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1.
Nat Commun ; 11(1): 4994, 2020 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-33020485

RESUMO

Serogroup B meningococcus (MenB) is a leading cause of meningitis and sepsis across the world and vaccination is the most effective way to protect against this disease. 4CMenB is a multi-component vaccine against MenB, which is now licensed for use in subjects >2 months of age in several countries. In this study, we describe the development and use of an ad hoc protein microarray to study the immune response induced by the three major 4CMenB antigenic components (fHbp, NHBA and NadA) in individual sera from vaccinated infants, adolescents and adults. The resulting 4CMenB protein antigen fingerprinting allowed the identification of specific human antibody repertoire correlating with the bactericidal response elicited in each subject. This work represents an example of epitope mapping of the immune response induced by a multicomponent vaccine in different age groups with the identification of protective signatures. It shows the high flexibility of this microarray based methodology in terms of high-throughput information and minimal volume of biological samples needed.


Assuntos
Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Infecções Meningocócicas/imunologia , Vacinas Meningocócicas/imunologia , Neisseria meningitidis Sorogrupo B/imunologia , Adolescente , Adulto , Anticorpos Antibacterianos/sangue , Criança , Pré-Escolar , Mapeamento de Epitopos , Humanos , Lactente , Infecções Meningocócicas/prevenção & controle , Biblioteca de Peptídeos , Análise Serial de Proteínas , Ensaios de Anticorpos Bactericidas Séricos , Adulto Jovem
2.
Cell Mol Immunol ; 17(10): 1095-1097, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32895485
3.
PLoS Pathog ; 16(8): e1008753, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866207

RESUMO

The induction of broad and potent immunity by vaccines is the key focus of research efforts aimed at protecting against HIV-1 infection. Soluble native-like HIV-1 envelope glycoproteins have shown promise as vaccine candidates as they can induce potent autologous neutralizing responses in rabbits and non-human primates. In this study, monoclonal antibodies were isolated and characterized from rhesus macaques immunized with the BG505 SOSIP.664 trimer to better understand vaccine-induced antibody responses. Our studies reveal a diverse landscape of antibodies recognizing immunodominant strain-specific epitopes and non-neutralizing neo-epitopes. Additionally, we isolated a subset of mAbs against an epitope cluster at the gp120-gp41 interface that recognize the highly conserved fusion peptide and the glycan at position 88 and have characteristics akin to several human-derived broadly neutralizing antibodies.


Assuntos
Vacinas contra a AIDS/imunologia , Mapeamento de Epitopos , Epitopos/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , Vacinas contra a AIDS/genética , Animais , Anticorpos Monoclonais Murinos/imunologia , Epitopos/genética , Anticorpos Anti-HIV/genética , Proteína gp41 do Envelope de HIV/genética , HIV-1/genética , Macaca mulatta , Multimerização Proteica/genética , Multimerização Proteica/imunologia
4.
PLoS Biol ; 18(9): e3000821, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32886672

RESUMO

As a novel alternative to established surface display or combinatorial chemistry approaches for the discovery of therapeutic peptides, we present a method for the isolation of small, cysteine-rich domains from bovine antibody ultralong complementarity-determining regions (CDRs). We show for the first time that isolated bovine antibody knob domains can function as autonomous entities by binding antigen outside the confines of the antibody scaffold. This yields antibody fragments so small as to be considered peptides, each stabilised by an intricate, bespoke arrangement of disulphide bonds. For drug discovery, cow immunisations harness the immune system to generate knob domains with affinities in the picomolar to low nanomolar range, orders of magnitude higher than unoptimized peptides from naïve library screening. Using this approach, knob domain peptides that tightly bound Complement component C5 were obtained, at scale, using conventional antibody discovery and peptide purification techniques.


Assuntos
Anticorpos/química , Dissulfetos/isolamento & purificação , Domínios de Imunoglobulina , Fragmentos de Peptídeos/isolamento & purificação , Domínios e Motivos de Interação entre Proteínas , Animais , Anticorpos/imunologia , Anticorpos/metabolismo , Afinidade de Anticorpos , Formação de Anticorpos , Especificidade de Anticorpos , Antígenos/genética , Antígenos/imunologia , Linfócitos B/fisiologia , Bovinos , Complemento C5/química , Complemento C5/genética , Complemento C5/imunologia , Regiões Determinantes de Complementaridade/química , Regiões Determinantes de Complementaridade/genética , Regiões Determinantes de Complementaridade/imunologia , Dissulfetos/química , Dissulfetos/imunologia , Mapeamento de Epitopos/métodos , Humanos , Imunização , Domínios de Imunoglobulina/genética , Modelos Moleculares , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Domínios e Motivos de Interação entre Proteínas/genética
5.
PLoS One ; 15(9): e0238089, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32903266

RESUMO

A novel severe acute respiratory syndrome coronavirus (SARS-CoV-2) is the source of a current pandemic (COVID-19) with devastating consequences in public health and economic stability. Using a peptide array to map the antibody response of plasma from healing patients (12) and heathy patients (6), we identified three immunodominant linear epitopes, two of which correspond to key proteolytic sites on the spike protein (S1/S2 and S2') known to be critical for cellular entry. We show biochemical evidence that plasma positive for the epitope adjacent to the S1/S2 cleavage site inhibits furin-mediated proteolysis of spike.


Assuntos
Infecções por Coronavirus/patologia , Epitopos/química , Pneumonia Viral/patologia , Sequência de Aminoácidos , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Betacoronavirus/imunologia , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/virologia , Mapeamento de Epitopos , Epitopos/sangue , Epitopos/imunologia , Furina/metabolismo , Humanos , Pandemias , Ácidos Nucleicos Peptídicos/química , Peptídeos/química , Pneumonia Viral/virologia , Análise Serial de Proteínas , Estrutura Terciária de Proteína , Proteólise , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Alinhamento de Sequência , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/metabolismo
6.
Nat Commun ; 11(1): 4303, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855401

RESUMO

The novel highly transmissible human coronavirus SARS-CoV-2 is the causative agent of the COVID-19 pandemic. Thus far, there is no approved therapeutic drug specifically targeting this emerging virus. Here we report the isolation and characterization of a panel of human neutralizing monoclonal antibodies targeting the SARS-CoV-2 receptor binding domain (RBD). These antibodies were selected from a phage display library constructed using peripheral circulatory lymphocytes collected from patients at the acute phase of the disease. These neutralizing antibodies are shown to recognize distinct epitopes on the viral spike RBD. A subset of the antibodies exert their inhibitory activity by abrogating binding of the RBD to the human ACE2 receptor. The human monoclonal antibodies described here represent a promising basis for the design of efficient combined post-exposure therapy for SARS-CoV-2 infection.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Betacoronavirus/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Anticorpos Monoclonais/metabolismo , Anticorpos Neutralizantes/metabolismo , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/metabolismo , Betacoronavirus/metabolismo , Chlorocebus aethiops , Mapeamento de Epitopos , Epitopos , Humanos , Biblioteca de Peptídeos , Peptidil Dipeptidase A/metabolismo , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Células Vero
7.
Science ; 370(6512): 89-94, 2020 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-32753554

RESUMO

Many unknowns exist about human immune responses to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus. SARS-CoV-2-reactive CD4+ T cells have been reported in unexposed individuals, suggesting preexisting cross-reactive T cell memory in 20 to 50% of people. However, the source of those T cells has been speculative. Using human blood samples derived before the SARS-CoV-2 virus was discovered in 2019, we mapped 142 T cell epitopes across the SARS-CoV-2 genome to facilitate precise interrogation of the SARS-CoV-2-specific CD4+ T cell repertoire. We demonstrate a range of preexisting memory CD4+ T cells that are cross-reactive with comparable affinity to SARS-CoV-2 and the common cold coronaviruses human coronavirus (HCoV)-OC43, HCoV-229E, HCoV-NL63, and HCoV-HKU1. Thus, variegated T cell memory to coronaviruses that cause the common cold may underlie at least some of the extensive heterogeneity observed in coronavirus disease 2019 (COVID-19) disease.


Assuntos
Betacoronavirus/imunologia , Linfócitos T CD4-Positivos/imunologia , Infecções por Coronavirus/imunologia , Epitopos de Linfócito T/imunologia , Memória Imunológica , Pneumonia Viral/imunologia , Betacoronavirus/genética , Doadores de Sangue , Reações Cruzadas , Mapeamento de Epitopos , Epitopos de Linfócito T/genética , Genoma Viral , Humanos , Fases de Leitura Aberta , Pandemias , Homologia de Sequência
8.
Cell Rep ; 32(3): 107918, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32668215

RESUMO

Coronavirus disease 2019 (COVID-19) has become a worldwide threat to humans, and neutralizing antibodies have therapeutic potential. We have purified more than 1,000 memory B cells specific to SARS-CoV-2 S1 or its RBD (receptor binding domain) and obtain 729 paired heavy- and light-chain fragments. Among these, 178 antibodies test positive for antigen binding, and the majority of the top 17 binders with EC50 below 1 nM are RBD binders. Furthermore, we identify 11 neutralizing antibodies, eight of which show IC50 within 10 nM, and the best one, 414-1, with IC50 of 1.75 nM. Through epitope mapping, we find three main epitopes in RBD recognized by these antibodies, and epitope-B antibody 553-15 could substantially enhance the neutralizing abilities of most of the other antibodies. We also find that 515-5 could cross neutralize the SARS-CoV pseudovirus. Altogether, our study provides 11 potent human neutralizing antibodies for COVID-19 as therapeutic candidates.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Betacoronavirus/imunologia , Receptores Virais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Monoclonais/uso terapêutico , Linfócitos B/imunologia , Infecções por Coronavirus/terapia , Mapeamento de Epitopos , Epitopos/imunologia , Humanos , Memória Imunológica/imunologia , Testes de Neutralização , Pandemias , Pneumonia Viral/terapia , Domínios Proteicos/imunologia
9.
Science ; 369(6510): 1505-1509, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32703908

RESUMO

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in an unprecedented public health crisis. There are no approved vaccines or therapeutics for treating COVID-19. Here we report a humanized monoclonal antibody, H014, that efficiently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2 at nanomolar concentrations by engaging the spike (S) receptor binding domain (RBD). H014 administration reduced SARS-CoV-2 titers in infected lungs and prevented pulmonary pathology in a human angiotensin-converting enzyme 2 mouse model. Cryo-electron microscopy characterization of the SARS-CoV-2 S trimer in complex with the H014 Fab fragment unveiled a previously uncharacterized conformational epitope, which was only accessible when the RBD was in an open conformation. Biochemical, cellular, virological, and structural studies demonstrated that H014 prevents attachment of SARS-CoV-2 to its host cell receptors. Epitope analysis of available neutralizing antibodies against SARS-CoV and SARS-CoV-2 uncovered broad cross-protective epitopes. Our results highlight a key role for antibody-based therapeutic interventions in the treatment of COVID-19.


Assuntos
Anticorpos Monoclonais Humanizados/química , Anticorpos Neutralizantes/química , Betacoronavirus/imunologia , Peptidil Dipeptidase A/imunologia , Receptores Virais/imunologia , Vírus da SARS/imunologia , Animais , Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Neutralizantes/uso terapêutico , Chlorocebus aethiops , Infecções por Coronavirus/terapia , Mapeamento de Epitopos , Humanos , Fragmentos Fab das Imunoglobulinas/química , Pulmão/imunologia , Camundongos , Pandemias , Pneumonia Viral/terapia , Domínios Proteicos , Multimerização Proteica , Células Vero
10.
Cell ; 182(4): 828-842.e16, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32645326

RESUMO

Neutralizing antibody responses to coronaviruses mainly target the receptor-binding domain (RBD) of the trimeric spike. Here, we characterized polyclonal immunoglobulin Gs (IgGs) and Fabs from COVID-19 convalescent individuals for recognition of coronavirus spikes. Plasma IgGs differed in their focus on RBD epitopes, recognition of alpha- and beta-coronaviruses, and contributions of avidity to increased binding/neutralization of IgGs over Fabs. Using electron microscopy, we examined specificities of polyclonal plasma Fabs, revealing recognition of both S1A and RBD epitopes on SARS-CoV-2 spike. Moreover, a 3.4 Å cryo-electron microscopy (cryo-EM) structure of a neutralizing monoclonal Fab-spike complex revealed an epitope that blocks ACE2 receptor binding. Modeling based on these structures suggested different potentials for inter-spike crosslinking by IgGs on viruses, and characterized IgGs would not be affected by identified SARS-CoV-2 spike mutations. Overall, our studies structurally define a recurrent anti-SARS-CoV-2 antibody class derived from VH3-53/VH3-66 and similarity to a SARS-CoV VH3-30 antibody, providing criteria for evaluating vaccine-elicited antibodies.


Assuntos
Anticorpos Neutralizantes/química , Betacoronavirus/química , Infecções por Coronavirus/imunologia , Fragmentos Fab das Imunoglobulinas/química , Imunoglobulina G/química , Pneumonia Viral/imunologia , Glicoproteína da Espícula de Coronavírus/química , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/isolamento & purificação , Betacoronavirus/imunologia , Infecções por Coronavirus/sangue , Infecções por Coronavirus/terapia , Reações Cruzadas , Microscopia Crioeletrônica , Mapeamento de Epitopos , Epitopos , Humanos , Imunização Passiva , Fragmentos Fab das Imunoglobulinas/sangue , Fragmentos Fab das Imunoglobulinas/isolamento & purificação , Fragmentos Fab das Imunoglobulinas/ultraestrutura , Imunoglobulina G/sangue , Imunoglobulina G/isolamento & purificação , Imunoglobulina G/ultraestrutura , Coronavírus da Síndrome Respiratória do Oriente Médio/química , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Modelos Moleculares , Pandemias , Pneumonia Viral/sangue , Vírus da SARS/química , Vírus da SARS/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia
11.
Nature ; 584(7821): 450-456, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32698192

RESUMO

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic continues, with devasting consequences for human lives and the global economy1,2. The discovery and development of virus-neutralizing monoclonal antibodies could be one approach to treat or prevent infection by this coronavirus. Here we report the isolation of sixty-one SARS-CoV-2-neutralizing monoclonal antibodies from five patients infected with SARS-CoV-2 and admitted to hospital with severe coronavirus disease 2019 (COVID-19). Among these are nineteen antibodies that potently neutralized authentic SARS-CoV-2 in vitro, nine of which exhibited very high potency, with 50% virus-inhibitory concentrations of 0.7 to 9 ng ml-1. Epitope mapping showed that this collection of nineteen antibodies was about equally divided between those directed against the receptor-binding domain (RBD) and those directed against the N-terminal domain (NTD), indicating that both of these regions at the top of the viral spike are immunogenic. In addition, two other powerful neutralizing antibodies recognized quaternary epitopes that overlap with the domains at the top of the spike. Cryo-electron microscopy reconstructions of one antibody that targets the RBD, a second that targets the NTD, and a third that bridges two separate RBDs showed that the antibodies recognize the closed, 'all RBD-down' conformation of the spike. Several of these monoclonal antibodies are promising candidates for clinical development as potential therapeutic and/or prophylactic agents against SARS-CoV-2.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Epitopos de Linfócito B/imunologia , Pneumonia Viral/imunologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Anticorpos Monoclonais/análise , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/ultraestrutura , Anticorpos Neutralizantes/análise , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/ultraestrutura , Anticorpos Antivirais/análise , Anticorpos Antivirais/química , Anticorpos Antivirais/ultraestrutura , Betacoronavirus/química , Betacoronavirus/ultraestrutura , Infecções por Coronavirus/prevenção & controle , Microscopia Crioeletrônica , Modelos Animais de Doenças , Mapeamento de Epitopos , Epitopos de Linfócito B/química , Epitopos de Linfócito B/ultraestrutura , Feminino , Humanos , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/ultraestrutura , Pulmão/patologia , Pulmão/virologia , Masculino , Mesocricetus , Modelos Moleculares , Testes de Neutralização , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Glicoproteína da Espícula de Coronavírus/ultraestrutura
12.
J Immunol ; 205(4): 915-922, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32591393

RESUMO

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.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/farmacologia , Chlorocebus aethiops , Infecções por Coronavirus/virologia , Modelos Animais de Doenças , Mapeamento de Epitopos , Feminino , Células HEK293 , Humanos , Epitopos Imunodominantes/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Pandemias , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/virologia , Domínios e Motivos de Interação entre Proteínas/genética , Domínios e Motivos de Interação entre Proteínas/imunologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Transfecção , Células Vero
13.
Nat Commun ; 11(1): 2601, 2020 05 20.
Artigo em Inglês | MEDLINE | ID: covidwho-326048

RESUMO

The coronavirus family member, SARS-CoV-2 has been identified as the causal agent for the pandemic viral pneumonia disease, COVID-19. At this time, no vaccine is available to control further dissemination of the disease. We have previously engineered a synthetic DNA vaccine targeting the MERS coronavirus Spike (S) protein, the major surface antigen of coronaviruses, which is currently in clinical study. Here we build on this prior experience to generate a synthetic DNA-based vaccine candidate targeting SARS-CoV-2 S protein. The engineered construct, INO-4800, results in robust expression of the S protein in vitro. Following immunization of mice and guinea pigs with INO-4800 we measure antigen-specific T cell responses, functional antibodies which neutralize the SARS-CoV-2 infection and block Spike protein binding to the ACE2 receptor, and biodistribution of SARS-CoV-2 targeting antibodies to the lungs. This preliminary dataset identifies INO-4800 as a potential COVID-19 vaccine candidate, supporting further translational study.


Assuntos
Antígenos Virais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Antígenos Virais/química , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Mapeamento de Epitopos , Cobaias , Imunidade Humoral , Imunoglobulina G/imunologia , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Coronavírus da Síndrome Respiratória do Oriente Médio , Modelos Animais , Peptidil Dipeptidase A/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Vacinas Virais/química
14.
Arch Virol ; 165(7): 1611-1620, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32405826

RESUMO

Infectious bursal disease virus (IBDV), which infects young chickens, is one of the most important pathogens that harm the poultry industry. Evaluation of the immune status of birds before and after vaccination is of great importance for controlling the disease caused by this virus. Therefore, the development of low-cost and easy-to-manufacture test systems for IBDV antibody detection remains an urgent issue. In this study, three expression systems (bacteria, yeast, and human cells) were used to produce recombinant VP3 protein of IBDV. VP3 is a group-specific antigen and hence may be a good candidate for use in diagnostic tests. Comparison of the antigenic properties of the obtained polypeptides showed that the titres of antibodies raised in chickens against bacteria- or human-cell-derived recombinant VP3 were high, whereas the antibody level against yeast-derived recombinant VP3 was low. The results of an enzyme-linked immunosorbent assay (ELISA) of sera from IBDV-infected chickens demonstrated that the recombinant VP3 produced in E. coli would be the best choice for use in test systems.


Assuntos
Infecções por Birnaviridae/veterinária , Vírus da Doença Infecciosa da Bursa/imunologia , Peptídeos/imunologia , Doenças das Aves Domésticas/virologia , Proteínas Estruturais Virais/imunologia , Animais , Anticorpos Antivirais/imunologia , Infecções por Birnaviridae/virologia , Galinhas , Ensaio de Imunoadsorção Enzimática , Mapeamento de Epitopos , Escherichia coli/genética , Escherichia coli/metabolismo , Vírus da Doença Infecciosa da Bursa/química , Vírus da Doença Infecciosa da Bursa/genética , Vírus da Doença Infecciosa da Bursa/isolamento & purificação , Peptídeos/química , Peptídeos/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Estruturais Virais/química , Proteínas Estruturais Virais/genética
15.
Eur J Pharm Sci ; 151: 105387, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32454128

RESUMO

The emergence and rapid expansion of the coronavirus disease (COVID-19) require the development of effective countermeasures especially a vaccine to provide active acquired immunity against the virus. This study presented a comprehensive vaccinomics approach applied to the complete protein data published so far in the National Center for Biotechnological Information (NCBI) coronavirus data hub. We identified non-structural protein 8 (Nsp8), 3C-like proteinase, and spike glycoprotein as potential targets for immune responses to COVID-19. Epitopes prediction illustrated both B-cell and T-cell epitopes associated with the mentioned proteins. The shared B and T-cell epitopes: DRDAAMQRK and QARSEDKRA of Nsp8, EDMLNPNYEDL and EFTPFDVVR of 3C-like proteinase, and VNNSYECDIPI of the spike glycoprotein are regions of high potential interest and have a high likelihood of being recognized by the human immune system. The vaccine construct of the epitopes shows stimulation of robust primary immune responses and high level of interferon gamma. Also, the construct has the best conformation with respect to the tested innate immune receptors involving vigorous molecular mechanics and solvation energy. Designing of vaccination strategies that target immune response focusing on these conserved epitopes could generate immunity that not only provide cross protection across Betacoronaviruses but additionally resistant to virus evolution.


Assuntos
Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Desenho de Fármacos , Epitopos/imunologia , Pandemias/prevenção & controle , Pneumonia Viral/imunologia , Pneumonia Viral/prevenção & controle , Vacinas Virais/imunologia , Zoonoses/imunologia , Sequência de Aminoácidos , Animais , Linfócitos B/imunologia , Mapeamento de Epitopos , Glicoproteínas/imunologia , Humanos , Modelos Moleculares , Simulação de Dinâmica Molecular , Receptores Imunológicos/química , Receptores Imunológicos/imunologia , Linfócitos T/imunologia , Proteínas não Estruturais Virais/imunologia , Proteínas Virais/química , Proteínas Virais/imunologia
16.
Microbes Infect ; 22(4-5): 182-187, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32446902

RESUMO

Envelope protein of coronaviruses is a structural protein existing in both monomeric and homo-pentameric form. It has been related to a multitude of roles including virus infection, replication, dissemination and immune response stimulation. In the present study, we employed an immunoinformatic approach to investigate the major immunogenic domains of the SARS-CoV-2 envelope protein and map them among the homologue proteins of coronaviruses with tropism for animal species that are closely inter-related with the human beings population all over the world. Also, when not available, we predicted the envelope protein structural folding and mapped SARS-CoV-2 epitopes. Envelope sequences alignment provides evidence of high sequence homology for some of the investigated virus specimens; while the structural mapping of epitopes resulted in the interesting maintenance of the structural folding and epitope sequence localization also in the envelope proteins scoring a lower alignment score. In line with the One-Health approach, our evidences provide a molecular structural rationale for a potential role of taxonomically related coronaviruses in conferring protection from SARS-CoV-2 infection and identifying potential candidates for the development of diagnostic tools and prophylactic-oriented strategies.


Assuntos
Betacoronavirus/metabolismo , Biologia Computacional/métodos , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Proteínas do Envelope Viral/imunologia , Animais , Betacoronavirus/classificação , Betacoronavirus/genética , Betacoronavirus/imunologia , Mapeamento de Epitopos , Regulação Viral da Expressão Gênica , Humanos , Modelos Moleculares , Saúde Única , Pandemias , Filogenia , Conformação Proteica , Alinhamento de Sequência , Análise de Sequência de Proteína
17.
Cancer Sci ; 111(7): 2508-2525, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32415868

RESUMO

Human epidermal growth factor receptor 4 (HER4) isoforms have oncogenic or tumor suppressor functions depending on their susceptibility to proteolytic cleavage and HER4 intracellular domain (4ICD) translocation. Here, we report that the neuregulin 1 (NRG1) tumor suppressor mechanism through the HER4 JMa/CYT1 isoform can be mimicked by the agonist anti-HER4 Ab C6. Neuregulin 1 induced cleavage of poly(ADP-ribose) polymerase (PARP) and sub-G1 DNA fragmentation, and also reduced the metabolic activity of HER3- /HER4+ cervical (C-33A) and ovarian (COV318) cancer cells. This effect was confirmed in HER4 JMa/CYT1-, but not JMa/CYT2-transfected BT549 triple-negative breast cancer cells. Neuregulin 1 favored 4ICD cleavage and retention in mitochondria in JMa/CYT1-transfected BT549 cells, leading to reactive oxygen species (ROS) production through mitochondrial depolarization. Similarly, the anti-HER4 Ab C6, which binds to a conformational epitope located on a.a. 575-592 and 605-620 of HER4 domain IV, induced 4ICD cleavage and retention in mitochondria, and mimicked NRG1-mediated effects on PARP cleavage, ROS production, and mitochondrial membrane depolarization in cancer cells. In vivo, C6 reduced growth of COV434 and HCC1187 tumor cell xenografts in nude mice. Biasing 4ICD trafficking to mitochondria with anti-HER4 Abs to mimic NRG1 suppressor functions could be an alternative anticancer strategy.


Assuntos
Anticorpos Monoclonais/farmacologia , Receptor ErbB-4/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Anticorpos Monoclonais/imunologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Mapeamento de Epitopos , Humanos , Espaço Intracelular/metabolismo , Camundongos , Mitocôndrias/metabolismo , Neuregulina-1/farmacologia , Transporte Proteico/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Receptor ErbB-4/imunologia
18.
Virus Res ; 285: 198021, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32416259

RESUMO

A new betacoronavirus named SARS-CoV-2 has emerged as a new threat to global health and economy. A promising target for both diagnosis and therapeutics treatments of the new disease named COVID-19 is the coronavirus (CoV) spike (S) glycoprotein. By constant-pH Monte Carlo simulations and the PROCEEDpKa method, we have mapped the electrostatic epitopes for four monoclonal antibodies and the angiotensin-converting enzyme 2 (ACE2) on both SARS-CoV-1 and the new SARS-CoV-2 S receptor binding domain (RBD) proteins. We also calculated free energy of interactions and shown that the S RBD proteins from both SARS viruses binds to ACE2 with similar affinities. However, the affinity between the S RBD protein from the new SARS-CoV-2 and ACE2 is higher than for any studied antibody previously found complexed with SARS-CoV-1. Based on physical chemical analysis and free energies estimates, we can shed some light on the involved molecular recognition processes, their clinical aspects, the implications for drug developments, and suggest structural modifications on the CR3022 antibody that would improve its binding affinities for SARS-CoV-2 and contribute to address the ongoing international health crisis.


Assuntos
Betacoronavirus/química , Peptidil Dipeptidase A/metabolismo , Receptores Virais/metabolismo , Vírus da SARS/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/metabolismo , Betacoronavirus/imunologia , Simulação por Computador , Mapeamento de Epitopos , Humanos , Modelos Moleculares , Método de Monte Carlo , Peptidil Dipeptidase A/química , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas , Receptores Virais/química , Vírus da SARS/imunologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Termodinâmica
19.
Nat Commun ; 11(1): 2601, 2020 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-32433465

RESUMO

The coronavirus family member, SARS-CoV-2 has been identified as the causal agent for the pandemic viral pneumonia disease, COVID-19. At this time, no vaccine is available to control further dissemination of the disease. We have previously engineered a synthetic DNA vaccine targeting the MERS coronavirus Spike (S) protein, the major surface antigen of coronaviruses, which is currently in clinical study. Here we build on this prior experience to generate a synthetic DNA-based vaccine candidate targeting SARS-CoV-2 S protein. The engineered construct, INO-4800, results in robust expression of the S protein in vitro. Following immunization of mice and guinea pigs with INO-4800 we measure antigen-specific T cell responses, functional antibodies which neutralize the SARS-CoV-2 infection and block Spike protein binding to the ACE2 receptor, and biodistribution of SARS-CoV-2 targeting antibodies to the lungs. This preliminary dataset identifies INO-4800 as a potential COVID-19 vaccine candidate, supporting further translational study.


Assuntos
Antígenos Virais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Antígenos Virais/química , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Mapeamento de Epitopos , Cobaias , Imunidade Humoral , Imunoglobulina G/imunologia , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Coronavírus da Síndrome Respiratória do Oriente Médio , Modelos Animais , Peptidil Dipeptidase A/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Vacinas Virais/química
20.
Virology ; 546: 1-12, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32275203

RESUMO

Broadly neutralizing antibodies (bNAbs) may constitute an essential component of a protective vaccine against HIV-1, yet no immunogen has been able to elicit them. To characterize the development of bNAbs in HIV-1 subtype C infected individuals, a panel of 18 Env-pseudotyped viruses was used to screen 18 study participants. The specificity of plasma neutralization was mapped against Env mutants and MPER chimeras. Envelope (env) gene sequence evolution was characterized by single genome amplification and sequencing. Three out of eighteen individuals developed broad plasma neutralizing activity (>60% breadth). Two of the three participants may target epitopes comprising glycans at position 276 of the D loop in the CD4 binding site and 332 glycan supersite, respectively. Deletion of these glycans was associated with neutralization resistance. Our study describes the kinetics of the development of plasma neutralizing activity and identified amino acid residue changes suggestive of immune pressure on putative epitopes. The study enhances our understanding of how neutralization breadth develops in the course of HIV-1 subtype C infection.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Sequência de Aminoácidos , Mapeamento de Epitopos , Infecções por HIV/sangue , Infecções por HIV/virologia , HIV-1/química , HIV-1/classificação , HIV-1/genética , Humanos , Produtos do Gene env do Vírus da Imunodeficiência Humana/química , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
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