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1.
Cell ; 184(9): 2316-2331.e15, 2021 04 29.
Artículo en Inglés | MEDLINE | ID: mdl-33773105

RESUMEN

Most human monoclonal antibodies (mAbs) neutralizing SARS-CoV-2 recognize the spike (S) protein receptor-binding domain and block virus interactions with the cellular receptor angiotensin-converting enzyme 2. We describe a panel of human mAbs binding to diverse epitopes on the N-terminal domain (NTD) of S protein from SARS-CoV-2 convalescent donors and found a minority of these possessed neutralizing activity. Two mAbs (COV2-2676 and COV2-2489) inhibited infection of authentic SARS-CoV-2 and recombinant VSV/SARS-CoV-2 viruses. We mapped their binding epitopes by alanine-scanning mutagenesis and selection of functional SARS-CoV-2 S neutralization escape variants. Mechanistic studies showed that these antibodies neutralize in part by inhibiting a post-attachment step in the infection cycle. COV2-2676 and COV2-2489 offered protection either as prophylaxis or therapy, and Fc effector functions were required for optimal protection. Thus, natural infection induces a subset of potent NTD-specific mAbs that leverage neutralizing and Fc-mediated activities to protect against SARS-CoV-2 infection using multiple functional attributes.


Asunto(s)
Anticuerpos Monoclonales/farmacología , Anticuerpos Neutralizantes/farmacología , Sustancias Protectoras/farmacología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/inmunología , Animales , Unión Competitiva , COVID-19/inmunología , COVID-19/virología , Quimiocinas/metabolismo , Chlorocebus aethiops , Células HEK293 , Humanos , Fragmentos Fab de Inmunoglobulinas/metabolismo , Inmunoglobulina G/metabolismo , Pulmón/metabolismo , Ratones Endogámicos C57BL , Modelos Moleculares , Mutagénesis/genética , Pruebas de Neutralización , Dominios Proteicos , Células Vero
2.
Cell ; 184(17): 4414-4429.e19, 2021 08 19.
Artículo en Inglés | MEDLINE | ID: mdl-34416146

RESUMEN

Alphaviruses are emerging, mosquito-transmitted pathogens that cause musculoskeletal and neurological disease in humans. Although neutralizing antibodies that inhibit individual alphaviruses have been described, broadly reactive antibodies that protect against both arthritogenic and encephalitic alphaviruses have not been reported. Here, we identify DC2.112 and DC2.315, two pan-protective yet poorly neutralizing human monoclonal antibodies (mAbs) that avidly bind to viral antigen on the surface of cells infected with arthritogenic and encephalitic alphaviruses. These mAbs engage a conserved epitope in domain II of the E1 protein proximal to and within the fusion peptide. Treatment with DC2.112 or DC2.315 protects mice against infection by both arthritogenic (chikungunya and Mayaro) and encephalitic (Venezuelan, Eastern, and Western equine encephalitis) alphaviruses through multiple mechanisms, including inhibition of viral egress and monocyte-dependent Fc effector functions. These findings define a conserved epitope recognized by weakly neutralizing yet protective antibodies that could be targeted for pan-alphavirus immunotherapy and vaccine design.


Asunto(s)
Alphavirus/inmunología , Anticuerpos Antivirales/inmunología , Secuencia Conservada/inmunología , Epítopos/inmunología , Proteínas Virales/inmunología , Infecciones por Alphavirus/inmunología , Infecciones por Alphavirus/virología , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales/inmunología , Fiebre Chikungunya/inmunología , Fiebre Chikungunya/virología , Virus Chikungunya/inmunología , Chlorocebus aethiops , Mapeo Epitopo , Epítopos/química , Humanos , Masculino , Ratones Endogámicos C57BL , Modelos Biológicos , Monocitos/metabolismo , Células Vero , Proteínas Virales/química , Liberación del Virus
3.
Cell ; 184(17): 4430-4446.e22, 2021 08 19.
Artículo en Inglés | MEDLINE | ID: mdl-34416147

RESUMEN

Alphaviruses cause severe arthritogenic or encephalitic disease. The E1 structural glycoprotein is highly conserved in these viruses and mediates viral fusion with host cells. However, the role of antibody responses to the E1 protein in immunity is poorly understood. We isolated E1-specific human monoclonal antibodies (mAbs) with diverse patterns of recognition for alphaviruses (ranging from Eastern equine encephalitis virus [EEEV]-specific to alphavirus cross-reactive) from survivors of natural EEEV infection. Antibody binding patterns and epitope mapping experiments identified differences in E1 reactivity based on exposure of epitopes on the glycoprotein through pH-dependent mechanisms or presentation on the cell surface prior to virus egress. Therapeutic efficacy in vivo of these mAbs corresponded with potency of virus egress inhibition in vitro and did not require Fc-mediated effector functions for treatment against subcutaneous EEEV challenge. These studies reveal the molecular basis for broad and protective antibody responses to alphavirus E1 proteins.


Asunto(s)
Alphavirus/inmunología , Anticuerpos Antivirales/inmunología , Reacciones Cruzadas/inmunología , Proteínas Virales/inmunología , Liberación del Virus/fisiología , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/aislamiento & purificación , Anticuerpos Neutralizantes/inmunología , Antígenos Virales/inmunología , Línea Celular , Virus Chikungunya/inmunología , Virus de la Encefalitis Equina del Este/inmunología , Encefalomielitis Equina/inmunología , Encefalomielitis Equina/virología , Mapeo Epitopo , Femenino , Caballos , Humanos , Concentración de Iones de Hidrógeno , Articulaciones/patología , Masculino , Ratones Endogámicos C57BL , Modelos Biológicos , Unión Proteica , ARN Viral/metabolismo , Receptores Fc/metabolismo , Temperatura , Virión/metabolismo , Internalización del Virus
4.
Cell ; 184(7): 1804-1820.e16, 2021 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-33691139

RESUMEN

SARS-CoV-2 has caused the global COVID-19 pandemic. Although passively delivered neutralizing antibodies against SARS-CoV-2 show promise in clinical trials, their mechanism of action in vivo is incompletely understood. Here, we define correlates of protection of neutralizing human monoclonal antibodies (mAbs) in SARS-CoV-2-infected animals. Whereas Fc effector functions are dispensable when representative neutralizing mAbs are administered as prophylaxis, they are required for optimal protection as therapy. When given after infection, intact mAbs reduce SARS-CoV-2 burden and lung disease in mice and hamsters better than loss-of-function Fc variant mAbs. Fc engagement of neutralizing antibodies mitigates inflammation and improves respiratory mechanics, and transcriptional profiling suggests these phenotypes are associated with diminished innate immune signaling and preserved tissue repair. Immune cell depletions establish that neutralizing mAbs require monocytes and CD8+ T cells for optimal clinical and virological benefit. Thus, potently neutralizing mAbs utilize Fc effector functions during therapy to mitigate lung infection and disease.


Asunto(s)
Anticuerpos Monoclonales , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Linfocitos T CD8-positivos , COVID-19 , Fragmentos Fc de Inmunoglobulinas/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/uso terapéutico , Anticuerpos Antivirales/inmunología , Anticuerpos Antivirales/uso terapéutico , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Células CHO , COVID-19/inmunología , COVID-19/terapia , Chlorocebus aethiops , Cricetulus , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , SARS-CoV-2/inmunología , Células Vero , Carga Viral
5.
Cell ; 182(4): 901-918.e18, 2020 08 20.
Artículo en Inglés | MEDLINE | ID: mdl-32668198

RESUMEN

Chikungunya virus (CHIKV), an emerging alphavirus, has infected millions of people. However, the factors modulating disease outcome remain poorly understood. Here, we show in germ-free mice or in oral antibiotic-treated conventionally housed mice with depleted intestinal microbiomes that greater CHIKV infection and spread occurs within 1 day of virus inoculation. Alteration of the microbiome alters TLR7-MyD88 signaling in plasmacytoid dendritic cells (pDCs) and blunts systemic production of type I interferon (IFN). Consequently, circulating monocytes express fewer IFN-stimulated genes and become permissive for CHIKV infection. Reconstitution with a single bacterial species, Clostridium scindens, or its derived metabolite, the secondary bile acid deoxycholic acid, can restore pDC- and MyD88-dependent type I IFN responses to restrict systemic CHIKV infection and transmission back to vector mosquitoes. Thus, symbiotic intestinal bacteria modulate antiviral immunity and levels of circulating alphaviruses within hours of infection through a bile acid-pDC-IFN signaling axis, which affects viremia, dissemination, and potentially transmission.


Asunto(s)
Ácidos y Sales Biliares/metabolismo , Fiebre Chikungunya/patología , Microbioma Gastrointestinal , Interferón Tipo I/metabolismo , Animales , Antibacterianos/farmacología , Fiebre Chikungunya/inmunología , Fiebre Chikungunya/veterinaria , Virus Chikungunya/genética , Virus Chikungunya/aislamiento & purificación , Clostridiales/fisiología , Células Dendríticas/citología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Trasplante de Microbiota Fecal , Microbioma Gastrointestinal/efectos de los fármacos , Masculino , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/citología , Monocitos/inmunología , Monocitos/metabolismo , Factor 88 de Diferenciación Mieloide/deficiencia , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , ARN Viral/sangre , Factor de Transcripción STAT1/deficiencia , Transducción de Señal , Receptor Toll-Like 7/metabolismo
6.
Cell ; 182(3): 744-753.e4, 2020 08 06.
Artículo en Inglés | MEDLINE | ID: mdl-32553273

RESUMEN

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic with millions of human infections. One limitation to the evaluation of potential therapies and vaccines to inhibit SARS-CoV-2 infection and ameliorate disease is the lack of susceptible small animals in large numbers. Commercially available laboratory strains of mice are not readily infected by SARS-CoV-2 because of species-specific differences in their angiotensin-converting enzyme 2 (ACE2) receptors. Here, we transduced replication-defective adenoviruses encoding human ACE2 via intranasal administration into BALB/c mice and established receptor expression in lung tissues. hACE2-transduced mice were productively infected with SARS-CoV-2, and this resulted in high viral titers in the lung, lung pathology, and weight loss. Passive transfer of a neutralizing monoclonal antibody reduced viral burden in the lung and mitigated inflammation and weight loss. The development of an accessible mouse model of SARS-CoV-2 infection and pathogenesis will expedite the testing and deployment of therapeutics and vaccines.


Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Neutralizantes/uso terapéutico , Anticuerpos Antivirales/uso terapéutico , Betacoronavirus/inmunología , Infecciones por Coronavirus/terapia , Modelos Animales de Enfermedad , Neumonía Viral/terapia , Enzima Convertidora de Angiotensina 2 , Animales , COVID-19 , Chlorocebus aethiops , Infecciones por Coronavirus/virología , Femenino , Células HEK293 , Humanos , Inmunización Pasiva/métodos , Pulmón/metabolismo , Pulmón/virología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Noqueados , Pandemias , Peptidil-Dipeptidasa A/genética , Peptidil-Dipeptidasa A/metabolismo , Neumonía Viral/virología , SARS-CoV-2 , Transducción Genética , Células Vero , Carga Viral/inmunología
7.
Cell ; 183(1): 169-184.e13, 2020 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-32931734

RESUMEN

The coronavirus disease 2019 pandemic has made deployment of an effective vaccine a global health priority. We evaluated the protective activity of a chimpanzee adenovirus-vectored vaccine encoding a prefusion stabilized spike protein (ChAd-SARS-CoV-2-S) in challenge studies with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and mice expressing the human angiotensin-converting enzyme 2 receptor. Intramuscular dosing of ChAd-SARS-CoV-2-S induces robust systemic humoral and cell-mediated immune responses and protects against lung infection, inflammation, and pathology but does not confer sterilizing immunity, as evidenced by detection of viral RNA and induction of anti-nucleoprotein antibodies after SARS-CoV-2 challenge. In contrast, a single intranasal dose of ChAd-SARS-CoV-2-S induces high levels of neutralizing antibodies, promotes systemic and mucosal immunoglobulin A (IgA) and T cell responses, and almost entirely prevents SARS-CoV-2 infection in both the upper and lower respiratory tracts. Intranasal administration of ChAd-SARS-CoV-2-S is a candidate for preventing SARS-CoV-2 infection and transmission and curtailing pandemic spread.


Asunto(s)
Infecciones por Coronavirus/inmunología , Inmunogenicidad Vacunal , Neumonía Viral/inmunología , Vacunas Virales/inmunología , Adenoviridae/genética , Administración Intranasal , Animales , Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , COVID-19 , Vacunas contra la COVID-19 , Chlorocebus aethiops , Infecciones por Coronavirus/patología , Infecciones por Coronavirus/prevención & control , Femenino , Células HEK293 , Humanos , Inyecciones Intramusculares , Ratones , Ratones Endogámicos BALB C , Pandemias , Neumonía Viral/patología , Mucosa Respiratoria/inmunología , Mucosa Respiratoria/patología , Mucosa Respiratoria/virología , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología , Células Vero , Vacunas Virales/administración & dosificación
8.
Nat Immunol ; 21(11): 1327-1335, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-32839612

RESUMEN

Although animal models have been evaluated for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, none have fully recapitulated the lung disease phenotypes seen in humans who have been hospitalized. Here, we evaluate transgenic mice expressing the human angiotensin I-converting enzyme 2 (ACE2) receptor driven by the cytokeratin-18 (K18) gene promoter (K18-hACE2) as a model of SARS-CoV-2 infection. Intranasal inoculation of SARS-CoV-2 in K18-hACE2 mice results in high levels of viral infection in lungs, with spread to other organs. A decline in pulmonary function occurs 4 days after peak viral titer and correlates with infiltration of monocytes, neutrophils and activated T cells. SARS-CoV-2-infected lung tissues show a massively upregulated innate immune response with signatures of nuclear factor-κB-dependent, type I and II interferon signaling, and leukocyte activation pathways. Thus, the K18-hACE2 model of SARS-CoV-2 infection shares many features of severe COVID-19 infection and can be used to define the basis of lung disease and test immune and antiviral-based countermeasures.


Asunto(s)
Betacoronavirus/inmunología , Infecciones por Coronavirus/patología , Inmunidad Innata/inmunología , Peptidil-Dipeptidasa A/genética , Neumonía Viral/patología , Neumonía/patología , Enzima Convertidora de Angiotensina 2 , Animales , COVID-19 , Chlorocebus aethiops , Infecciones por Coronavirus/inmunología , Modelos Animales de Enfermedad , Femenino , Humanos , Interferón Tipo I/inmunología , Interferón gamma/inmunología , Queratina-18/genética , Leucocitos/inmunología , Activación de Linfocitos/inmunología , Masculino , Ratones , Ratones Transgénicos , Monocitos/inmunología , FN-kappa B/inmunología , Infiltración Neutrófila/inmunología , Neutrófilos/inmunología , Pandemias , Neumonía/genética , Neumonía/virología , Neumonía Viral/inmunología , Regiones Promotoras Genéticas/genética , SARS-CoV-2 , Linfocitos T/inmunología , Células Vero , Replicación Viral/inmunología
10.
Nat Immunol ; 20(10): 1291-1298, 2019 10.
Artículo en Inglés | MEDLINE | ID: mdl-31477918

RESUMEN

Infections with dengue virus (DENV) and Zika virus (ZIKV) can induce cross-reactive antibody responses. Two immunodominant epitopes-one to precursor membrane protein and one to the fusion loop epitope on envelope (E) protein-are recognized by cross-reactive antibodies1-3 that are not only poorly neutralizing, but can also promote increased viral replication and disease severity via Fcγ receptor-mediated infection of myeloid cells-a process termed antibody-dependent enhancement (ADE)1,4,5. ADE is a significant concern for both ZIKV and DENV vaccines as the induction of poorly neutralizing cross-reactive antibodies may prime an individual for ADE on natural infection. In this report, we describe the design and production of covalently stabilized ZIKV E dimers, which lack precursor membrane protein and do not expose the immunodominant fusion loop epitope. Immunization of mice with ZIKV E dimers induces dimer-specific antibodies, which protect against ZIKV challenge during pregnancy. Importantly, the ZIKV E-dimer-induced response does not cross-react with DENV or induce ADE of DENV infection.


Asunto(s)
Virus del Dengue/fisiología , Dengue/inmunología , Vacunas Virales/inmunología , Infección por el Virus Zika/inmunología , Virus Zika/fisiología , Animales , Anticuerpos Neutralizantes/metabolismo , Anticuerpos Antivirales/metabolismo , Reacciones Cruzadas , Dimerización , Epítopos/genética , Femenino , Ingeniería Genética , Células HEK293 , Humanos , Epítopos Inmunodominantes/genética , Ratones , Ratones Endogámicos BALB C , Receptores de IgG/metabolismo , Vacunación , Proteínas del Envoltorio Viral/genética , Vacunas Virales/genética , Replicación Viral
11.
Immunity ; 54(10): 2399-2416.e6, 2021 10 12.
Artículo en Inglés | MEDLINE | ID: mdl-34481543

RESUMEN

With the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants with increased transmissibility and potential resistance, antibodies and vaccines with broadly inhibitory activity are needed. Here, we developed a panel of neutralizing anti-SARS-CoV-2 monoclonal antibodies (mAbs) that bound the receptor binding domain of the spike protein at distinct epitopes and blocked virus attachment to its host receptor, human angiotensin converting enzyme-2 (hACE2). Although several potently neutralizing mAbs protected K18-hACE2 transgenic mice against infection caused by ancestral SARS-CoV-2 strains, others induced escape variants in vivo or lost neutralizing activity against emerging strains. One mAb, SARS2-38, potently neutralized all tested SARS-CoV-2 variants of concern and protected mice against challenge by multiple SARS-CoV-2 strains. Structural analysis showed that SARS2-38 engaged a conserved epitope proximal to the receptor binding motif. Thus, treatment with or induction of neutralizing antibodies that bind conserved spike epitopes may limit the loss of potency of therapies or vaccines against emerging SARS-CoV-2 variants.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Epítopos/inmunología , SARS-CoV-2/inmunología , Secuencias de Aminoácidos , Animales , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Neutralizantes/química , Anticuerpos Neutralizantes/uso terapéutico , COVID-19/prevención & control , COVID-19/virología , Epítopos/química , Epítopos/metabolismo , Humanos , Cadenas Ligeras de Inmunoglobulina/química , Cadenas Ligeras de Inmunoglobulina/metabolismo , Ratones , Pruebas de Neutralización , Dominios Proteicos , SARS-CoV-2/genética , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología
12.
Immunity ; 54(6): 1290-1303.e7, 2021 06 08.
Artículo en Inglés | MEDLINE | ID: mdl-34022127

RESUMEN

Dissecting the evolution of memory B cells (MBCs) against SARS-CoV-2 is critical for understanding antibody recall upon secondary exposure. Here, we used single-cell sequencing to profile SARS-CoV-2-reactive B cells in 38 COVID-19 patients. Using oligo-tagged antigen baits, we isolated B cells specific to the SARS-CoV-2 spike, nucleoprotein (NP), open reading frame 8 (ORF8), and endemic human coronavirus (HCoV) spike proteins. SARS-CoV-2 spike-specific cells were enriched in the memory compartment of acutely infected and convalescent patients several months post symptom onset. With severe acute infection, substantial populations of endemic HCoV-reactive antibody-secreting cells were identified and possessed highly mutated variable genes, signifying preexisting immunity. Finally, MBCs exhibited pronounced maturation to NP and ORF8 over time, especially in older patients. Monoclonal antibodies against these targets were non-neutralizing and non-protective in vivo. These findings reveal antibody adaptation to non-neutralizing intracellular antigens during infection, emphasizing the importance of vaccination for inducing neutralizing spike-specific MBCs.


Asunto(s)
Anticuerpos Antivirales/inmunología , Formación de Anticuerpos/inmunología , Linfocitos B/inmunología , COVID-19/inmunología , Interacciones Huésped-Patógeno/inmunología , Epítopos Inmunodominantes/inmunología , SARS-CoV-2/inmunología , Anticuerpos Neutralizantes/inmunología , Formación de Anticuerpos/genética , Linfocitos B/metabolismo , Biología Computacional/métodos , Reacciones Cruzadas/inmunología , Mapeo Epitopo , Femenino , Perfilación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Interacciones Huésped-Patógeno/genética , Humanos , Epítopos Inmunodominantes/genética , Memoria Inmunológica , Masculino , Pruebas de Neutralización , Análisis de la Célula Individual/métodos , Glicoproteína de la Espiga del Coronavirus/inmunología , Transcriptoma
13.
Nature ; 591(7849): 293-299, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33494095

RESUMEN

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-a new coronavirus that has led to a worldwide pandemic1-has a furin cleavage site (PRRAR) in its spike protein that is absent in other group-2B coronaviruses2. To explore whether the furin cleavage site contributes to infection and pathogenesis in this virus, we generated a mutant SARS-CoV-2 that lacks the furin cleavage site (ΔPRRA). Here we report that replicates of ΔPRRA SARS-CoV-2 had faster kinetics, improved fitness in Vero E6 cells and reduced spike protein processing, as compared to parental SARS-CoV-2. However, the ΔPRRA mutant had reduced replication in a human respiratory cell line and was attenuated in both hamster and K18-hACE2 transgenic mouse models of SARS-CoV-2 pathogenesis. Despite reduced disease, the ΔPRRA mutant conferred protection against rechallenge with the parental SARS-CoV-2. Importantly, the neutralization values of sera from patients with coronavirus disease 2019 (COVID-19) and monoclonal antibodies against the receptor-binding domain of SARS-CoV-2 were lower against the ΔPRRA mutant than against parental SARS-CoV-2, probably owing to an increased ratio of particles to plaque-forming units in infections with the former. Together, our results demonstrate a critical role for the furin cleavage site in infection with SARS-CoV-2 and highlight the importance of this site for evaluating the neutralization activities of antibodies.


Asunto(s)
COVID-19/virología , Furina/metabolismo , Mutación , SARS-CoV-2/genética , SARS-CoV-2/patogenicidad , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Secuencia de Aminoácidos , Animales , Anticuerpos Neutralizantes/inmunología , COVID-19/patología , COVID-19/fisiopatología , Línea Celular , Chlorocebus aethiops , Cricetinae , Femenino , Humanos , Enfermedades Pulmonares/patología , Enfermedades Pulmonares/fisiopatología , Enfermedades Pulmonares/virología , Masculino , Ratones , Ratones Transgénicos , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteolisis , SARS-CoV-2/química , SARS-CoV-2/metabolismo , Serina Endopeptidasas/metabolismo , Glicoproteína de la Espiga del Coronavirus/metabolismo , Células Vero , Replicación Viral/genética
14.
Nature ; 596(7870): 103-108, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34153975

RESUMEN

Rapidly emerging SARS-CoV-2 variants jeopardize antibody-based countermeasures. Although cell culture experiments have demonstrated a loss of potency of several anti-spike neutralizing antibodies against variant strains of SARS-CoV-21-3, the in vivo importance of these results remains uncertain. Here we report the in vitro and in vivo activity of a panel of monoclonal antibodies (mAbs), which correspond to many in advanced clinical development by Vir Biotechnology, AbbVie, AstraZeneca, Regeneron and Lilly, against SARS-CoV-2 variant viruses. Although some individual mAbs showed reduced or abrogated neutralizing activity in cell culture against B.1.351, B.1.1.28, B.1.617.1 and B.1.526 viruses with mutations at residue E484 of the spike protein, low prophylactic doses of mAb combinations protected against infection by many variants in K18-hACE2 transgenic mice, 129S2 immunocompetent mice and hamsters, without the emergence of resistance. Exceptions were LY-CoV555 monotherapy and LY-CoV555 and LY-CoV016 combination therapy, both of which lost all protective activity, and the combination of AbbVie 2B04 and 47D11, which showed a partial loss of activity. When administered after infection, higher doses of several mAb cocktails protected in vivo against viruses with a B.1.351 spike gene. Therefore, many-but not all-of the antibody products with Emergency Use Authorization should retain substantial efficacy against the prevailing variant strains of SARS-CoV-2.


Asunto(s)
Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Antivirales/farmacología , Anticuerpos Antivirales/uso terapéutico , COVID-19/virología , Pruebas de Neutralización , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/inmunología , Enzima Convertidora de Angiotensina 2/genética , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/farmacología , Anticuerpos Neutralizantes/uso terapéutico , Anticuerpos Antivirales/inmunología , COVID-19/genética , COVID-19/inmunología , COVID-19/prevención & control , Chlorocebus aethiops , Femenino , Humanos , Masculino , Mesocricetus/inmunología , Mesocricetus/virología , Ratones , Ratones Transgénicos , Profilaxis Posexposición , Profilaxis Pre-Exposición , SARS-CoV-2/genética , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología , Células Vero
15.
Nature ; 584(7821): 443-449, 2020 08.
Artículo en Inglés | MEDLINE | ID: mdl-32668443

RESUMEN

The ongoing pandemic of coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a major threat to global health1 and the medical countermeasures available so far are limited2,3. Moreover, we currently lack a thorough understanding of the mechanisms of humoral immunity to SARS-CoV-24. Here we analyse a large panel of human monoclonal antibodies that target the spike (S) glycoprotein5, and identify several that exhibit potent neutralizing activity and fully block the receptor-binding domain of the S protein (SRBD) from interacting with human angiotensin-converting enzyme 2 (ACE2). Using competition-binding, structural and functional studies, we show that the monoclonal antibodies can be clustered into classes that recognize distinct epitopes on the SRBD, as well as distinct conformational states of the S trimer. Two potently neutralizing monoclonal antibodies, COV2-2196 and COV2-2130, which recognize non-overlapping sites, bound simultaneously to the S protein and neutralized wild-type SARS-CoV-2 virus in a synergistic manner. In two mouse models of SARS-CoV-2 infection, passive transfer of COV2-2196, COV2-2130 or a combination of both of these antibodies protected mice from weight loss and reduced the viral burden and levels of inflammation in the lungs. In addition, passive transfer of either of two of the most potent ACE2-blocking monoclonal antibodies (COV2-2196 or COV2-2381) as monotherapy protected rhesus macaques from SARS-CoV-2 infection. These results identify protective epitopes on the SRBD and provide a structure-based framework for rational vaccine design and the selection of robust immunotherapeutic agents.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Betacoronavirus/inmunología , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/prevención & control , Pandemias/prevención & control , Neumonía Viral/inmunología , Neumonía Viral/prevención & control , Enzima Convertidora de Angiotensina 2 , Animales , Anticuerpos Monoclonales/inmunología , Betacoronavirus/química , Unión Competitiva , COVID-19 , Línea Celular , Reacciones Cruzadas , Modelos Animales de Enfermedad , Epítopos de Linfocito B/química , Epítopos de Linfocito B/inmunología , Femenino , Humanos , Macaca mulatta , Masculino , Ratones , Persona de Mediana Edad , Pruebas de Neutralización , Peptidil-Dipeptidasa A/genética , Peptidil-Dipeptidasa A/metabolismo , Profilaxis Pre-Exposición , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/química , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/inmunología , SARS-CoV-2 , Síndrome Respiratorio Agudo Grave/inmunología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/metabolismo
16.
J Virol ; 96(1): e0151121, 2022 01 12.
Artículo en Inglés | MEDLINE | ID: mdl-34668780

RESUMEN

The development of mouse models for coronavirus disease 2019 (COVID-19) has enabled testing of vaccines and therapeutics and defining aspects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pathogenesis. SARS-CoV-2 disease is severe in K18 transgenic mice (K18-hACE2 Tg) expressing human angiotensin-converting enzyme 2 (hACE2), the SARS-CoV-2 receptor, under an ectopic cytokeratin promoter, with high levels of infection measured in the lung and brain. Here, we evaluated SARS-CoV-2 infection in hACE2 knock-in (KI) mice that express hACE2 under an endogenous promoter in place of murine ACE2 (mACE2). Intranasal inoculation of hACE2 KI mice with SARS-CoV-2 WA1/2020 resulted in substantial viral replication within the upper and lower respiratory tracts with limited spread to extrapulmonary organs. However, SARS-CoV-2-infected hACE2 KI mice did not lose weight and developed limited pathology. Moreover, no significant differences in viral burden were observed in hACE2 KI mice infected with B.1.1.7 or B.1.351 variants compared to the WA1/2020 strain. Because the entry mechanisms of SARS-CoV-2 in mice remain uncertain, we evaluated the impact of the naturally occurring, mouse-adapting N501Y mutation by comparing infection of hACE2 KI, K18-hACE2 Tg, ACE2-deficient, and wild-type C57BL/6 mice. The N501Y mutation minimally affected SARS-CoV-2 infection in hACE2 KI mice but was required for viral replication in wild-type C57BL/6 mice in a mACE2-dependent manner and augmented pathogenesis in the K18-hACE2 Tg mice. Thus, the N501Y mutation likely enhances interactions with mACE2 or hACE2 in vivo. Overall, our study highlights the hACE2 KI mice as a model of mild SARS-CoV-2 infection and disease and clarifies the requirement of the N501Y mutation in mice. IMPORTANCE Mouse models of SARS-CoV-2 pathogenesis have facilitated the rapid evaluation of countermeasures. While the first generation of models developed pneumonia and severe disease after SARS-CoV-2 infection, they relied on ectopic expression of supraphysiological levels of human ACE2 (hACE2). This has raised issues with their relevance to humans, as the hACE2 receptor shows a more restricted expression pattern in the respiratory tract. Here, we evaluated SARS-CoV-2 infection and disease with viruses containing or lacking a key mouse-adapting mutation in the spike gene in hACE2 KI mice, which express hACE2 under an endogenous promoter in place of murine ACE2. While infection of hACE2 KI mice with multiple strains of SARS-CoV-2 including variants of concern resulted in viral replication within the upper and lower respiratory tracts, the animals did not sustain severe lung injury. Thus, hACE2 KI mice serve as a model of mild infection with both ancestral and emerging SARS-CoV-2 variant strains.


Asunto(s)
Enzima Convertidora de Angiotensina 2/genética , COVID-19/virología , Pulmón/virología , SARS-CoV-2/patogenicidad , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , COVID-19/patología , Modelos Animales de Enfermedad , Expresión Génica , Técnicas de Sustitución del Gen , Humanos , Inflamación , Pulmón/metabolismo , Pulmón/patología , Ratones , Ratones Transgénicos , Mutación , SARS-CoV-2/genética , Carga Viral , Replicación Viral
17.
PLoS Pathog ; 16(1): e1008292, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31999809

RESUMEN

Humoral immune responses initiate in the lymph node draining the site of viral infection (dLN). Some viruses subvert LN B cell activation; however, our knowledge of viral hindrance of B cell responses of important human pathogens is lacking. Here, we define mechanisms whereby chikungunya virus (CHIKV), a mosquito-transmitted RNA virus that causes outbreaks of acute and chronic arthritis in humans, hinders dLN antiviral B cell responses. Infection of WT mice with pathogenic, but not acutely cleared CHIKV, induced MyD88-dependent recruitment of monocytes and neutrophils to the dLN. Blocking this influx improved lymphocyte accumulation, dLN organization, and CHIKV-specific B cell responses. Both inducible nitric oxide synthase (iNOS) and the phagocyte NADPH oxidase (Nox2) contributed to impaired dLN organization and function. Infiltrating monocytes expressed iNOS through a local IRF5- and IFNAR1-dependent pathway that was partially TLR7-dependent. Together, our data suggest that pathogenic CHIKV triggers the influx and activation of monocytes and neutrophils in the dLN that impairs virus-specific B cell responses.


Asunto(s)
Linfocitos B/inmunología , Fiebre Chikungunya/inmunología , Factores Reguladores del Interferón/inmunología , Monocitos/inmunología , Factor 88 de Diferenciación Mieloide/inmunología , NADPH Oxidasa 2/inmunología , Neutrófilos/inmunología , Óxido Nítrico Sintasa de Tipo II/inmunología , Animales , Fiebre Chikungunya/virología , Virus Chikungunya/fisiología , Humanos , Factores Reguladores del Interferón/genética , Ganglios Linfáticos/inmunología , Ganglios Linfáticos/virología , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 88 de Diferenciación Mieloide/genética , NADPH Oxidasa 2/genética , Óxido Nítrico Sintasa de Tipo II/genética
18.
J Immunol ; 205(4): 915-922, 2020 08 15.
Artículo en Inglés | MEDLINE | ID: mdl-32591393

RESUMEN

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for millions of infections and hundreds of thousands of deaths globally. There are no widely available licensed therapeutics against SARS-CoV-2, highlighting an urgent need for effective interventions. The virus enters host cells through binding of a receptor-binding domain within its trimeric spike glycoprotein to human angiotensin-converting enzyme 2. In this article, we describe the generation and characterization of a panel of murine mAbs directed against the receptor-binding domain. One mAb, 2B04, neutralized wild-type SARS-CoV-2 in vitro with remarkable potency (half-maximal inhibitory concentration of <2 ng/ml). In a murine model of SARS-CoV-2 infection, 2B04 protected challenged animals from weight loss, reduced lung viral load, and blocked systemic dissemination. Thus, 2B04 is a promising candidate for an effective antiviral that can be used to prevent SARS-CoV-2 infection.


Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Neutralizantes/uso terapéutico , Anticuerpos Antivirales/uso terapéutico , Betacoronavirus/efectos de los fármacos , Infecciones por Coronavirus/tratamiento farmacológico , Infecciones por Coronavirus/inmunología , Neumonía Viral/tratamiento farmacológico , Neumonía Viral/inmunología , Enzima Convertidora de Angiotensina 2 , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/farmacología , Anticuerpos Antivirales/inmunología , Anticuerpos Antivirales/farmacología , COVID-19 , Chlorocebus aethiops , Infecciones por Coronavirus/virología , Modelos Animales de Enfermedad , Mapeo Epitopo , Femenino , Células HEK293 , Humanos , Epítopos Inmunodominantes/inmunología , Ratones , Ratones Endogámicos C57BL , Pandemias , Peptidil-Dipeptidasa A/genética , Peptidil-Dipeptidasa A/metabolismo , Neumonía Viral/virología , Dominios y Motivos de Interacción de Proteínas/genética , Dominios y Motivos de Interacción de Proteínas/inmunología , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/metabolismo , Transfección , Células Vero
19.
J Infect Dis ; 221(9): 1506-1517, 2020 04 07.
Artículo en Inglés | MEDLINE | ID: mdl-31616920

RESUMEN

BACKGROUND: Sexual transmission and persistence of Zika virus (ZIKV) in the male reproductive tract has raised concerned for potential damaging effects on function. Animal studies have demonstrated that ZIKV virus can infect and damage the testis and epididymis, and these results has been correlated to lower sperm counts in ZIKV-infected humans. The prostate plays a vital role in the male reproductive tract, with acute and chronic prostatitis linked to male infertility. METHODS: In this study, we evaluated the effects of ZIKV virus on the prostate in mice and nonhuman primates. RESULTS: In mice, ZIKV infected the prostate and triggered inflammation that persisted even after virus clearance. Evidence of chronic prostatitis associated with ZIKV infection remained for several months. Similar histological findings were observed in the prostate of ZIKV-infected rhesus macaques. CONCLUSIONS: These studies establish that ZIKV replicates in the prostate and can cause acute and chronic inflammatory and proliferative changes in mouse and nonhuman primate models.


Asunto(s)
Prostatitis/virología , Testículo/virología , Infección por el Virus Zika/complicaciones , Enfermedad Aguda , Animales , Enfermedad Crónica , Modelos Animales de Enfermedad , Epidídimo , Macaca mulatta , Masculino , Ratones , Ratones Endogámicos C57BL , Prostatitis/patología , Semen/virología , Testículo/patología , Virus Zika , Infección por el Virus Zika/transmisión
20.
bioRxiv ; 2022 Jan 24.
Artículo en Inglés | MEDLINE | ID: mdl-35118466

RESUMEN

Although vaccines and monoclonal antibody countermeasures have reduced the morbidity and mortality associated with SARS-CoV-2 infection, variants with constellations of mutations in the spike gene threaten their efficacy. Accordingly, antiviral interventions that are resistant to further virus evolution are needed. The host-derived cytokine IFN-λ has been proposed as a possible treatment based on correlative studies in human COVID-19 patients. Here, we show IFN-λ protects against SARS-CoV-2 B.1.351 (Beta) and B.1.1.529 (Omicron)variants in three strains of conventional and human ACE2 transgenic mice. Prophylaxis or therapy with nasally-delivered IFN-λ2 limited infection of historical or variant (B.1.351 and B.1.1.529) SARS-CoV-2 strains in the upper and lower respiratory tracts without causing excessive inflammation. In the lung, IFN-λ was produced preferentially in epithelial cells and acted on radio-resistant cells to protect against of SARS-CoV-2 infection. Thus, inhaled IFN-λ may have promise as a treatment for evolving SARS-CoV-2 variants that develop resistance to antibody-based countermeasures.

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