High Throughput Screening Targeting the Dengue NS3-NS5 Interface Identifies Antivirals against Dengue, Zika and West Nile Viruses.

Sundy N. Y. Yang,David A. Jans,Johanna E. Fraser,Kylie M. Wagstaff,Chunxiao Wang,Belinda Maher

doi:10.3390/cells11040730

Abstract

Dengue virus (DENV) threatens almost 70% of the world’s population, with no effective therapeutic currently available and controversy surrounding the one approved vaccine. A key factor in dengue viral replication is the interaction between DENV nonstructural proteins (NS) 5 and 3 (NS3) in the infected cell. Here, we perform a proof-of-principle high-throughput screen to identify compounds targeting the NS5-NS3 binding interface. We use a range of approaches to show for the first time that two small molecules–repurposed drugs I-OMe tyrphostin AG538 (I-OMe-AG238) and suramin hexasodium (SHS)–inhibit NS5-NS3 binding at low μM concentration through direct binding to NS5 that impacts thermostability. Importantly, both have strong antiviral activity at low μM concentrations against not only DENV-2, but also Zika virus (ZIKV) and West Nile virus (WNV). This work highlights the NS5-NS3 binding interface as a viable target for the development of anti-flaviviral therapeutics.

Highlights

Dengue fever is an acute, mosquito-transmitted viral disease caused by the flavivirus dengue virus (DENV) that threatens more than two-thirds of the world population [1]
We previously showed that DENV NS5 traffics into and out of the cell nucleus during infection through interaction with specific members of the host importin (IMP) superfamily of nuclear transporters [5,13], and that this is essential to the DENV infectious cycle, whereby prevention of interaction through mutation leads to viral attenuation [14] and various small molecule inhibitors of the interaction reduce infectious virus production [13,15,16,17,18,19]
Complex formation by NS5 and NS3 is essential for DENV replication [10,11,12]; we speculated that disruption of this protein–protein interaction may be a viable antiviral strategy

Summary

Introduction

Dengue fever is an acute, mosquito-transmitted viral disease caused by the flavivirus dengue virus (DENV) that threatens more than two-thirds of the world population [1]. Despite almost 100 million symptomatic cases annually, with 500,000 hospitalisations and ca. There is a compelling case to devise effective new therapies. Related to Zika virus (ZIKV), West Nile virus (WNV), and Japanese encephalitis virus (JEV) [4], DENV possesses a positive-sense, single-stranded RNA genome that encodes three structural and seven nonstructural (NS) proteins. Conserved among the four DENV serotypes (DENV 1–4) as well as ZIKV/WNV/JEV [4,5], the dual function NS3 serine protease/RNA helicase and NS5 methyltransferase/RNA-dependent RNA polymerase (RdRp) proteins provide the key enzymatic activities required to synthesise the viral RNA genome. Specific mutation to prevent NS3 binding to NS5 abolishes infectious DENV virus production [11], consistent with an essential role of the NS5-NS3 interaction in DENV replication [10,11,12] and highlighting the potential of the DENV NS5-NS3 interface as an antiviral strategy

Methods

Results

Conclusion