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1.
Viruses ; 14(2)2022 01 19.
Article in English | MEDLINE | ID: covidwho-1625815

ABSTRACT

SARS-CoV-2, a member of the coronavirus family, is the causative agent of the COVID-19 pandemic. Currently, there is still an urgent need in developing an efficient therapeutic intervention. In this study, we aimed at evaluating the therapeutic effect of a single intranasal treatment of the TLR3/MDA5 synthetic agonist Poly(I:C) against a lethal dose of SARS-CoV-2 in K18-hACE2 transgenic mice. We demonstrate here that early Poly(I:C) treatment acts synergistically with SARS-CoV-2 to induce an intense, immediate and transient upregulation of innate immunity-related genes in lungs. This effect is accompanied by viral load reduction, lung and brain cytokine storms prevention and increased levels of macrophages and NK cells, resulting in 83% mice survival, concomitantly with long-term immunization. Thus, priming the lung innate immunity by Poly(I:C) or alike may provide an immediate, efficient and safe protective measure against SARS-CoV-2 infection.


Subject(s)
COVID-19/immunology , COVID-19/prevention & control , Immunity, Innate , Poly I-C/immunology , Poly I-C/therapeutic use , SARS-CoV-2/drug effects , Toll-Like Receptor 3/agonists , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/immunology , Animals , COVID-19/drug therapy , Cytokine Release Syndrome/immunology , Cytokine Release Syndrome/prevention & control , Disease Models, Animal , Female , Humans , Lung/immunology , Lung/virology , Mice , Mice, Transgenic , SARS-CoV-2/immunology , Toll-Like Receptor 3/immunology , Viral Load/drug effects
2.
J Med Virol ; 93(9): 5376-5389, 2021 09.
Article in English | MEDLINE | ID: covidwho-1363676

ABSTRACT

The suppression of types I and III interferon (IFN) responses by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributes to the pathogenesis of coronavirus disease 2019 (COVID-19). The strategy used by SARS-CoV-2 to evade antiviral immunity needs further investigation. Here, we reported that SARS-CoV-2 ORF9b inhibited types I and III IFN production by targeting multiple molecules of innate antiviral signaling pathways. SARS-CoV-2 ORF9b impaired the induction of types I and III IFNs by Sendai virus and poly (I:C). SARS-CoV-2 ORF9b inhibited the activation of types I and III IFNs induced by the components of cytosolic dsRNA-sensing pathways of RIG-I/MDA5-MAVS signaling, including RIG-I, MDA-5, MAVS, TBK1, and IKKε, rather than IRF3-5D, which is the active form of IRF3. SARS-CoV-2 ORF9b also suppressed the induction of types I and III IFNs by TRIF and STING, which are the adaptor protein of the endosome RNA-sensing pathway of TLR3-TRIF signaling and the adaptor protein of the cytosolic DNA-sensing pathway of cGAS-STING signaling, respectively. A mechanistic analysis revealed that the SARS-CoV-2 ORF9b protein interacted with RIG-I, MDA-5, MAVS, TRIF, STING, and TBK1 and impeded the phosphorylation and nuclear translocation of IRF3. In addition, SARS-CoV-2 ORF9b facilitated the replication of the vesicular stomatitis virus. Therefore, the results showed that SARS-CoV-2 ORF9b negatively regulates antiviral immunity and thus facilitates viral replication. This study contributes to our understanding of the molecular mechanism through which SARS-CoV-2 impairs antiviral immunity and provides an essential clue to the pathogenesis of COVID-19.


Subject(s)
DEAD Box Protein 58/immunology , Immune Evasion/genetics , Interferons/immunology , Nucleotidyltransferases/immunology , Receptors, Immunologic/immunology , SARS-CoV-2/immunology , Toll-Like Receptor 3/immunology , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/immunology , Adaptor Proteins, Vesicular Transport/genetics , Adaptor Proteins, Vesicular Transport/immunology , Animals , Chlorocebus aethiops , Coronavirus Nucleocapsid Proteins/genetics , Coronavirus Nucleocapsid Proteins/immunology , DEAD Box Protein 58/genetics , Gene Expression Regulation , HEK293 Cells , HeLa Cells , Humans , I-kappa B Kinase/genetics , I-kappa B Kinase/immunology , Immunity, Innate , Interferon Regulatory Factor-3/genetics , Interferon Regulatory Factor-3/immunology , Interferon-Induced Helicase, IFIH1/genetics , Interferon-Induced Helicase, IFIH1/immunology , Interferons/genetics , Membrane Proteins/genetics , Membrane Proteins/immunology , Nucleotidyltransferases/genetics , Phosphoproteins/genetics , Phosphoproteins/immunology , Plasmids/chemistry , Plasmids/metabolism , /immunology , Receptors, Immunologic/genetics , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Signal Transduction/genetics , Signal Transduction/immunology , Toll-Like Receptor 3/genetics , Transfection , Vero Cells , Virus Replication/immunology
3.
J Med Virol ; 93(9): 5376-5389, 2021 09.
Article in English | MEDLINE | ID: covidwho-1206842

ABSTRACT

The suppression of types I and III interferon (IFN) responses by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributes to the pathogenesis of coronavirus disease 2019 (COVID-19). The strategy used by SARS-CoV-2 to evade antiviral immunity needs further investigation. Here, we reported that SARS-CoV-2 ORF9b inhibited types I and III IFN production by targeting multiple molecules of innate antiviral signaling pathways. SARS-CoV-2 ORF9b impaired the induction of types I and III IFNs by Sendai virus and poly (I:C). SARS-CoV-2 ORF9b inhibited the activation of types I and III IFNs induced by the components of cytosolic dsRNA-sensing pathways of RIG-I/MDA5-MAVS signaling, including RIG-I, MDA-5, MAVS, TBK1, and IKKε, rather than IRF3-5D, which is the active form of IRF3. SARS-CoV-2 ORF9b also suppressed the induction of types I and III IFNs by TRIF and STING, which are the adaptor protein of the endosome RNA-sensing pathway of TLR3-TRIF signaling and the adaptor protein of the cytosolic DNA-sensing pathway of cGAS-STING signaling, respectively. A mechanistic analysis revealed that the SARS-CoV-2 ORF9b protein interacted with RIG-I, MDA-5, MAVS, TRIF, STING, and TBK1 and impeded the phosphorylation and nuclear translocation of IRF3. In addition, SARS-CoV-2 ORF9b facilitated the replication of the vesicular stomatitis virus. Therefore, the results showed that SARS-CoV-2 ORF9b negatively regulates antiviral immunity and thus facilitates viral replication. This study contributes to our understanding of the molecular mechanism through which SARS-CoV-2 impairs antiviral immunity and provides an essential clue to the pathogenesis of COVID-19.


Subject(s)
DEAD Box Protein 58/immunology , Immune Evasion/genetics , Interferons/immunology , Nucleotidyltransferases/immunology , Receptors, Immunologic/immunology , SARS-CoV-2/immunology , Toll-Like Receptor 3/immunology , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/immunology , Adaptor Proteins, Vesicular Transport/genetics , Adaptor Proteins, Vesicular Transport/immunology , Animals , Chlorocebus aethiops , Coronavirus Nucleocapsid Proteins/genetics , Coronavirus Nucleocapsid Proteins/immunology , DEAD Box Protein 58/genetics , Gene Expression Regulation , HEK293 Cells , HeLa Cells , Humans , I-kappa B Kinase/genetics , I-kappa B Kinase/immunology , Immunity, Innate , Interferon Regulatory Factor-3/genetics , Interferon Regulatory Factor-3/immunology , Interferon-Induced Helicase, IFIH1/genetics , Interferon-Induced Helicase, IFIH1/immunology , Interferons/genetics , Membrane Proteins/genetics , Membrane Proteins/immunology , Nucleotidyltransferases/genetics , Phosphoproteins/genetics , Phosphoproteins/immunology , Plasmids/chemistry , Plasmids/metabolism , /immunology , Receptors, Immunologic/genetics , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Signal Transduction/genetics , Signal Transduction/immunology , Toll-Like Receptor 3/genetics , Transfection , Vero Cells , Virus Replication/immunology
4.
J Med Virol ; 92(10): 2114-2123, 2020 10.
Article in English | MEDLINE | ID: covidwho-777547

ABSTRACT

The present study provides the first multiepitope vaccine construct using the 3CL hydrolase protein of SARS-CoV-2. The coronavirus 3CL hydrolase (Mpro) enzyme is essential for proteolytic maturation of the virus. This study was based on immunoinformatics and structural vaccinology strategies. The design of the multiepitope vaccine was built using helper T-cell and cytotoxic T-cell epitopes from the 3CL hydrolase protein along with an adjuvant to enhance immune response; these are joined to each other by short peptide linkers. The vaccine also carries potential B-cell linear epitope regions, B-cell discontinuous epitopes, and interferon-γ-inducing epitopes. Epitopes of the constructed multiepitope vaccine were found to be antigenic, nonallergic, nontoxic, and covering large human populations worldwide. The vaccine construct was modeled, validated, and refined by different programs to achieve a high-quality three-dimensional structure. The resulting high-quality model was applied for conformational B-cell epitope selection and docking analyses with toll-like receptor-3 for understanding the capability of the vaccine to elicit an immune response. In silico cloning and codon adaptation were also performed with the pET-19b plasmid vector. The designed multiepitope peptide vaccine may prompt the development of a vaccine to control SARS-CoV-2 infection.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , Coronavirus 3C Proteases/immunology , Epitopes, B-Lymphocyte/immunology , Epitopes, T-Lymphocyte/immunology , SARS-CoV-2/immunology , Toll-Like Receptor 3/immunology , Amino Acid Sequence , Binding Sites , COVID-19/immunology , COVID-19/virology , COVID-19 Vaccines/genetics , Cloning, Molecular/methods , Computational Biology/methods , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/genetics , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/genetics , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/genetics , Genetic Vectors/chemistry , Genetic Vectors/immunology , HLA Antigens/chemistry , HLA Antigens/genetics , HLA Antigens/immunology , Humans , Immunity, Innate/drug effects , Immunogenicity, Vaccine , Interferon-gamma/genetics , Interferon-gamma/immunology , Molecular Docking Simulation , Protein Binding , Protein Interaction Domains and Motifs , SARS-CoV-2/drug effects , SARS-CoV-2/pathogenicity , T-Lymphocytes, Cytotoxic/chemistry , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/virology , T-Lymphocytes, Helper-Inducer/chemistry , T-Lymphocytes, Helper-Inducer/immunology , T-Lymphocytes, Helper-Inducer/virology , Toll-Like Receptor 3/chemistry , Toll-Like Receptor 3/genetics , User-Computer Interface , Vaccines, Subunit
5.
Infect Dis Poverty ; 9(1): 132, 2020 Sep 16.
Article in English | MEDLINE | ID: covidwho-768657

ABSTRACT

BACKGROUND: Coronavirus disease 2019 (COVID-19) linked with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cause severe illness and life-threatening pneumonia in humans. The current COVID-19 pandemic demands an effective vaccine to acquire protection against the infection. Therefore, the present study was aimed to design a multiepitope-based subunit vaccine (MESV) against COVID-19. METHODS: Structural proteins (Surface glycoprotein, Envelope protein, and Membrane glycoprotein) of SARS-CoV-2 are responsible for its prime functions. Sequences of proteins were downloaded from GenBank and several immunoinformatics coupled with computational approaches were employed to forecast B- and T- cell epitopes from the SARS-CoV-2 highly antigenic structural proteins to design an effective MESV. RESULTS: Predicted epitopes suggested high antigenicity, conserveness, substantial interactions with the human leukocyte antigen (HLA) binding alleles, and collective global population coverage of 88.40%. Taken together, 276 amino acids long MESV was designed by connecting 3 cytotoxic T lymphocytes (CTL), 6 helper T lymphocyte (HTL) and 4 B-cell epitopes with suitable adjuvant and linkers. The MESV construct was non-allergenic, stable, and highly antigenic. Molecular docking showed a stable and high binding affinity of MESV with human pathogenic toll-like receptors-3 (TLR3). Furthermore, in silico immune simulation revealed significant immunogenic response of MESV. Finally, MEV codons were optimized for its in silico cloning into the Escherichia coli K-12 system, to ensure its increased expression. CONCLUSION: The MESV developed in this study is capable of generating immune response against COVID-19. Therefore, if designed MESV further investigated experimentally, it would be an effective vaccine candidate against SARS-CoV-2 to control and prevent COVID-19.


Subject(s)
Betacoronavirus/immunology , Coronavirus Infections/prevention & control , Epitopes, B-Lymphocyte/immunology , Epitopes, T-Lymphocyte/immunology , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Viral Vaccines/immunology , COVID-19 , COVID-19 Vaccines , Coronavirus Infections/genetics , Coronavirus Infections/immunology , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/genetics , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/genetics , Humans , Immunogenicity, Vaccine/immunology , Molecular Docking Simulation , Pneumonia, Viral/immunology , SARS-CoV-2 , Sequence Analysis, Protein , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Toll-Like Receptor 3/chemistry , Toll-Like Receptor 3/genetics , Toll-Like Receptor 3/immunology , Vaccines, Subunit/chemistry , Vaccines, Subunit/genetics , Vaccines, Subunit/immunology , Vaccinology/methods , Viral Matrix Proteins/chemistry , Viral Matrix Proteins/genetics , Viral Matrix Proteins/immunology , Viral Vaccines/chemistry , Viral Vaccines/genetics
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