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1.
Cell Rep ; 15(7): 1514-1526, 2016 05 17.
Article in English | MEDLINE | ID: mdl-27160900

ABSTRACT

Previous efforts to identify cross-neutralizing antibodies to the receptor-binding site (RBS) of ebolavirus glycoproteins have been unsuccessful, largely because the RBS is occluded on the viral surface. We report a monoclonal antibody (FVM04) that targets a uniquely exposed epitope within the RBS; cross-neutralizes Ebola (EBOV), Sudan (SUDV), and, to a lesser extent, Bundibugyo viruses; and shows protection against EBOV and SUDV in mice and guinea pigs. The antibody cocktail ZMapp™ is remarkably effective against EBOV (Zaire) but does not cross-neutralize other ebolaviruses. By replacing one of the ZMapp™ components with FVM04, we retained the anti-EBOV efficacy while extending the breadth of protection to SUDV, thereby generating a cross-protective antibody cocktail. In addition, we report several mutations at the base of the ebolavirus glycoprotein that enhance the binding of FVM04 and other cross-reactive antibodies. These findings have important implications for pan-ebolavirus vaccine development and defining broadly protective antibody cocktails.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Ebolavirus/physiology , Epitopes/immunology , Glycoproteins/metabolism , Hemorrhagic Fever, Ebola/immunology , Receptors, Virus/metabolism , Amino Acid Sequence , Animals , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/ultrastructure , Antibodies, Neutralizing , Antibodies, Viral/chemistry , Binding Sites , Disease Models, Animal , Female , Glycoproteins/chemistry , Glycoproteins/genetics , Glycoproteins/ultrastructure , Guinea Pigs , HEK293 Cells , Humans , Kinetics , Mice, Inbred BALB C , Models, Molecular , Mutation/genetics , Negative Staining , Neutralization Tests , Treatment Outcome
2.
Viruses ; 8(3): 71, 2016 Mar 07.
Article in English | MEDLINE | ID: mdl-27072420

ABSTRACT

Iminosugars that are competitive inhibitors of endoplasmic reticulum (ER) α-glucosidases have been demonstrated to have antiviral activity against a diverse set of viruses. A novel iminosugar, UV-4B, has recently been shown to provide protection against lethal infections with dengue and influenza A (H1N1) viruses in mice. In the current study, the breadth of activity of UV-4B against influenza was examined ex vivo and in vivo. Efficacy of UV-4B against influenza A and B viruses was shown in primary human bronchial epithelial cells, a principal target tissue for influenza. Efficacy of UV-4B against influenza A (H1N1 and H3N2 subtypes) and influenza B was demonstrated using multiple lethal mouse models with readouts including mortality and weight loss. Clinical trials are ongoing to demonstrate safety of UV-4B and future studies to evaluate antiviral activity against influenza in humans are planned.


Subject(s)
1-Deoxynojirimycin/analogs & derivatives , Antiviral Agents/administration & dosage , Influenza A virus/drug effects , Influenza B virus/drug effects , Orthomyxoviridae Infections/drug therapy , 1-Deoxynojirimycin/administration & dosage , 1-Deoxynojirimycin/pharmacology , Animals , Antiviral Agents/pharmacology , Body Weight , Cells, Cultured , Disease Models, Animal , Epithelial Cells/virology , Humans , Mice , Orthomyxoviridae Infections/pathology , Orthomyxoviridae Infections/virology , Survival Analysis , Treatment Outcome
3.
Antiviral Res ; 129: 93-98, 2016 May.
Article in English | MEDLINE | ID: mdl-26946111

ABSTRACT

The antiviral activity of UV-4 was previously demonstrated against dengue virus serotype 2 (DENV2) in multiple mouse models. Herein, step-wise minimal effective dose and therapeutic window of efficacy studies of UV-4B (UV-4 hydrochloride salt) were conducted in an antibody-dependent enhancement (ADE) mouse model of severe DENV2 infection in AG129 mice lacking types I and II interferon receptors. Significant survival benefit was demonstrated with 10-20 mg/kg of UV-4B administered thrice daily (TID) for seven days with initiation of treatment up to 48 h after infection. UV-4B also reduced infectious virus production in in vitro antiviral activity assays against all four DENV serotypes, including clinical isolates. A set of purified enzyme, in vitro, and in vivo studies demonstrated that inhibition of endoplasmic reticulum (ER) α-glucosidases and not the glycosphingolipid pathway appears to be responsible for the antiviral activity of UV-4B against DENV. Along with a comprehensive safety package, these and previously published data provided support for an Investigational New Drug (IND) filing and Phases 1 and 2 clinical trials for UV-4B with an indication of acute dengue disease.


Subject(s)
1-Deoxynojirimycin/analogs & derivatives , Antiviral Agents/pharmacology , Dengue Virus/drug effects , Glycoside Hydrolase Inhibitors/pharmacology , Severe Dengue/drug therapy , alpha-Glucosidases/metabolism , 1-Deoxynojirimycin/administration & dosage , 1-Deoxynojirimycin/pharmacology , 1-Deoxynojirimycin/therapeutic use , Animals , Antibodies, Viral/blood , Antibody-Dependent Enhancement/drug effects , Antiviral Agents/administration & dosage , Antiviral Agents/therapeutic use , Cells, Cultured , Chlorocebus aethiops , Clinical Trials as Topic , Disease Models, Animal , Drugs, Investigational , Endoplasmic Reticulum/drug effects , Endoplasmic Reticulum/enzymology , Glycoside Hydrolase Inhibitors/administration & dosage , Glycoside Hydrolase Inhibitors/chemistry , Glycoside Hydrolase Inhibitors/therapeutic use , Humans , Inhibitory Concentration 50 , Mice , Monocytes/virology , Receptors, Interferon/deficiency , Serogroup , Severe Dengue/virology , Vero Cells
4.
J Virol ; 90(1): 279-91, 2016 01 01.
Article in English | MEDLINE | ID: mdl-26468532

ABSTRACT

UNLABELLED: Filoviruses cause highly lethal viral hemorrhagic fever in humans and nonhuman primates. Current immunotherapeutic options for filoviruses are mostly specific to Ebola virus (EBOV), although other members of Filoviridae such as Sudan virus (SUDV), Bundibugyo virus (BDBV), and Marburg virus (MARV) have also caused sizeable human outbreaks. Here we report a set of pan-ebolavirus and pan-filovirus monoclonal antibodies (MAbs) derived from cynomolgus macaques immunized repeatedly with a mixture of engineered glycoproteins (GPs) and virus-like particles (VLPs) for three different filovirus species. The antibodies recognize novel neutralizing and nonneutralizing epitopes on the filovirus glycoprotein, including conserved conformational epitopes within the core regions of the GP1 subunit and a novel linear epitope within the glycan cap. We further report the first filovirus antibody binding to a highly conserved epitope within the fusion loop of ebolavirus and marburgvirus species. One of the antibodies binding to the core GP1 region of all ebolavirus species and with lower affinity to MARV GP cross neutralized both SUDV and EBOV, the most divergent ebolavirus species. In a mouse model of EBOV infection, this antibody provided 100% protection when administered in two doses and partial, but significant, protection when given once at the peak of viremia 3 days postinfection. Furthermore, we describe novel cocktails of antibodies with enhanced protective efficacy compared to individual MAbs. In summary, the present work describes multiple novel, cross-reactive filovirus epitopes and innovative combination concepts that challenge the current therapeutic models. IMPORTANCE: Filoviruses are among the most deadly human pathogens. The 2014-2015 outbreak of Ebola virus disease (EVD) led to more than 27,000 cases and 11,000 fatalities. While there are five species of Ebolavirus and several strains of marburgvirus, the current immunotherapeutics primarily target Ebola virus. Since the nature of future outbreaks cannot be predicted, there is an urgent need for therapeutics with broad protective efficacy against multiple filoviruses. Here we describe a set of monoclonal antibodies cross-reactive with multiple filovirus species. These antibodies target novel conserved epitopes within the envelope glycoprotein and exhibit protective efficacy in mice. We further present novel concepts for combination of cross-reactive antibodies against multiple epitopes that show enhanced efficacy compared to monotherapy and provide complete protection in mice. These findings set the stage for further evaluation of these antibodies in nonhuman primates and development of effective pan-filovirus immunotherapeutics for use in future outbreaks.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Viral/immunology , Epitopes/immunology , Filoviridae/immunology , Glycoproteins/immunology , Hemorrhagic Fever, Ebola/prevention & control , Viral Proteins/immunology , Animals , Antibodies, Monoclonal/isolation & purification , Antibodies, Monoclonal/therapeutic use , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/isolation & purification , Antibodies, Neutralizing/therapeutic use , Antibodies, Viral/isolation & purification , Antibodies, Viral/therapeutic use , Cross Reactions , Disease Models, Animal , Female , Immunization, Passive , Macaca , Mice, Inbred BALB C , Survival Analysis , Treatment Outcome
5.
Viruses ; 7(5): 2404-27, 2015 May 13.
Article in English | MEDLINE | ID: mdl-25984714

ABSTRACT

Iminosugars are capable of targeting the life cycles of multiple viruses by blocking host endoplasmic reticulum α-glucosidase enzymes that are required for competent replication of a variety of enveloped, glycosylated viruses. Iminosugars as a class are approved for use in humans with diseases such as diabetes and Gaucher's disease, providing evidence for safety of this class of compounds. The in vitro antiviral activity of iminosugars has been described in several publications with a subset of these demonstrating in vivo activity against flaviviruses, herpesviruses, retroviruses and filoviruses. Although there is compelling non-clinical in vivo evidence of antiviral efficacy, the efficacy of iminosugars as antivirals has yet to be demonstrated in humans. In the current study, we report a novel iminosugar, UV-12, which has efficacy against dengue and influenza in mouse models. UV-12 exhibits drug-like properties including oral bioavailability and good safety profile in mice and guinea pigs. UV-12 is an example of an iminosugar with activity against multiple virus families that should be investigated in further safety and efficacy studies and demonstrates potential value of this drug class as antiviral therapeutics.


Subject(s)
Antiviral Agents/therapeutic use , Dengue/drug therapy , Imino Sugars/therapeutic use , Orthomyxoviridae Infections/drug therapy , Animals , Antiviral Agents/pharmacology , Dengue Virus/drug effects , Disease Models, Animal , Female , Guinea Pigs , Imino Sugars/pharmacology , Male , Mice , Microbial Sensitivity Tests , Orthomyxoviridae/drug effects , Treatment Outcome
6.
J Pharm Sci ; 100(12): 5156-73, 2011 Dec.
Article in English | MEDLINE | ID: mdl-21858822

ABSTRACT

The filoviruses, Ebola virus and Marburg virus, cause severe hemorrhagic fever with up to 90% human mortality. Virus-like particles of EBOV (eVLPs) and MARV (mVLPs) are attractive vaccine candidates. For the development of stable vaccines, the conformational stability of these two enveloped VLPs produced in insect cells was characterized by various spectroscopic techniques over a wide pH and temperature range. Temperature-induced aggregation of the VLPs at various pH values was monitored by light scattering. Temperature/pH empirical phase diagrams (EPDs) of the two VLPs were constructed to summarize the large volume of data generated. The EPDs show that both VLPs lose their conformational integrity above about 50°C-60°C, depending on solution pH. The VLPs were maximally thermal stable in solution at pH 7-8, with a significant reduction in stability at pH 5 and 6. They were much less stable in solution at pH 3-4 due to increased susceptibility of the VLPs to aggregation. The characterization data and conformational stability profiles from these studies provide a basis for selection of optimized solution conditions for further vaccine formulation and long-term stability studies of eVLPs and mVLPs.


Subject(s)
Ebolavirus/metabolism , Marburg Virus Disease/metabolism , Marburgvirus/metabolism , Virion/chemistry , Animals , Antibodies, Viral , Baculoviridae/genetics , Baculoviridae/metabolism , Cell Line , Ebola Vaccines/analysis , Ebola Vaccines/chemistry , Ebola Vaccines/metabolism , Ebolavirus/chemistry , Ebolavirus/genetics , Ebolavirus/immunology , Genetic Vectors , Hemorrhagic Fever, Ebola/immunology , Hemorrhagic Fever, Ebola/metabolism , Humans , Hydrogen-Ion Concentration , Insecta , Marburg Virus Disease/immunology , Marburgvirus/chemistry , Marburgvirus/genetics , Marburgvirus/immunology , Molecular Conformation , Temperature , Vaccines, Virus-Like Particle/analysis , Vaccines, Virus-Like Particle/chemistry , Vaccines, Virus-Like Particle/immunology , Vaccines, Virus-Like Particle/metabolism
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