA genome of approximately 7400 nucleotides [1]. The genus Enterovirus, one of most common genera within the family Picornaviridae, comprises 72 serotypes: e.g., poliovirus, Coxsackie A virus (CVA), Coxsackie. B virus (CVB), echovirus, EV-A71 [2,3,4]. EV-A71 was first isolated and characterized from cases of neurological disease in California as of 1969 [5], usually infecting children by direct contact with virus shed from the upper respiratory or gastrointestinal tract. The genome contains a single long open reading frame (ORF). The ORF encodes a polyprotein precursor cleaved into many consecutive functional parts by viral 2A and 3C proteases, such as P1, P2 and P3 fragments firstly cleaved by 2A protease. The P1 fragment yields structural proteins VP1, VP2, VP3 and VP4, while P2 and P3 fragments divide into non-structural 2A, 2B, 2C, 3A, 3B, 3C and 3D

Methods

Results

Conclusion