Human transbodies that interfere with the functions of Ebola virus VP35 protein in genome replication and transcription and innate immune antagonism

Watee Seesuay,Surasak Jittavisutthikul,Nawannaporn Sae-lim,Nitat Sookrung,Yuwaporn Sakolvaree,Wanpen Chaicumpa

doi:10.1038/s41426-018-0031-3

Watee Seesuay, Surasak Jittavisutthikul + Show 4 more

Open Access

https://doi.org/10.1038/s41426-018-0031-3

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Abstract

Small molecular inhibitors and passive immunization against Ebola virus disease (EVD) have been tested in animal models, including rodents and non-human primates, as well as in clinical trials. Nevertheless, there is currently no Food and Drug Administration (FDA)-approved therapy, and alternative strategies must be pursued. The aim of this study was to produce cell-penetrable human single-chain antibodies (transbodies) that are able to interfere with the activities of interferon inhibitory domain (IID) of the VP35 protein, a multifunctional virulence factor of Ebola virus (EBOV). We speculated that effective VP35-IID-specific transbodies could inspire further studies to identify an alternative to conventional antibody therapies. Phage display technology was used to generate Escherichia coli-derived human single-chain antibodies (HuscFvs) that bind to IID. HuscFvs were linked to nona-arginine (R9) to make them cell penetrable. Transbodies of transformed E. coli clones 13 and 3, which were predicted to interact with first basic patch residues (R9-HuscFv13), central basic patch, and end-cap residues (R9-HuscFv3), effectively inhibited EBOV minigenome activity. Transbodies of E. coli clones 3 and 8 antagonized VP35-mediated interferon suppression in VP35-transduced cells. We postulate that these transbodies formed an interface contact with the IID central basic patch, end-cap, and/or residues that are important for IID multimeric formation for dsRNA binding. These transbodies should be evaluated further in vitro using authentic EBOV and in vivo in animal models of EVD before their therapeutic/prophylactic effectiveness is clinically evaluated.

Highlights

VP35 protein is a multifunctional virulence factor forEbola virus (EBOV) replication[1]
Given that VP35 is associated with several pivotal activities in the EBOV infectious cycle[1], the aim of this study was to generate cell-penetrable human scFvs (R9-HuscFvs) or transbodies that can effectively interfere with VP35-inhibitory domain (IID) functions
Passive immunization and previously tested immunotherapy could not interfere with the replication of virus that has gained entry into host cells

Summary

Introduction

VP35 protein is a multifunctional virulence factor forEbola virus (EBOV) replication[1]. One of the functions of this protein is its polymerase co-factor activity that VP35 protein and other EBOV proteins, such as NP (nucleoprotein), VP30 (transcription factor), and L (RNAdependent RNA polymerase) and viral RNA, form the. The EBOV VP35 comprises 340 residues that contain the N-terminal oligomerization domain (NOD) 1–220) and C-terminal portion that has IFN antagonistic activity (termed the IFN-inhibitory domain, IID)[12–14]. NOD, which facilitates the interferon-antagonist activity of IID15. Residues 20–48 bind NP and regulate NP assembly on viral RNA to facilitate RNA synthesis[16,17]. The IID consists of two subdomains: an N-terminal α-helical subdomain (residues 221–283) that contains four alpha helices (α1–α4) and a C-terminal β-sheet subdomain (residues 294–340) that comprises four antiparallel strands (β1–β4), a small α5, and a type II polyproline helix[13]. Within the IID, there are two conserved basic patches and additional border basic residues that play different but cooperative roles in EBOV replication[19].

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