Filoviruses infect humans, non-human primates, bats and other mammals. Several filoviruses cause hemorrhagic fever diseases in humans, including the recent Ebola virus (EBOV) outbreak in western Africa. Promising interventions are on the horizon, but the need for new strategies is highlighted by the continued absence of an effective and widely-available vaccine or antiviral drug regimen, by the difficulty of assuring patient recovery even in modern hospital settings, and by the continuing risk of the emergence and rapid international spread of new diseases. This project will reveal new opportunities to target glycoproteins (GPs) by identifying and characterizing nucleic acid aptamers that recognize and inhibit filoviral GPs. Filoviruses display a trimeric GP on their surface membrane, which they acquire from infected cells during assembly. GPs are the binding targets for both neutralizing and non-neutralizing antibodies (Ab), and the molecular mechanisms of neutralization are beginning to emerge. However, the non-conserved and heavily- glycosylated mucin-like domain (MLD) blocks Ab access to much of the GP surface, including the NCP1 binding site in the case of EBOV GP (?GPEBOV?). Smaller ligands such as aptamers could potentially reach and block neutralization surfaces that are inaccessible to antibodies, and they could aid identification of new neutralization sites. Our long-term objective is to develop aptamers as tools for identifying new neutralizing epitopes and for dissecting molecular and cellular events in viral pathogenesis. The current proposal will establish feasibility of that approach and identify initial leads for mechanistic studies. It capitalizes on complementary expertise in virology and RNA biochemistry in the participating labs; on our recent achievements in advanced aptamer selection and informatics methods; on our recent critical insights into the mechanisms and versatility of glycoprotein incorporation during viral assembly; and on the addition of new collaborators with EBOV expertise.
The first Aim will identify aptamers that neutralize GP-pseudotyped and infectious virus by recognizing epitopes that are shared between (or unique to) GPEBOV and GPMARV.
The second Aim will identify and evaluate aptamers that target known neutralizing epitopes.
Health relevance: This study may lead to better understanding of strategies for blocking replication of Ebola virus and related viruses: 1) it could identify new sites on the viral glycoprotein that neutralize viral infectivity and that could be targeted in vaccination strategies, 2) reagents developed in this study could identify neutralization surfaces that are not normally accessible to antibodies but that could be targeted by small molecule drugs, 3) the mode of action of those reagents could illuminate molecular mechanism of viral entry and the identification of new strategies to block the virus. Also, the innovative experimental approaches could be generalized for future studies to target other serious emerging viruses such as Lassa, Nipah, and Hendra.
| Porciani, David; Cardwell, Leah N; Tawiah, Kwaku D et al. (2018) Modular cell-internalizing aptamer nanostructure enables targeted delivery of large functional RNAs in cancer cell lines. Nat Commun 9:2283 |
