Enhancement of Ebola virus infection by seminal amyloid fibrils

The 2014 western Africa Ebola virus (EBOV) epidemic was unprecedented in magnitude, infecting over 28,000 and causing over 11,000 deaths. During this outbreak, multiple instances of EBOV sexual transmission were reported, including cases where the infectious individual had recovered from EBOV disease months before transmission. Potential human host factors in EBOV sexual transmission remain unstudied. Several basic seminal amyloids, most notably semen-derived enhancer of viral infection (SEVI), enhance in vitro infection by HIV and several other viruses. To test the ability of these peptides to enhance EBOV infection, viruses bearing the EBOV glycoprotein (EboGP) were preincubated with physiological concentrations of SEVI before infection of physiologically relevant cell lines and primary cells. Preincubation with SEVI significantly increased EboGP-mediated infectivity and replication in epithelium- and monocyte-derived cell lines. This enhancement was dependent upon amyloidogenesis and positive charge, and infection results were observed with both viruses carrying EboGP and authentic EBOV as well as with semen. SEVI enhanced binding of virus to cells and markedly increased its subsequent internalization. SEVI also stimulated uptake of a fluid phase marker by macropinocytosis, a critical mechanism by which cells internalize EBOV. We report a previously unrecognized ability of SEVI and semen to significantly alter viral physical properties critical for transmissibility by increasing the stability of EboGP-bearing recombinant viruses during incubation at elevated temperature and providing resistance to desiccation. Given the potential for EBOV sexual transmission to spark new transmission chains, these findings represent an important interrogation of factors potentially important for this EBOV transmission route.

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Inflammatory and Humoral Immune Response during Ebola Virus Infection in Survivor and Fatal Cases Occurred in Sierra Leone during the 2014–2016 Outbreak in West Africa

Viruses ◽

10.3390/v11040373 ◽

2019 ◽

Vol 11 (4) ◽

pp. 373 ◽

Cited By ~ 8

Author(s):

Francesca Colavita ◽

Mirella Biava ◽

Concetta Castilletti ◽

Simone Lanini ◽

Rossella Miccio ◽

...

Keyword(s):

Immune Response ◽

Inflammatory Response ◽

Sierra Leone ◽

Ebola Virus ◽

Virus Disease ◽

Late Phase ◽

Treatment Center ◽

Humoral Immune ◽

Western Africa ◽

Ebov Infection

Ebola virus (EBOV) infection is characterized by an excessive inflammatory response, a loss of lymphocytes and a general paralysis of the immune system, however pathophysiological mechanisms are not fully understood. In a cohort of 23 fatal and 21 survivors of ebola virus disease (EVD) cases admitted to the Emergency Ebola-Treatment-Center in Goderich (Freetown, Sierra Leone) during the 2014 to 2016 EBOV epidemic in Western Africa, we analyzed the pathway-focused gene expression profile of secreted proteins involved in the immune response and the levels of specific anti-EBOV IgM and IgG from the time of admission till discharge or death. We observed a dysregulated inflammatory response in fatal patients as compared to survivors, mainly consisting of the upregulation of inflammatory mediators, whose extent directly correlated with viremia levels. The upregulation persisted and intensified during the late phase of infection. Relevant differences were also found in humoral immunity, as an earlier and more robust EBOV antibody response was observed in survivor patients.

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Quantitative Expression and Virus Transmission Analysis of DC-SIGN on Monocyte-Derived Dendritic Cells

Journal of Virology ◽

10.1128/jvi.76.18.9135-9142.2002 ◽

2002 ◽

Vol 76 (18) ◽

pp. 9135-9142 ◽

Cited By ~ 86

Author(s):

Frédéric Baribaud ◽

Stefan Pöhlmann ◽

George Leslie ◽

Frank Mortari ◽

Robert W. Doms

Keyword(s):

Dendritic Cells ◽

Cell Lines ◽

Ebola Virus ◽

Virus Transmission ◽

Virus Binding ◽

Expression Levels ◽

Quantitative Expression ◽

Immunodeficiency Virus

ABSTRACT The C-type lectins DC-SIGN and DC-SIGNR efficiently bind human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) strains and can transmit bound virus to adjacent CD4-positive cells. DC-SIGN also binds efficiently to the Ebola virus glycoprotein, enhancing Ebola virus infection. DC-SIGN is thought to be responsible for the ability of dendritic cells (DCs) to capture HIV and transmit it to T cells, thus promoting HIV dissemination in vitro and perhaps in vivo as well. To investigate DC-SIGN function and expression levels on DCs, we characterized a panel of monoclonal antibodies (MAbs) directed against the carbohydrate recognition domain of DC-SIGN. Using quantitative fluorescence-activated cell sorter technology, we found that DC-SIGN is highly expressed on immature monocyte-derived DCs, with at least 100,000 copies and often in excess of 250,000 copies per DC. There was modest variation (three- to fourfold) in DC-SIGN expression levels between individuals and between DCs isolated from the same individual at different times. Several MAbs efficiently blocked virus binding to cell lines expressing human or rhesus DC-SIGN, preventing HIV and SIV transmission. Interactions with Ebola virus pseudotypes were also blocked efficiently. Despite their ability to block virus-DC-SIGN interactions on cell lines, these antibodies only inhibited transmission of virus from DCs by approximately 50% or less. These results indicate that factors other than DC-SIGN may play important roles in the ability of DCs to capture and transmit HIV.

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Repurposing the Ebola and Marburg Virus Inhibitors Tilorone, Quinacrine and Pyronaridine: In vitro Activity Against SARS-CoV-2 and Potential Mechanisms

10.1101/2020.12.01.407361 ◽

2020 ◽

Author(s):

Ana C. Puhl ◽

Ethan James Fritch ◽

Thomas R. Lane ◽

Longping V. Tse ◽

Boyd L. Yount ◽

...

Keyword(s):

Cell Lines ◽

Small Molecule ◽

Ebola Virus ◽

Early Stage ◽

Drug Repurposing ◽

Marburg Virus ◽

Small Molecule Drugs ◽

Speed Up

AbstractSARS-CoV-2 is a newly identified virus that has resulted in over 1.3 M deaths globally and over 59 M cases globally to date. Small molecule inhibitors that reverse disease severity have proven difficult to discover. One of the key approaches that has been widely applied in an effort to speed up the translation of drugs is drug repurposing. A few drugs have shown in vitro activity against Ebola virus and demonstrated activity against SARS-CoV-2 in vivo. Most notably the RNA polymerase targeting remdesivir demonstrated activity in vitro and efficacy in the early stage of the disease in humans. Testing other small molecule drugs that are active against Ebola virus would seem a reasonable strategy to evaluate their potential for SARS-CoV-2. We have previously repurposed pyronaridine, tilorone and quinacrine (from malaria, influenza, and antiprotozoal uses, respectively) as inhibitors of Ebola and Marburg virus in vitro in HeLa cells and of mouse adapted Ebola virus in mouse in vivo. We have now tested these three drugs in various cell lines (VeroE6, Vero76, Caco-2, Calu-3, A549-ACE2, HUH-7 and monocytes) infected with SARS-CoV-2 as well as other viruses (including MHV and HCoV 229E). The compilation of these results indicated considerable variability in antiviral activity observed across cell lines. We found that tilorone and pyronaridine inhibited the virus replication in A549-ACE2 cells with IC50 values of 180 nM and IC50 198 nM, respectively. We have also tested them in a pseudovirus assay and used microscale thermophoresis to test the binding of these molecules to the spike protein. They bind to spike RBD protein with Kd values of 339 nM and 647 nM, respectively. Human Cmax for pyronaridine and quinacrine is greater than the IC50 hence justifying in vivo evaluation. We also provide novel insights into their mechanism which is likely lysosomotropic.

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Human antibody cocktail deploys multiple functions to confer pan-ebolavirus protection

10.1101/395525 ◽

2018 ◽

Author(s):

Anna Z. Wec ◽

Zachary A. Bornholdt ◽

Shihua He ◽

Andrew S. Herbert ◽

Eileen Goodwin ◽

...

Keyword(s):

Ebola Virus ◽

Virus Disease ◽

Protective Efficacy ◽

Affinity Maturation ◽

Surface Glycoprotein ◽

Human Antibody ◽

Human Mabs ◽

Western Africa ◽

Guinea Pig Model

During the unprecedented 2013–2016 Ebola virus disease (EVD) epidemic in Western Africa and in its aftermath, the passive administration of monoclonal antibodies (mAbs) emerged as a promising treatment approach1–7. However, all antibody-based therapeutics currently in advanced development are specific for a single member of the Ebolavirus genus, Ebola virus (EBOV), and ineffective against divergent outbreak-causing ebolaviruses, including Bundibugyo virus (BDBV) and Sudan virus (SUDV)2,3,5,7. Here we advance MBP134, a cocktail of two broadly neutralizing human mAbs targeting the filovirus surface glycoprotein, GP, as a candidate pan-ebolavirus therapeutic. One component of this cocktail is a pan-ebolavirus neutralizing mAb, ADI-15878, isolated from a human EVD survivor8,9. The second, ADI-23774, was derived by affinity maturation of a human mAb8,9 via yeast display to enhance its potency against SUDV. MBP134 afforded exceptionally potent pan-ebolavirus neutralization in vitro and demonstrated greater protective efficacy than ADI-15878 alone in the guinea pig model of lethal EBOV challenge. A second-generation cocktail, MBP134AF, engineered to effectively harness natural killer (NK) cells afforded additional, unprecedented improvements in protective efficacy against EBOV and SUDV in guinea pigs relative to both its precursor and to any mAbs or mAb cocktails tested previously. MBP134AF is a best-in-class mAb cocktail suitable for evaluation as a pan-ebolavirus therapeutic in nonhuman primates.

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Canine Morbillivirus from Colombian Lineage Exhibits In Silico and In Vitro Potential to Infect Human Cells

Pathogens ◽

10.3390/pathogens10091199 ◽

2021 ◽

Vol 10 (9) ◽

pp. 1199

Author(s):

Santiago Rendon-Marin ◽

Carolina Quintero-Gil ◽

Diego Guerra ◽

Carlos Muskus ◽

Julian Ruiz-Saenz

Keyword(s):

Cell Lines ◽

Protein Interactions ◽

In Silico ◽

Measles Virus ◽

Mononuclear Cells ◽

Viral Agent ◽

Cellular Receptors ◽

Transmission Potential ◽

H Protein

Canine morbillivirus (CDV) is a viral agent that infects domestic dogs and a vast array of wildlife species. It belongs to the Paramyxoviridae family, genus Morbillivirus, which is shared with the Measles virus (MeV). Both viruses employ orthologous cellular receptors, SLAM in mononuclear cells and Nectin-4 in epithelial cells, to enter the cells. Although CDV and MeV hemagglutinin (H) have similar functions in viral pathogenesis and cell tropism, the potential interaction of CDV-H protein with human cellular receptors is still uncertain. Considering that CDV is classified as a multi-host pathogen, the potential risk of CDV transmission to humans has not been fully discarded. In this study, we aimed to evaluate both in silico and in vitro, whether there is a cross-species transmission potential from CDV to humans. To accomplish this, the CDV-H protein belonging to the Colombian lineage was modelled. After model validations, molecular docking and molecular dynamics simulations were carried out between Colombian CDV-H protein and canine and human cellular receptors to determine different aspects of the protein–protein interactions. Moreover, cell lines expressing orthologous cellular receptors, with both reference and wild-type CDV strains, were conducted to determine the CDV cross-species transmission potential from an in vitro model. This in silico and in vitro approach suggests the possibility that CDV interacts with ortholog human SLAM (hSLAM) and human Nectin-4 receptors to infect human cell lines, which could imply a potential cross-species transmission of CDV from dogs to humans.

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Mechanism of Cellular Formation and In Vivo Seeding Effects of Hexameric β-Amyloid Assemblies

Molecular Neurobiology ◽

10.1007/s12035-021-02567-8 ◽

2021 ◽

Author(s):

Céline Vrancx ◽

Devkee M. Vadukul ◽

Nuria Suelves ◽

Sabrina Contino ◽

Ludovic D’Auria ◽

...

Keyword(s):

Cell Lines ◽

Self Assembly ◽

Amyloid Fibrils ◽

Senile Plaques ◽

Amyloid Peptide ◽

Cellular Models ◽

Aβ Aggregation ◽

Β Amyloid

AbstractThe β-amyloid peptide (Aβ) is found as amyloid fibrils in senile plaques, a typical hallmark of Alzheimer’s disease (AD). However, intermediate soluble oligomers of Aβ are now recognized as initiators of the pathogenic cascade leading to AD. Studies using recombinant Aβ have shown that hexameric Aβ in particular acts as a critical nucleus for Aβ self-assembly. We recently isolated hexameric Aβ assemblies from a cellular model, and demonstrated their ability to enhance Aβ aggregation in vitro. Here, we report the presence of similar hexameric-like Aβ assemblies across several cellular models, including neuronal-like cell lines. In order to better understand how they are produced in a cellular context, we investigated the role of presenilin-1 (PS1) and presenilin-2 (PS2) in their formation. PS1 and PS2 are the catalytic subunits of the γ-secretase complex that generates Aβ. Using CRISPR-Cas9 to knockdown each of the two presenilins in neuronal-like cell lines, we observed a direct link between the PS2-dependent processing pathway and the release of hexameric-like Aβ assemblies in extracellular vesicles. Further, we assessed the contribution of hexameric Aβ to the development of amyloid pathology. We report the early presence of hexameric-like Aβ assemblies in both transgenic mice brains exhibiting human Aβ pathology and in the cerebrospinal fluid of AD patients, suggesting hexameric Aβ as a potential early AD biomarker. Finally, cell-derived hexameric Aβ was found to seed other human Aβ forms, resulting in the aggravation of amyloid deposition in vivo and neuronal toxicity in vitro.

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Mechanisms of phosphatidylserine influence on viral production: a computational model of Ebola virus matrix protein assembly

10.1101/2021.07.22.453424 ◽

2021 ◽

Author(s):

Xiao Liu ◽

Ethan J Pappas ◽

Monica L Husby ◽

Robert V Stahelin ◽

Elsje Pienaar

Keyword(s):

Cell Membrane ◽

Computational Model ◽

Host Cell ◽

Matrix Protein ◽

Ebola Virus ◽

Global Public Health ◽

Matrix Assembly ◽

Host Cell Membrane ◽

Western Africa

Ebola virus (EBOV) infections continue to pose a global public health threat, with high mortality rates and sporadic outbreaks in Central and Western Africa. A quantitative understanding of the key processes driving EBOV assembly and budding could provide valuable insights to inform drug development. Here we used a computational model to evaluate EBOV matrix assembly. Our model focused on the assembly kinetics of VP40, the matrix protein in EBOV, and its interaction with phosphatidylserine (PS) in the host cell membrane. Human cells transfected with VP40-expressing plasmids are capable of producing virus-like particles (VLPs) that closely resemble EBOV virions. We used data from this in vitro VP40 system to calibrate our computational model. PS levels in the host cell membrane had been shown to affect VP40 dynamics as well as VLP production through recruiting VP40 dimers to plasma membrane inner leaflet. Our computational results indicated that PS may have direct influence on VP40 filament growth and affect multiple steps in the assembly and budding of VP40 VLPs. We also proposed that the assembly of VP40 filaments may follow the nucleation-elongation theory where initialization and oligomerization of VP40 are two separate and distinct steps in the assembly process. This work illustrated how computational and experimental approaches can be combined to allow for additional analysis and hypothesis generation. Our findings advanced understanding of the molecular process of EBOV assembly and budding processes and may help the development of new EBOV treatments targeting VP40 matrix assembly.

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Repurposing of Berbamine Hydrochloride to Inhibit Ebola Virus by Targeting Viral Glycoprotein

10.21203/rs.3.rs-111854/v1 ◽

2020 ◽

Author(s):

Shan Cen ◽

Dongrong Yi ◽

Quanjie Li ◽

Xiangguo Qiu ◽

Han Wang ◽

...

Keyword(s):

Ebola Virus ◽

Low Cost ◽

Great Promise ◽

Viral Glycoprotein ◽

Viral Receptor ◽

Small Molecule Drugs ◽

Ebov Infection ◽

Direct Binding

Abstract Ebola virus (EBOV) infection leads to staggeringly high mortality rates. Effective and low-cost treatments are urgently needed to control frequent EBOV outbreaks in Africa. In this study, we report that a natural compound called berbamine hydrochloride strongly inhibits EBOV replication in vitro and in vivo. Our work further showed that berbamine hydrochloride acts by direct binding to the cleaved EBOV glycoprotein (GPcl), disrupting GPcl interaction with viral receptor NPC1, thus blocking the fusion of viral and cellular membranes. Our data support the probability of developing anti-EBOV small molecule drugs by targeting viral GPcl. More importantly, since berbamine hydrochloride has been used in clinic to treat leukopenia, it holds great promise of being quickly repurposed as an anti-EBOV drug.

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