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1.
Int J Mol Sci ; 22(11)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34206009

RESUMO

Toll-like receptor (TLR) signaling plays a critical role in the induction and progression of autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematous, experimental autoimmune encephalitis, type 1 diabetes mellitus and neurodegenerative diseases. Deciphering antigen recognition by antibodies provides insights and defines the mechanism of action into the progression of immune responses. Multiple strategies, including phage display and hybridoma technologies, have been used to enhance the affinity of antibodies for their respective epitopes. Here, we investigate the TLR4 antibody-binding epitope by computational-driven approach. We demonstrate that three important residues, i.e., Y328, N329, and K349 of TLR4 antibody binding epitope identified upon in silico mutagenesis, affect not only the interaction and binding affinity of antibody but also influence the structural integrity of TLR4. Furthermore, we predict a novel epitope at the TLR4-MD2 interface which can be targeted and explored for therapeutic antibodies and small molecules. This technique provides an in-depth insight into antibody-antigen interactions at the resolution and will be beneficial for the development of new monoclonal antibodies. Computational techniques, if coupled with experimental methods, will shorten the duration of rational design and development of antibody therapeutics.


Assuntos
Anticorpos Monoclonais/imunologia , Artrite Reumatoide/imunologia , Encefalite/imunologia , Epitopos/genética , Doença de Hashimoto/imunologia , Doenças Neurodegenerativas/imunologia , Receptor 4 Toll-Like/genética , Sequência de Aminoácidos/genética , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/genética , Técnicas de Visualização da Superfície Celular , Encefalite/genética , Encefalite/patologia , Mapeamento de Epitopos/métodos , Epitopos/imunologia , Doença de Hashimoto/genética , Doença de Hashimoto/patologia , Humanos , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/patologia , Ligação Proteica/genética , Receptor 4 Toll-Like/antagonistas & inibidores , Receptor 4 Toll-Like/imunologia
2.
Front Immunol ; 12: 678570, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34211469

RESUMO

Passive immunization using monoclonal antibodies will play a vital role in the fight against COVID-19. The recent emergence of viral variants with reduced sensitivity to some current antibodies and vaccines highlights the importance of broad cross-reactivity. This study describes deep-mining of the antibody repertoires of hospitalized COVID-19 patients using phage display technology and B cell receptor (BCR) repertoire sequencing to isolate neutralizing antibodies and gain insights into the early antibody response. This comprehensive discovery approach has yielded a panel of potent neutralizing antibodies which bind distinct viral epitopes including epitopes conserved in SARS-CoV-1. Structural determination of a non-ACE2 receptor blocking antibody reveals a previously undescribed binding epitope, which is unlikely to be affected by the mutations in any of the recently reported major viral variants including B.1.1.7 (from the UK), B.1.351 (from South Africa) and B.1.1.28 (from Brazil). Finally, by combining sequences of the RBD binding and neutralizing antibodies with the B cell receptor repertoire sequencing, we also describe a highly convergent early antibody response. Similar IgM-derived sequences occur within this study group and also within patient responses described by multiple independent studies published previously.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Anticorpos Neutralizantes/uso terapêutico , COVID-19/prevenção & controle , COVID-19/terapia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/imunologia , Técnicas de Visualização da Superfície Celular/métodos , Mineração de Dados/métodos , Epitopos/imunologia , Humanos , Imunização Passiva/métodos
3.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 37(8): 710-715, 2021 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-34236030

RESUMO

Objective To study the bioinformatics characteristics of HSP70 domain proteins derived from pollen of Populus deltoides (P. deltoides), optimize the prokaryotic expression methods, and identify the biological activity of these proteins. Methods Physicochemical characteristics of three kinds of HSP70 domain-containing proteins were analyzed by bioinformatics software. The T/B cell epitopes of these proteins were predicted by Immune Epitope Database and Analysis Resource (IEDB). According to the amino acid sequence provided by Uniprot database, their nucleotide sequences were synthesized and cloned into pET28a(+) plasmid for prokaryotic expression. Protein expression was detected by SDS-PAGE, then the expressed products were purified by nickel column and identified by Western blotting. The protein concentration was measured by protein quantitative kit. Then the three proteins were used as antigens to prepare mouse asthma models, and the concentration of serum total IgE antibody was determined by ELISA. Results The bioinformatics analysis showed that the relative molecular mass (Mr) of B9N9W6, B9GX02 and A0A2K2AYN8 were 71 900, 94 600 and 75 200, respectively. The 13 T-cell epitopes and 14 B-cell epitopes were identified in the three proteins which had high hydrophilia and stability. SDS-PAGE analysis revealed that the genes encoding the three proteins were expressed with three specific bands of approximately Mr 72 000, 95 000 and 75 000, respectively. Western blotting showed the specific bands at the corresponding sites. ELISA showed that the IgE level in the extract group and the A0A2K2AYN8 group were higher than that in the PBS group. Compared with the A0A2K2AYN8 group, the IgE concentration in the B9N9W6 group and B9GX02 group increased significantly. Conclusion The soluble HSP70 domain-containing proteins A0A2K2AYN8, B9GX02 and B9N9W6 derived from pollen of P. deltoides can be expressed as well as purified, and have the biological activity of producing IgE antibodies.


Assuntos
Populus , Alérgenos , Animais , Western Blotting , Biologia Computacional , Epitopos , Camundongos , Pólen/genética , Populus/genética
4.
JCI Insight ; 6(13)2021 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-34081630

RESUMO

BACKGROUNDThe role of humoral immunity in COVID-19 is not fully understood, owing, in large part, to the complexity of antibodies produced in response to the SARS-CoV-2 infection. There is a pressing need for serology tests to assess patient-specific antibody response and predict clinical outcome.METHODSUsing SARS-CoV-2 proteome and peptide microarrays, we screened 146 COVID-19 patients' plasma samples to identify antigens and epitopes. This enabled us to develop a master epitope array and an epitope-specific agglutination assay to gauge antibody responses systematically and with high resolution.RESULTSWe identified linear epitopes from the spike (S) and nucleocapsid (N) proteins and showed that the epitopes enabled higher resolution antibody profiling than the S or N protein antigen. Specifically, we found that antibody responses to the S-811-825, S-881-895, and N-156-170 epitopes negatively or positively correlated with clinical severity or patient survival. Moreover, we found that the P681H and S235F mutations associated with the coronavirus variant of concern B.1.1.7 altered the specificity of the corresponding epitopes.CONCLUSIONEpitope-resolved antibody testing not only affords a high-resolution alternative to conventional immunoassays to delineate the complex humoral immunity to SARS-CoV-2 and differentiate between neutralizing and non-neutralizing antibodies, but it also may potentially be used to predict clinical outcome. The epitope peptides can be readily modified to detect antibodies against variants of concern in both the peptide array and latex agglutination formats.FUNDINGOntario Research Fund (ORF) COVID-19 Rapid Research Fund, Toronto COVID-19 Action Fund, Western University, Lawson Health Research Institute, London Health Sciences Foundation, and Academic Medical Organization of Southwestern Ontario (AMOSO) Innovation Fund.


Assuntos
Testes de Aglutinação/métodos , Formação de Anticorpos/imunologia , Teste Sorológico para COVID-19/métodos , COVID-19/imunologia , Epitopos de Linfócito B/imunologia , SARS-CoV-2/imunologia , Sequência de Aminoácidos , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Especificidade de Anticorpos/imunologia , COVID-19/sangue , COVID-19/mortalidade , Epitopos/imunologia , Epitopos de Linfócito B/química , Epitopos de Linfócito B/genética , Humanos , Imunidade Humoral , Análise em Microsséries/métodos , Nucleocapsídeo/química , Nucleocapsídeo/genética , Nucleocapsídeo/imunologia , Peptídeos/imunologia , SARS-CoV-2/genética , Índice de Gravidade de Doença , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia
5.
MAbs ; 13(1): 1930636, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34097570

RESUMO

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes coronavirus disease-2019 (COVID-19), interacts with the host cell receptor angiotensin-converting enzyme 2 (hACE2) via its spike 1 protein during infection. After the virus sequence was published, we identified two potent antibodies against the SARS-CoV-2 receptor binding domain (RBD) from antibody libraries using a phage-to-yeast (PtY) display platform in only 10 days. Our lead antibody JMB2002, now in a Phase 1 clinical trial (ChiCTR2100042150), showed broad-spectrum in vitro blocking activity against hACE2 binding to the RBD of multiple SARS-CoV-2 variants, including B.1.351 that was reportedly much more resistant to neutralization by convalescent plasma, vaccine sera and some clinical-stage neutralizing antibodies. Furthermore, JMB2002 has demonstrated complete prophylactic and potent therapeutic efficacy in a rhesus macaque disease model. Prophylactic and therapeutic countermeasure intervention of SARS-CoV-2 using JMB2002 would likely slow down the transmission of currently emerged SARS-CoV-2 variants and result in more efficient control of the COVID-19 pandemic.


Assuntos
Enzima de Conversão de Angiotensina 2/antagonistas & inibidores , Anticorpos Neutralizantes/farmacologia , Antivirais/farmacologia , COVID-19/tratamento farmacológico , COVID-19/prevenção & controle , SARS-CoV-2/efeitos dos fármacos , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/imunologia , Animais , Especificidade de Anticorpos , Sítios de Ligação de Anticorpos , Células CHO , COVID-19/imunologia , COVID-19/metabolismo , COVID-19/virologia , Chlorocebus aethiops , Cricetulus , Modelos Animais de Doenças , Epitopos , Macaca mulatta , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Células Vero
6.
Nat Commun ; 12(1): 3789, 2021 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-34145279

RESUMO

Influenza viruses are a major public health problem. Vaccines are the best available countermeasure to induce effective immunity against infection with seasonal influenza viruses; however, the breadth of antibody responses in infection versus vaccination is quite different. Here, we show that nasal infection controls two sequential processes to induce neutralizing IgG antibodies recognizing the hemagglutinin (HA) of heterotypic strains. The first is viral replication in the lung, which facilitates exposure of shared epitopes that are otherwise hidden from the immune system. The second process is the germinal center (GC) response, in particular, IL-4 derived from follicular helper T cells has an essential role in the expansion of rare GC-B cells recognizing the shared epitopes. Therefore, the combination of exposure of the shared epitopes and efficient proliferation of GC-B cells is critical for generating broadly-protective antibodies. These observations provide insight into mechanisms promoting broad protection from virus infection.


Assuntos
Anticorpos Antivirais/imunologia , Linfócitos B/imunologia , Anticorpos Amplamente Neutralizantes/imunologia , Hemaglutininas Virais/imunologia , Interleucina-4/imunologia , Infecções por Orthomyxoviridae/imunologia , Animais , Anticorpos Antivirais/sangue , Anticorpos Amplamente Neutralizantes/sangue , Epitopos/imunologia , Feminino , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A Subtipo H2N2/imunologia , Vírus da Influenza A Subtipo H3N2/imunologia , Vacinas contra Influenza/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/imunologia , Células T Auxiliares Foliculares/imunologia , Vacinação
8.
Nat Commun ; 12(1): 3573, 2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-34117223

RESUMO

O-GalNAc glycans (or mucin O-glycans) play pivotal roles in diverse biological and pathological processes, including tumor growth and progression. Structurally defined O-GalNAc glycans are essential for functional studies but synthetic challenges and their inherent structural diversity and complexity have limited access to these compounds. Herein, we report an efficient and robust chemoenzymatic modular assembly (CEMA) strategy to construct structurally diverse O-GalNAc glycans. The key to this strategy is the convergent assembly of O-GalNAc cores 1-4 and 6 from three chemical building blocks, followed by enzymatic diversification of the cores by 13 well-tailored enzyme modules. A total of 83 O-GalNAc glycans presenting various natural glycan epitopes are obtained and used to generate a unique synthetic mucin O-glycan microarray. Binding specificities of glycan-binding proteins (GBPs) including plant lectins and selected anti-glycan antibodies towards these O-GalNAc glycans are revealed by this microarray, promoting their applicability in functional O-glycomics. Serum samples from colorectal cancer patients and healthy controls are assayed using the array reveal higher bindings towards less common cores 3, 4, and 6 than abundant cores 1 and 2, providing insights into O-GalNAc glycan structure-activity relationships.


Assuntos
Glicômica , Mucinas/química , Mucinas/metabolismo , Polissacarídeos/química , Polissacarídeos/metabolismo , Carboidratos , Proteínas de Transporte/metabolismo , Epitopos , Glicosilação , Humanos , Análise em Microsséries
9.
Front Public Health ; 9: 690017, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34123998

RESUMO

Despite substantial progress in confronting the global HIV-1 epidemic since its inception in the 1980s, better approaches for both treatment and prevention will be necessary to end the epidemic and remain a top public health priority. Antiretroviral therapy (ART) has been effective in extending lives, but at a cost of lifelong adherence to treatment. Broadly neutralizing antibodies (bNAbs) are directed to conserved regions of the HIV-1 envelope glycoprotein trimer (Env) and can block infection if present at the time of viral exposure. The therapeutic application of bNAbs holds great promise, and progress is being made toward their development for widespread clinical use. Compared to the current standard of care of small molecule-based ART, bNAbs offer: (1) reduced toxicity; (2) the advantages of extended half-lives that would bypass daily dosing requirements; and (3) the potential to incorporate a wider immune response through Fc signaling. Recent advances in discovery technology can enable system-wide mining of the immunoglobulin repertoire and will continue to accelerate isolation of next generation potent bNAbs. Passive transfer studies in pre-clinical models and clinical trials have demonstrated the utility of bNAbs in blocking or limiting transmission and achieving viral suppression. These studies have helped to define the window of opportunity for optimal intervention to achieve viral clearance, either using bNAbs alone or in combination with ART. None of these advances with bNAbs would be possible without technological advancements and expanding the cohorts of donor participation. Together these elements fueled the remarkable growth in bNAb development. Here, we review the development of bNAbs as therapies for HIV-1, exploring advances in discovery, insights from animal models and early clinical trials, and innovations to optimize their clinical potential through efforts to extend half-life, maximize the contribution of Fc effector functions, preclude escape through multiepitope targeting, and the potential for sustained delivery.


Assuntos
Infecções por HIV , HIV-1 , Animais , Anticorpos Neutralizantes/uso terapêutico , Anticorpos Amplamente Neutralizantes , Epitopos , Anticorpos Anti-HIV , Infecções por HIV/tratamento farmacológico , Produtos do Gene env do Vírus da Imunodeficiência Humana
10.
Int J Mol Sci ; 22(11)2021 May 22.
Artigo em Inglês | MEDLINE | ID: covidwho-1244041

RESUMO

The COVID-19 pandemic is caused by SARS-CoV-2. Currently, most of the research efforts towards the development of vaccines and antibodies against SARS-CoV-2 were mainly focused on the spike (S) protein, which mediates virus entry into the host cell by binding to ACE2. As the virus SARS-CoV-2 continues to spread globally, variants have emerged, characterized by multiple mutations of the S glycoprotein. Herein, we employed microsecond-long molecular dynamics simulations to study the impact of the mutations of the S glycoprotein in SARS-CoV-2 Variant of Concern 202012/01 (B.1.1.7), termed the "UK variant", in comparison with the wild type, with the aim to decipher the structural basis of the reported increased infectivity and virulence. The simulations provided insights on the different dynamics of UK and wild-type S glycoprotein, regarding in particular the Receptor Binding Domain (RBD). In addition, we investigated the role of glycans in modulating the conformational transitions of the RBD. The overall results showed that the UK mutant experiences higher flexibility in the RBD with respect to wild type; this behavior might be correlated with the increased transmission reported for this variant. Our work also adds useful structural information on antigenic "hotspots" and epitopes targeted by neutralizing antibodies.


Assuntos
COVID-19/virologia , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Anticorpos Neutralizantes/imunologia , Sítios de Ligação , Epitopos , Humanos , Ligação de Hidrogênio , Simulação de Dinâmica Molecular , Polissacarídeos/química , Polissacarídeos/metabolismo , Domínios Proteicos , Domínios e Motivos de Interação entre Proteínas , SARS-CoV-2/patogenicidade , Glicoproteína da Espícula de Coronavírus/metabolismo , Reino Unido
11.
Nat Rev Microbiol ; 19(7): 409-424, 2021 07.
Artigo em Inglês | MEDLINE | ID: covidwho-1253944

RESUMO

Although most mutations in the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome are expected to be either deleterious and swiftly purged or relatively neutral, a small proportion will affect functional properties and may alter infectivity, disease severity or interactions with host immunity. The emergence of SARS-CoV-2 in late 2019 was followed by a period of relative evolutionary stasis lasting about 11 months. Since late 2020, however, SARS-CoV-2 evolution has been characterized by the emergence of sets of mutations, in the context of 'variants of concern', that impact virus characteristics, including transmissibility and antigenicity, probably in response to the changing immune profile of the human population. There is emerging evidence of reduced neutralization of some SARS-CoV-2 variants by postvaccination serum; however, a greater understanding of correlates of protection is required to evaluate how this may impact vaccine effectiveness. Nonetheless, manufacturers are preparing platforms for a possible update of vaccine sequences, and it is crucial that surveillance of genetic and antigenic changes in the global virus population is done alongside experiments to elucidate the phenotypic impacts of mutations. In this Review, we summarize the literature on mutations of the SARS-CoV-2 spike protein, the primary antigen, focusing on their impacts on antigenicity and contextualizing them in the protein structure, and discuss them in the context of observed mutation frequencies in global sequence datasets.


Assuntos
COVID-19/virologia , Evasão da Resposta Imune , SARS-CoV-2/genética , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/classificação , Aminoácidos/química , Aminoácidos/genética , Variação Antigênica/genética , Variação Antigênica/fisiologia , COVID-19/imunologia , COVID-19/prevenção & controle , COVID-19/transmissão , Vacinas contra COVID-19/imunologia , Vacinas contra COVID-19/normas , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Humanos , Evasão da Resposta Imune/genética , Mutação , Conformação Proteica , SARS-CoV-2/classificação , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia
13.
Front Immunol ; 12: 690742, 2021.
Artigo em Inglês | MEDLINE | ID: covidwho-1268253

RESUMO

Since December 2019, the SARS-CoV-2 has erupted on a large scale worldwide and spread rapidly. Passive immunization of antibody-related molecules provides opportunities for prevention and treatment of high-risk patients and children. Nanobodies (Nbs) have many strong physical and chemical properties. They can be atomized, administered by inhalation, and can be directly applied to the infected site, with fast onset, high local drug concentration/high bioavailability, and high patient compliance (no needles). It has very attractive potential in the treatment of respiratory viruses. Rapid and low-cost development of Nbs targeting SARS-CoV-2 can quickly be achieved. Nbs against SARS-CoV-2 mutant strains also can be utilized quickly to prevent the virus from escaping. It provides important technical supports for the treatment of the SARS-CoV-2 and has the potential to become an essential medicine in the toolbox against the SARS-CoV-2.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/imunologia , COVID-19/virologia , SARS-CoV-2/imunologia , Anticorpos de Domínio Único/imunologia , Animais , Afinidade de Anticorpos/imunologia , Sítios de Ligação , Epitopos/imunologia , Humanos , Testes de Neutralização , Biblioteca de Peptídeos , Ligação Proteica
14.
Hum Immunol ; 82(8): 551-560, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: covidwho-1265675

RESUMO

Given the highly polymorphic nature of Human Leukocyte Antigen (HLA) molecules, it is not surprising that they function as key regulators of the host immune response to almost all invading pathogens, including SARS-CoV-2, the etiological agent responsible for the recent COVID-19 pandemic. Several correlations have already been established between the expression of a specific HLA allele/haplotype and susceptibility/progression of SARS-CoV-2 infection and new ones are continuously emerging. Protective and harmful HLA variants have been described in both mild and severe forms of the disease, but considering the huge amount of existing variants, the data gathered in such a brief span of time are to some extent confusing and contradictory. The aim of this mini-review is to provide a snap-shot of the main findings so far collected on the HLA-SARS-CoV-2 interaction, so as to partially untangle this intricate yarn. As key factors in the generation of antigenic peptides to be presented by HLA molecules, ERAP1 and ERAP2 role in SARS-CoV-2 infection will be revised as well.


Assuntos
Aminopeptidases/genética , Apresentação do Antígeno , Antígenos Virais/imunologia , COVID-19/genética , Antígenos HLA/genética , Antígenos de Histocompatibilidade Menor/genética , Polimorfismo Genético , SARS-CoV-2/imunologia , Aminopeptidases/imunologia , Animais , COVID-19/diagnóstico , COVID-19/imunologia , COVID-19/virologia , Epitopos , Antígenos HLA/imunologia , Interações Hospedeiro-Patógeno , Humanos , Antígenos de Histocompatibilidade Menor/imunologia , SARS-CoV-2/patogenicidade
15.
MAbs ; 13(1): 1930636, 2021.
Artigo em Inglês | MEDLINE | ID: covidwho-1258715

RESUMO

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes coronavirus disease-2019 (COVID-19), interacts with the host cell receptor angiotensin-converting enzyme 2 (hACE2) via its spike 1 protein during infection. After the virus sequence was published, we identified two potent antibodies against the SARS-CoV-2 receptor binding domain (RBD) from antibody libraries using a phage-to-yeast (PtY) display platform in only 10 days. Our lead antibody JMB2002, now in a Phase 1 clinical trial (ChiCTR2100042150), showed broad-spectrum in vitro blocking activity against hACE2 binding to the RBD of multiple SARS-CoV-2 variants, including B.1.351 that was reportedly much more resistant to neutralization by convalescent plasma, vaccine sera and some clinical-stage neutralizing antibodies. Furthermore, JMB2002 has demonstrated complete prophylactic and potent therapeutic efficacy in a rhesus macaque disease model. Prophylactic and therapeutic countermeasure intervention of SARS-CoV-2 using JMB2002 would likely slow down the transmission of currently emerged SARS-CoV-2 variants and result in more efficient control of the COVID-19 pandemic.


Assuntos
Enzima de Conversão de Angiotensina 2/antagonistas & inibidores , Anticorpos Neutralizantes/farmacologia , Antivirais/farmacologia , COVID-19/tratamento farmacológico , COVID-19/prevenção & controle , SARS-CoV-2/efeitos dos fármacos , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/imunologia , Animais , Especificidade de Anticorpos , Sítios de Ligação de Anticorpos , Células CHO , COVID-19/imunologia , COVID-19/metabolismo , COVID-19/virologia , Chlorocebus aethiops , Cricetulus , Modelos Animais de Doenças , Epitopos , Macaca mulatta , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Células Vero
16.
Emerg Microbes Infect ; 10(1): 1390-1403, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34120577

RESUMO

Global concerns arose as the emerged and rapidly spreading SARS-CoV-2 variants might escape host immunity induced by vaccination. In this study, a heterologous prime-boost immunization strategy for COVID-19 was designed to prime with a DNA vaccine encoding wild type (WT) spike protein receptor-binding domain (RBD) followed by S1 protein-based vaccine in rabbits. Four vaccine-elicited rabbit monoclonal antibodies (RmAbs), including 1H1, 9H1, 7G5, and 5E1, were isolated for biophysical property, neutralization potency and sequence analysis. All RmAbs recognized RBD or S1 protein with KD in the low nM or sub nM range. 1H1 and 9H1, but neither 7G5 nor 5E1, can bind to all RBD protein variants derived from B.1.351. All four RmAbs were able to neutralize wild type (WT) SARS-CoV-2 strain in pseudovirus assay, and 1H1 and 9H1 could neutralize the SARS-CoV-2 WT authentic virus with IC50 values of 0.136 and 0.026 µg/mL, respectively. Notably, 1H1 was able to neutralize all 6 emerging SARS-CoV-2 variants tested including D614G, B.1.1.7, B.1.429, P.1, B.1.526, and B.1.351 variants, and 5E1 could neutralize against the above 5 variants except P.1. Epitope binning analysis revealed that 9H1, 5E1 and 1H1 recognized distinct epitopes, while 9H1 and 7G5 may have overlapping but not identical epitope. In conclusion, DNA priming protein boost vaccination was an effective strategy to induce RmAbs with potent neutralization capability against not only SARS-CoV-2 WT strain but also emergent variants, which may provide a new avenue for effective therapeutics and point-of-care diagnostic measures.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Variação Genética , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/genética , Vacinas de DNA/imunologia , Animais , Anticorpos Antivirais/sangue , Epitopos , Humanos , Imunização Secundária , Domínios Proteicos/imunologia , Domínios Proteicos/fisiologia , Coelhos , SARS-CoV-2/imunologia , Vacinação , Vacinas Sintéticas , Ligação Viral
17.
Sci Rep ; 11(1): 12448, 2021 06 14.
Artigo em Inglês | MEDLINE | ID: covidwho-1268001

RESUMO

The SARS-CoV-2 spike glycoprotein is a focal point for vaccine immunogen and therapeutic antibody design, and also serves as a critical antigen in the evaluation of immune responses to COVID-19. A common feature amongst enveloped viruses such as SARS-CoV-2 and HIV-1 is the propensity for displaying host-derived glycans on entry spike proteins. Similarly displayed glycosylation motifs can serve as the basis for glyco-epitope mediated cross-reactivity by antibodies, which can have important implications on virus neutralization, antibody-dependent enhancement (ADE) of infection, and the interpretation of antibody titers in serological assays. From a panel of nine anti-HIV-1 gp120 reactive antibodies, we selected two (PGT126 and PGT128) that displayed high levels of cross-reactivity with the SARS-CoV-2 spike. We report that these antibodies are incapable of neutralizing pseudoviruses expressing SARS-CoV-2 spike proteins and are unlikely to mediate ADE via FcγRII receptor engagement. Nevertheless, ELISA and other immunoreactivity experiments demonstrate these antibodies are capable of binding the SARS-CoV-2 spike in a glycan-dependent manner. These results contribute to the growing literature surrounding SARS-CoV-2 S cross-reactivity, as we demonstrate the ability for cross-reactive antibodies to interfere in immunoassays.


Assuntos
Anticorpos Anti-HIV/imunologia , Polissacarídeos/imunologia , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/imunologia , Reações Antígeno-Anticorpo , COVID-19/patologia , COVID-19/virologia , Reações Cruzadas , Epitopos/imunologia , Anticorpos Anti-HIV/farmacologia , Humanos , SARS-CoV-2/isolamento & purificação , Internalização do Vírus/efeitos dos fármacos
18.
Sci Transl Med ; 13(596)2021 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-34078743

RESUMO

Broadly neutralizing antibodies are critical for protection against both drifted and shifted influenza viruses. Here, we reveal that first exposure to the 2009 pandemic H1N1 influenza virus recalls memory B cells that are specific to the conserved receptor-binding site (RBS) or lateral patch epitopes of the hemagglutinin (HA) head domain. Monoclonal antibodies (mAbs) generated against these epitopes are broadly neutralizing against H1N1 viruses spanning 40 years of viral evolution and provide potent protection in vivo. Lateral patch-targeting antibodies demonstrated near universal binding to H1 viruses, and RBS-binding antibodies commonly cross-reacted with H3N2 viruses and influenza B viruses. Lateral patch-targeting mAbs were restricted to expressing the variable heavy-chain gene VH3-23 with or without the variable kappa-chain gene VK1-33 and often had a Y-x-R motif within the heavy-chain complementarity determining region 3 to make key contacts with HA. Moreover, lateral patch antibodies that used both VH3-23 and VK1-33 maintained neutralizing capability with recent pH1N1 strains that acquired mutations near the lateral patch. RBS-binding mAbs used a diverse repertoire but targeted the RBS epitope similarly and made extensive contacts with the major antigenic site Sb. Together, our data indicate that RBS- and lateral patch-targeting clones are abundant within the human memory B cell pool, and universal vaccine strategies should aim to drive antibodies against both conserved head and stalk epitopes.


Assuntos
Vírus da Influenza A Subtipo H1N1 , Vacinas contra Influenza , Influenza Humana , Anticorpos Neutralizantes , Anticorpos Antivirais , Anticorpos Amplamente Neutralizantes , Epitopos , Glicoproteínas de Hemaglutininação de Vírus da Influenza , Hemaglutininas , Humanos , Vírus da Influenza A Subtipo H3N2
19.
Structure ; 29(7): 655-663.e4, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34111408

RESUMO

Emerging SARS-CoV-2 strains, B.1.1.7 and B.1.351, from the UK and South Africa, respectively, show decreased neutralization by monoclonal antibodies and convalescent or vaccinee sera raised against the original wild-type virus, and are thus of clinical concern. However, the neutralization potency of two antibodies, 1-57 and 2-7, which target the receptor-binding domain (RBD) of the spike, was unaffected by these emerging strains. Here, we report cryo-EM structures of 1-57 and 2-7 in complex with spike, revealing each of these antibodies to utilize a distinct mechanism to bypass or accommodate RBD mutations. Notably, each antibody represented an immune response with recognition distinct from those of frequent antibody classes. Moreover, many epitope residues recognized by 1-57 and 2-7 were outside hotspots of evolutionary pressure for ACE2 binding and neutralizing antibody escape. We suggest the therapeutic use of antibodies, such as 1-57 and 2-7, which target less prevalent epitopes, could ameliorate issues of monoclonal antibody escape.


Assuntos
Enzima de Conversão de Angiotensina 2/química , Anticorpos Monoclonais/química , Anticorpos Neutralizantes/química , Anticorpos Antivirais/química , Receptores Virais/química , SARS-CoV-2/química , Glicoproteína da Espícula de Coronavírus/química , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/imunologia , Enzima de Conversão de Angiotensina 2/metabolismo , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/metabolismo , Anticorpos Antivirais/genética , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/metabolismo , Sítios de Ligação , Clonagem Molecular , Microscopia Crioeletrônica , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Epitopos/metabolismo , Expressão Gênica , Células HEK293 , Humanos , Modelos Moleculares , Mutação , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Receptores Virais/genética , Receptores Virais/imunologia , Receptores Virais/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , SARS-CoV-2/genética , SARS-CoV-2/imunologia , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo
20.
J Med Case Rep ; 15(1): 313, 2021 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-34088358

RESUMO

BACKGROUND: Sugammadex is a synthetic γ-cyclodextrin derivative designed to selectively bind to steroidal neuromuscular blocking agents and reverse their effects. Although many cases of sugammadex-induced anaphylaxis have been reported, few studies have investigated the underlying mechanism. CASE PRESENTATION: A 55-year-old Japanese man underwent a laryngectomy under general anesthesia. One month before laryngectomy, he had undergone laryngoscopy under general anesthesia and received sugammadex administration without causing hypersensitivity. He had no history of allergies. The operation was finished without complications. Shortly after sugammadex administration, his blood pressure dropped to approximately 70 mmHg, and his heart rate increased to 110 beats/minute with systemic erythema. Suspecting anaphylaxis, he was treated with the intravenous injection of phenylephrine, D-chlorpheniramine, and hydrocortisone. After these treatments, his cardiovascular condition stabilized. Eight months after the event, skin prick tests and intradermal tests with all agents used during general anesthesia were performed. Intradermal tests showed positive results only for sugammadex. Subsequently, basophil activation tests with CD203c were performed using sugammadex, γ-cyclodextrin, and positive controls (anti-immunoglobulin-E and formyl-methionyl-leucyl-phenylalanine). In addition to both controls, sugammadex, but not γ-cyclodextrin, induced significant upregulation of CD203c expression. We performed additional basophil activation tests with wortmannin, an inhibitor of phosphoinositide 3-kinase, to investigate the mechanism underlying sugammadex-induced basophil activation. The inhibitory effect of wortmannin on basophil activation due to sugammadex was similar to that of anti-immunoglobulin-E, suggesting an immunoglobulin-E-dependent mechanism. Although the patient showed no hypersensitivity after the first exposure of sugammadex, anaphylaxis appeared after the second administration. Because most cases of sugammadex-induced anaphylaxis reportedly appeared after first administration, this seems to be a rare case. CONCLUSIONS: In the present case, sugammadex-induced anaphylaxis might have occurred through an immunoglobulin-E-dependent mechanism and not involve γ-cyclodextrin as an epitope. Physicians should pay attention to the occurrence of sugammadex-induced anaphylaxis even when the patient has a history of safe administration of sugammadex.


Assuntos
Anafilaxia , gama-Ciclodextrinas , Anafilaxia/induzido quimicamente , Epitopos , Humanos , Imunoglobulina E , Masculino , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases , Sugammadex , gama-Ciclodextrinas/efeitos adversos
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