Approaches to demonstrating the effectiveness of filovirus vaccines: Lessons from Ebola and COVID-19.

Zaire ebolavirus (EBOV), Sudan ebolavirus (SUDV) and Marburg virus (MARV), are members of the Filoviridae family that can cause severe disease and death in humans and animals. The reemergence of Ebola, Sudan and Marburg virus disease highlight the need for continued availability of safe and effectives vaccines as well as development of new vaccines. While randomized controlled trials using disease endpoints provide the most robust assessment of vaccine effectiveness, challenges to this approach include the unpredictable size, location, occurrence and duration of filovirus disease outbreaks. Thus, other approaches to demonstrating vaccine effectiveness have been considered. These approaches are discussed using examples of preventive vaccines against other infectious diseases. In addition, this article proposes a clinical immunobridging strategy using licensed EBOV vaccines as comparators for demonstrating the effectiveness of filovirus vaccine candidates that are based on the same licensed vaccine platform technology.

Inactivation of Ebola Virus and SARS-CoV-2 in Cell Culture Supernatants and Cell Pellets by Gamma Irradiation.

Viral pathogens with the potential to cause widespread disruption to human health and society continue to emerge or re-emerge around the world. Research on such viruses often involves high biocontainment laboratories (BSL3 or BSL4), but the development of diagnostics, vaccines and therapeutics often uses assays that are best performed at lower biocontainment. Reliable inactivation is necessary to allow removal of materials to these spaces and to ensure personnel safety. Here, we validate the use of gamma irradiation to inactivate culture supernatants and pellets of cells infected with a representative member of the Filovirus and Coronavirus families. We show that supernatants and cell pellets containing SARS-CoV-2 are readily inactivated with 1.9 MRad, while Ebola virus requires higher doses of 2.6 MRad for supernatants and 3.8 MRad for pellets. While these doses of radiation inactivate viruses, proinflammatory cytokines that are common markers of virus infection are still detected with low losses. The doses required for virus inactivation of supernatants are in line with previously reported values, but the inactivation of cell pellets has not been previously reported and enables new approaches for analysis of protein-based host responses to infection.

ß-arrestins and G protein-coupled receptor kinases in viral entry: A graphical review.

Viruses rely on host-cell machinery in order to invade host cells and carry out a successful infection. G-protein coupled receptor (GPCR)-mediated signaling pathways are master regulators of cellular physiological processing and are an attractive target for viruses to rewire cells during infection. In particular, the GPCR-associated scaffolding proteins ß-arrestins and GPCR signaling effectors G-protein receptor kinases (GRKs) have been identified as key cellular factors that mediate viral entry and orchestrate signaling pathways that reprogram cells for viral replication. Interestingly, a broad range of viruses have been identified to activate and/or require GPCR-mediated pathways for infection, including polyomaviruses, flaviviruses, influenza virus, and SARS-CoV-2, demonstrating that these viruses may have conserved mechanisms of host-cell invasion. Thus, GPCR-mediated pathways highlight an attractive target for the development of broad antiviral therapies.

The Evolution of Medical Countermeasures for Ebola Virus Disease: Lessons Learned and Next Steps.

The Ebola virus disease outbreak that occurred in Western Africa from 2013-2016, and subsequent smaller but increasingly frequent outbreaks of Ebola virus disease in recent years, spurred an unprecedented effort to develop and deploy effective vaccines, therapeutics, and diagnostics. This effort led to the U.S. regulatory approval of a diagnostic test, two vaccines, and two therapeutics for Ebola virus disease indications. Moreover, the establishment of fieldable diagnostic tests improved the speed with which patients can be diagnosed and public health resources mobilized. The United States government has played and continues to play a key role in funding and coordinating these medical countermeasure efforts. Here, we describe the coordinated U.S. government response to develop medical countermeasures for Ebola virus disease and we identify lessons learned that may improve future efforts to develop and deploy effective countermeasures against other filoviruses, such as Sudan virus and Marburg virus.

Broad-Spectrum In Vitro Antiviral Activity of ODBG-P-RVn: An Orally-Available, Lipid-Modified Monophosphate Prodrug of Remdesivir Parent Nucleoside (GS-441524).

The necessity for intravenous administration of remdesivir confines its utility for treatment of coronavirus disease 2019 (COVID-19) to hospitalized patients. We evaluated the broad-spectrum antiviral activity of ODBG-P-RVn, an orally available, lipid-modified monophosphate prodrug of the remdesivir parent nucleoside (GS-441524), against viruses that cause diseases of human public health concern, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). ODBG-P-RVn showed 20-fold greater antiviral activity than GS-441524 and had activity nearly equivalent to that of remdesivir in primary-like human small airway epithelial cells. Our results warrant in vivo efficacy evaluation of ODBG-P-RVn. IMPORTANCE While remdesivir remains one of the few drugs approved by the FDA to treat coronavirus disease 2019 (COVID-19), its intravenous route of administration limits its use to hospital settings. Optimizing the stability and absorption of remdesivir may lead to a more accessible and clinically potent therapeutic. Here, we describe an orally available lipid-modified version of remdesivir with activity nearly equivalent to that of remdesivir against emerging viruses that cause significant disease, including Ebola and Nipah viruses. Our work highlights the importance of such modifications to optimize drug delivery to relevant and appropriate human tissues that are most affected by such diseases.

Review: Insights on Current FDA-Approved Monoclonal Antibodies Against Ebola Virus Infection.

The unprecedented 2013-2016 West Africa Ebola outbreak accelerated several medical countermeasures (MCMs) against Ebola virus disease (EVD). Several investigational products (IPs) were used throughout the outbreak but were not conclusive for efficacy results. Only the Randomized Controlled Trial (RCT) on ZMapp was promising but inconclusive. More recently, during the second-largest Ebola outbreak in North Kivu and Ituri provinces, Democratic Republic of the Congo (DRC), four IPs, including one small molecule (Remdesivir), two monoclonal antibody (mAb) cocktails (ZMapp and REGN-EB3) and a single mAb (mAb114), were evaluated in an RCT, the Pamoja Tulinde Maisha (PALM) study. Two products (REGN-EB3 and mAb114) demonstrated efficacy as compared to the control arm, ZMapp. There were remarkably few side effects recorded in the trial. The FDA approved both medications in this scientifically sound study, marking a watershed moment in the field of EVD therapy. These products can be produced relatively inexpensively and can be stockpiled. The administration of mAbs in EVD patients appears to be safe and effective, while several critical knowledge gaps remain; the impact of early administration of Ebola-specific mAbs on developing a robust immune response for future Ebola virus exposure is unknown. The viral mutation escape, leading to resistance, presents a potential limitation for single mAb therapy; further improvements need to be explored. Understanding the contribution of Fc-mediated antibody functions such as antibody-dependent cellular cytotoxicity (ADCC) of those approved mAbs is still critical. The potential merit of combination therapy and post-exposure prophylaxis (PEP) need to be demonstrated. Furthermore, the PALM trial has accounted for 30% of mortality despite the administration of specific treatments. The putative role of EBOV soluble Glycoprotein (sGP) as a decoy to the immune system, the virus persistence, and relapses might be investigated for treatment failure. The development of pan-filovirus or pan-species mAbs remains essential for protection. The interaction between FDA-approved mAbs and vaccines remains unclear and needs to be investigated. In this review, we summarize the efficacy and safety results of the PALM study and review current research questions for the further development of mAbs in pre-exposure or emergency post-exposure use.

Ebola Virus Glycoprotein Domains Associated with Protective Efficacy.

Ebola virus (EBOV) is the cause of sporadic outbreaks of human hemorrhagic disease in Africa, and the best-characterized virus in the filovirus family. The West African epidemic accelerated the clinical development of vaccines and therapeutics, leading to licensure of vaccines and antibody-based therapeutics for human use in recent years. The most widely used vaccine is based on vesicular stomatitis virus (VSV) expressing the EBOV glycoprotein (GP) (VSV-EBOV). Due to its favorable immune cell targeting, this vaccine has also been used as a base vector for the development of second generation VSV-based vaccines against Influenza, Nipah, and Zika viruses. However, in these situations, it may be beneficial if the immunogenicity against EBOV GP is minimized to induce a better protective immune response against the other foreign immunogen. Here, we analyzed if EBOV GP can be truncated to be less immunogenic, yet still able to drive replication of the vaccine vector. We found that the EBOV GP glycan cap and the mucin-like domain are both dispensable for VSV-EBOV replication. The glycan cap, however, appears critical for mediating a protective immune response against lethal EBOV challenge in mice.

The SKI complex is a broad-spectrum, host-directed antiviral drug target for coronaviruses, influenza, and filoviruses.

The SARS-CoV-2 pandemic has made it clear that we have a desperate need for antivirals. We present work that the mammalian SKI complex is a broad-spectrum, host-directed, antiviral drug target. Yeast suppressor screening was utilized to find a functional genetic interaction between proteins from influenza A virus (IAV) and Middle East respiratory syndrome coronavirus (MERS-CoV) with eukaryotic proteins that may be potential host factors involved in replication. This screening identified the SKI complex as a potential host factor for both viruses. In mammalian systems siRNA-mediated knockdown of SKI genes inhibited replication of IAV and MERS-CoV. In silico modeling and database screening identified a binding pocket on the SKI complex and compounds predicted to bind. Experimental assays of those compounds identified three chemical structures that were antiviral against IAV and MERS-CoV along with the filoviruses Ebola and Marburg and two further coronaviruses, SARS-CoV and SARS-CoV-2. The mechanism of antiviral activity is through inhibition of viral RNA production. This work defines the mammalian SKI complex as a broad-spectrum antiviral drug target and identifies lead compounds for further development.