RESUMO
The omicron variants of SARS-CoV-2 have substantial ability to escape infection- and vaccine-elicited antibody immunity. Here, we investigated the extent of such escape in nine convalescent patients infected with the wild-type SARS-CoV-2 during the first wave of the pandemic. Among the total of 476 monoclonal antibodies (mAbs) isolated from peripheral memory B cells, we identified seven mAbs with broad neutralizing activity to all variants tested, including various omicron subvariants. Biochemical and structural analysis indicated the majority of these mAbs bound to the receptor-binding domain, mimicked the receptor ACE2 and were able to accommodate or inadvertently improve recognition of omicron substitutions. Passive delivery of representative antibodies protected K18-hACE2 mice from infection with omicron and beta SARS-CoV-2. A deeper understanding of how the memory B cells that produce these antibodies could be selectively boosted or recalled can augment antibody immunity against SARS-CoV-2 variants.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , Camundongos , Anticorpos Monoclonais , Anticorpos Antivirais , Anticorpos NeutralizantesRESUMO
The 2003 severe acute respiratory syndrome coronavirus (SARS-CoV-1) causes more severe disease than SARS-CoV-2, which is responsible for COVID-19. However, our understanding of antibody response to SARS-CoV-1 infection remains incomplete. Herein, we studied the antibody responses in 25 SARS-CoV-1 convalescent patients. Plasma neutralization was higher and lasted longer in SARS-CoV-1 patients than in severe SARS-CoV-2 patients. Among 77 monoclonal antibodies (mAbs) isolated, 60 targeted the receptor-binding domain (RBD) and formed 7 groups (RBD-1 to RBD-7) based on their distinct binding and structural profiles. Notably, RBD-7 antibodies bound to a unique RBD region interfaced with the N-terminal domain of the neighboring protomer (NTD proximal) and were more prevalent in SARS-CoV-1 patients. Broadly neutralizing antibodies for SARS-CoV-1, SARS-CoV-2, and bat and pangolin coronaviruses were also identified. These results provide further insights into the antibody response to SARS-CoV-1 and inform the design of more effective strategies against diverse human and animal coronaviruses.
Assuntos
COVID-19 , Animais , Humanos , Anticorpos Antivirais , Formação de Anticorpos , SARS-CoV-2 , Anticorpos NeutralizantesRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge during the global pandemic and may facilitate escape from current antibody therapies and vaccine protection. Here we showed that the South African variant B.1.351 was the most resistant to current monoclonal antibodies and convalescent plasma from coronavirus disease 2019 (COVID-19)-infected individuals, followed by the Brazilian variant P.1 and the United Kingdom variant B.1.1.7. This resistance hierarchy corresponded with Y144del and 242-244del mutations in the N-terminal domain and K417N/T, E484K, and N501Y mutations in the receptor-binding domain (RBD) of SARS-CoV-2. Crystal structure analysis of the B.1.351 triple mutant (417N-484K-501Y) RBD complexed with the monoclonal antibody P2C-1F11 revealed the molecular basis for antibody neutralization and escape. B.1.351 and P.1 also acquired the ability to use mouse and mink ACE2 receptors for entry. Our results demonstrate major antigenic shifts and potential broadening of the host range for B.1.351 and P.1 variants, which poses serious challenges to current antibody therapies and vaccine protection.
Assuntos
Enzima de Conversão de Angiotensina 2/metabolismo , Anticorpos Neutralizantes/imunologia , Evasão da Resposta Imune , SARS-CoV-2/imunologia , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/química , Variação Antigênica/genética , COVID-19/imunologia , COVID-19/virologia , Especificidade de Hospedeiro , Humanos , Evasão da Resposta Imune/genética , Camundongos , Vison , Mutação , Ligação Proteica , SARS-CoV-2/genética , SARS-CoV-2/fisiologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Internalização do VírusRESUMO
The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) presents a global health emergency that is in urgent need of intervention1-3. The entry of SARS-CoV-2 into its target cells depends on binding between the receptor-binding domain (RBD) of the viral spike protein and its cellular receptor, angiotensin-converting enzyme 2 (ACE2)2,4-6. Here we report the isolation and characterization of 206 RBD-specific monoclonal antibodies derived from single B cells from 8 individuals infected with SARS-CoV-2. We identified antibodies that potently neutralize SARS-CoV-2; this activity correlates with competition with ACE2 for binding to RBD. Unexpectedly, the anti-SARS-CoV-2 antibodies and the infected plasma did not cross-react with the RBDs of SARS-CoV or Middle East respiratory syndrome-related coronavirus (MERS-CoV), although there was substantial plasma cross-reactivity to their trimeric spike proteins. Analysis of the crystal structure of RBD-bound antibody revealed that steric hindrance inhibits viral engagement with ACE2, thereby blocking viral entry. These findings suggest that anti-RBD antibodies are largely viral-species-specific inhibitors. The antibodies identified here may be candidates for development of clinical interventions against SARS-CoV-2.
Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto , Idoso , Enzima de Conversão de Angiotensina 2 , Anticorpos Neutralizantes/química , Anticorpos Antivirais/química , Linfócitos B/citologia , Linfócitos B/imunologia , Betacoronavirus/química , COVID-19 , Criança , Células Clonais/citologia , Células Clonais/imunologia , Reações Cruzadas , Cristalização , Cristalografia por Raios X , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Modelos Moleculares , Testes de Neutralização , Pandemias , Peptidil Dipeptidase A/química , Peptidil Dipeptidase A/metabolismo , Plasma/imunologia , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismoRESUMO
Exosomes have been extensively studied as liquid biopsy biomarkers in the past decade. However, the origin and molecular heterogeneity of exosomes hinder the research development moving from proof-of-concept to clinical applications. Herein, we report an integrated microfluidic platform termed Sub-ExoProfile chip, to achieve the selective isolation and subsequent proteomic profiling of multiplex exosome subpopulations simultaneously. The Sub-ExoProfile chip comprises three cylindrical self-assembled nanopillars, on which specific exosome capture antibodies (CD81, EpCAM, HER2) were immobilized to capture and sort different exosome subpopulations. It is worth noting that the 3D porous nanopillars afford enhanced interface binding efficiency; thus, a tumor-specific exosome subpopulation with lowly-expressed surface marker was still isolated with satisfactory capture efficiency. Moreover, the amphiphilic mesoporous silica nanoparticle self-assembled nanopillars also served as a nanoreactor for the enrichment and in situ digestion of exosomal proteins, providing improved performance for the mass-spectrometry based molecular characterization of exosome subpopulations. The Sub-ExoProfile chip was investigated on standard exosome samples from different breast cancer cell lines. The isolation and quantitative detection of exosome subpopulations were in line with the molecular subtype of breast cancer cell lines, and the molecular makeup was confirmed using classic microplate ELISA. Clinical samples from HER2-positive and triple-negative breast cancer patients were also examined using the Sub-ExoProfile chip. The quantitative results of three exosome subpopulations distinguished the three subtypes of breast cancer significantly. Most importantly, the molecular characterization of three exosome subpopulations revealed that the distinct exosome subpopulation participated in a different signaling pathway and performed distinct biological functions. It is envisioned that the analysis of the exosome subpopulation on the Sub-ExoProfile chip will facilitate the screening of tumor-specific exosomal biomarkers and open a new avenue for exosome-based liquid biopsy.
Assuntos
Neoplasias da Mama , Exossomos , Biomarcadores Tumorais/análise , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/metabolismo , Molécula de Adesão da Célula Epitelial/análise , Exossomos/metabolismo , Feminino , Humanos , Proteômica , Dióxido de Silício/análiseRESUMO
Despite advancements in antiretroviral therapy (ART) suppressing HIV-1 replication, existing antiviral drugs pose limitations, including lifelong medication, frequent administration, side effects and viral resistance, necessitating novel HIV-1 treatment approaches. CD4, pivotal for HIV-1 entry, poses challenges for drug development due to neutralization and cytotoxicity concerns. Nevertheless, Ibalizumab, the sole approved CD4-specific antibody for HIV-1 treatment, reignites interest in exploring alternative anti-HIV targets, emphasizing CD4's potential value for effective drug development. Here, we explore anti-CD4 nanobodies, particularly Nb457 from a CD4-immunized alpaca. Nb457 displays high potency and broad-spectrum activity against HIV-1, surpassing Ibalizumab's efficacy. Strikingly, engineered trimeric Nb457 nanobodies achieve complete inhibition against live HIV-1, outperforming Ibalizumab and parental Nb457. Structural analysis unveils Nb457-induced CD4 conformational changes impeding viral entry. Notably, Nb457 demonstrates therapeutic efficacy in humanized female mouse models. Our findings highlight anti-CD4 nanobodies as promising HIV-1 therapeutics, with potential implications for advancing clinical treatment against this global health challenge.
Assuntos
Antígenos CD4 , Camelídeos Americanos , Anticorpos Anti-HIV , Infecções por HIV , HIV-1 , Anticorpos de Domínio Único , HIV-1/imunologia , HIV-1/efeitos dos fármacos , Anticorpos de Domínio Único/farmacologia , Anticorpos de Domínio Único/imunologia , Animais , Antígenos CD4/imunologia , Antígenos CD4/metabolismo , Humanos , Infecções por HIV/imunologia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Camelídeos Americanos/imunologia , Anticorpos Anti-HIV/imunologia , Anticorpos Anti-HIV/farmacologia , Camundongos , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Conformação Proteica , Feminino , Internalização do Vírus/efeitos dos fármacos , Células HEK293 , Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/uso terapêutico , Anticorpos MonoclonaisRESUMO
Striking number of mutations found in the spike protein of recently emerged SARS-CoV-2 Omicron subvariants BA.1, BA.2, BA.3 and BA.4/5 has raised serious concerns regarding the escape from current antibody therapies and vaccine protection. Here, we conducted comprehensive analysis on the extent of two major Omicron lineages BA.1/BA.1.1 and BA.2 to escape neutralization from the therapeutic antibodies approved by the regulatory authorities and convalescent plasma from SARS-CoV-2 patients infected during initial wave of pandemic in early 2020. We showed that Omicron BA.1/BA.1.1 were the most resistant in both magnitude and breadth against antibodies and convalescent plasma, followed by Beta, BA.2, Gamma, Delta and Alpha. While the majority of therapeutic antibodies lost binding and neutralization to Omicron variants, BRII combo (BRII-196 + BRII-198), S309, and AZ combo (COV2-2196 + COV2-2130) maintained neutralization despite of reduction due to either conserved epitope or combinational effect between the two designated antibodies. A single intraperitoneal injection of BRII combo as a prophylactic treatment protected animals from Omicron infection. Treated animals manifested normal body weight, survived infection up to 14 days, undetectable levels of infectious viruses in the lungs, and reduced lung pathology compared to the controls. Analyzing ACE2 from diverse host species showed that Omicron variants acquired ability to use mouse ACE2 for entry. These results demonstrate major antigenic shifts and potentially broadening the host range of two major Omicron lineages BA.1/BA.1.1 and BA.2, posing serious challenges to current antibody therapies and vaccine protection as well as increasing danger of spillover into the wildlife.
Assuntos
Enzima de Conversão de Angiotensina 2 , COVID-19 , Animais , Animais Selvagens , Anticorpos Monoclonais , Anticorpos Neutralizantes , COVID-19/terapia , Imunização Passiva , Camundongos , SARS-CoV-2/genética , Soroterapia para COVID-19RESUMO
Monoclonal antibodies (mAbs) targeting the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein have demonstrated clinical efficacy in preventing or treating coronavirus disease 2019 (COVID-19), resulting in the emergency use authorization (EUA) for several SARS-CoV-2 targeting mAb by regulatory authority. However, the continuous virus evolution requires diverse mAb options to combat variants. Here we describe two fully human mAbs, amubarvimab (BRII-196) and romlusevimab (BRII-198) that bind to non-competing epitopes on the receptor binding domain (RBD) of spike protein and effectively neutralize SARS-CoV-2 variants. A YTE modification was introduced to the fragment crystallizable (Fc) region of both mAbs to prolong serum half-life and reduce effector function. The amubarvimab and romlusevimab combination retained activity against most mutations associated with reduced susceptibility to previously authorized mAbs and against variants containing amino acid substitutions in their epitope regions. Consistently, the combination of amubarvimab and romlusevimab effectively neutralized a wide range of viruses including most variants of concern and interest in vitro. In a Syrian golden hamster model of SARS-CoV-2 infection, animals receiving combination of amubarvimab and romlusevimab either pre- or post-infection demonstrated less weight loss, significantly decreased viral load in the lungs, and reduced lung pathology compared to controls. These preclinical findings support their development as an antibody cocktail therapeutic option against COVID-19 in the clinic.
Assuntos
Tratamento Farmacológico da COVID-19 , SARS-CoV-2 , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais , Epitopos , Humanos , Testes de Neutralização , Glicoproteína da Espícula de CoronavírusRESUMO
As SARS-CoV-2 Omicron and other variants of concern (VOCs) continue spreading worldwide, development of antibodies and vaccines to confer broad and protective activity is a global priority. Here, we report on the identification of a special group of nanobodies from immunized alpaca with potency against diverse VOCs including Omicron subvariants BA.1, BA.2 and BA.4/5, SARS-CoV-1, and major sarbecoviruses. Crystal structure analysis of one representative nanobody, 3-2A2-4, discovers a highly conserved epitope located between the cryptic and the outer face of the receptor binding domain (RBD), distinctive from the receptor ACE2 binding site. Cryo-EM and biochemical evaluation reveal that 3-2A2-4 interferes structural alteration of RBD required for ACE2 binding. Passive delivery of 3-2A2-4 protects K18-hACE2 mice from infection of authentic SARS-CoV-2 Delta and Omicron. Identification of these unique nanobodies will inform the development of next generation antibody therapies and design of pan-sarbecovirus vaccines.
Assuntos
COVID-19 , Camelídeos Americanos , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave , Anticorpos de Domínio Único , Animais , Camundongos , SARS-CoV-2 , Enzima de Conversão de Angiotensina 2 , COVID-19/prevenção & controle , Anticorpos Neutralizantes , Glicoproteína da Espícula de Coronavírus , Anticorpos AntiviraisRESUMO
With the rapid emergence and spread of SARS-CoV-2 variants, development of vaccines with broad and potent protectivity has become a global priority. Here, we designed a lipid nanoparticle-encapsulated, nucleoside-unmodified mRNA (mRNA-LNP) vaccine encoding the trimerized receptor-binding domain (RBD trimer) and showed its robust capability in inducing broad and protective immune responses against wild-type and major variants of concern (VOCs) in the mouse model of SARS-CoV-2 infection. The protectivity was correlated with RBD-specific B cell responses especially the long-lived plasma B cells in bone marrow, strong ability in triggering BCR clustering, and downstream signaling. Monoclonal antibodies isolated from vaccinated animals demonstrated broad and potent neutralizing activity against VOCs tested. Structure analysis of one representative antibody identified a novel epitope with a high degree of conservation among different variants. Collectively, these results demonstrate that the RBD trimer mRNA vaccine serves as a promising vaccine candidate against SARS-CoV-2 variants and beyond.
RESUMO
Although progress has been made on constructing potent bi-specific broadly neutralizing antibody (bi-bNAb), few bi-bNAbs have been evaluated against HIV-1/AIDS in non-human primates (NHPs). Here, we report the efficacy of a tandem bi-bNAb, namely BiIA-SG, in Chinese-origin rhesus macaques (CRM) against the CRM-adapted R5-tropic pathogenic SHIVSF162P3CN challenge. Pre-exposure BiIA-SG injection prevents productive viral infection in 6 of 6 CRMs with unmeasurable proviral load, T cell responses, and seroconversion. Single BiIA-SG injection, at day 1 or 3 post viral challenge, significantly reduces peak viremia, achieves undetectable setpoint viremia in 8 of 13 CRMs, and delays disease progression for years in treated CRMs. In contrast, 6 of 8 untreated CRMs develop simian AIDS within 2 years. BiIA-SG-induced long-term protection is associated with CD8+ T cells as determined by anti-CD8ß antibody depletion experiments. Our findings provide a proof-of-concept that bi-bNAb treatment elicits T cell immunity in NHPs, which warrant the clinical development of BiIA-SG for HIV-1 prevention and immunotherapy.
Assuntos
Anticorpos Biespecíficos/farmacologia , Linfócitos T CD8-Positivos/virologia , Infecções por HIV/prevenção & controle , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Animais , Anticorpos Neutralizantes/imunologia , Linfócitos T CD8-Positivos/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/imunologia , Macaca mulatta , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Vírus da Imunodeficiência Símia/imunologiaRESUMO
As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge and spread around the world, antibodies and vaccines to confer broad and potent neutralizing activity are urgently needed. Through the isolation and characterization of monoclonal antibodies (mAbs) from individuals infected with SARS-CoV-2, we identified one antibody, P36-5D2, capable of neutralizing the major SARS-CoV-2 variants of concern. Crystal and electron cryo-microscopy (cryo-EM) structure analyses revealed that P36-5D2 targeted to a conserved epitope on the receptor-binding domain of the spike protein, withstanding the three key mutations-K417N, E484K, and N501Y-found in the variants that are responsible for escape from many potent neutralizing mAbs, including some already approved for emergency use authorization (EUA). A single intraperitoneal (IP) injection of P36-5D2 as a prophylactic treatment completely protected animals from challenge of infectious SARS-CoV-2 Alpha and Beta. Treated animals manifested normal body weight and were devoid of infection-associated death up to 14 days. A substantial decrease of the infectious virus in the lungs and brain, as well as reduced lung pathology, was found in these animals compared to the controls. Thus, P36-5D2 represents a new and desirable human antibody against the current and emerging SARS-CoV-2 variants.
Assuntos
Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/farmacologia , Anticorpos Antivirais/farmacologia , Tratamento Farmacológico da COVID-19 , SARS-CoV-2/efeitos dos fármacos , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/química , Anticorpos Antivirais/imunologia , Células HEK293 , Humanos , Imunização Passiva , CamundongosRESUMO
Understanding the mechanism for antibody neutralization of SARS-CoV-2 is critical for the development of effective therapeutics and vaccines. We recently isolated a large number of monoclonal antibodies from SARS-CoV-2 infected individuals. Here we select the top three most potent yet variable neutralizing antibodies for in-depth structural and functional analyses. Crystal structural comparisons reveal differences in the angles of approach to the receptor binding domain (RBD), the size of the buried surface areas, and the key binding residues on the RBD of the viral spike glycoprotein. One antibody, P2C-1F11, most closely mimics binding of receptor ACE2, displays the most potent neutralizing activity in vitro and conferred strong protection against SARS-CoV-2 infection in Ad5-hACE2-sensitized mice. It also occupies the largest binding surface and demonstrates the highest binding affinity to RBD. More interestingly, P2C-1F11 triggers rapid and extensive shedding of S1 from the cell-surface expressed spike glycoprotein, with only minimal such effect by the remaining two antibodies. These results offer a structural and functional basis for potent neutralization via disruption of the very first and critical steps for SARS-CoV-2 cell entry.
Assuntos
Enzima de Conversão de Angiotensina 2/química , Enzima de Conversão de Angiotensina 2/metabolismo , Anticorpos Monoclonais/química , Anticorpos Neutralizantes/imunologia , COVID-19/imunologia , SARS-CoV-2/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais/imunologia , Sítios de Ligação , COVID-19/virologia , Modelos Animais de Doenças , Epitopos , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Receptores Virais/imunologia , Receptores Virais/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Internalização do Vírus , Tratamento Farmacológico da COVID-19RESUMO
Neutralizing antibodies (nAbs) to SARS-CoV-2 hold powerful potentials for clinical interventions against COVID-19 disease. However, their common genetic and biologic features remain elusive. Here we interrogate a total of 165 antibodies from eight COVID-19 patients, and find that potent nAbs from different patients have disproportionally high representation of IGHV3-53/3-66 usage, and therefore termed as public antibodies. Crystal structural comparison of these antibodies reveals they share similar angle of approach to RBD, overlap in buried surface and binding residues on RBD, and have substantial spatial clash with receptor angiotensin-converting enzyme-2 (ACE2) in binding to RBD. Site-directed mutagenesis confirms these common binding features although some minor differences are found. One representative antibody, P5A-3C8, demonstrates extraordinarily protective efficacy in a golden Syrian hamster model against SARS-CoV-2 infection. However, virus escape analysis identifies a single natural mutation in RBD, namely K417N found in B.1.351 variant from South Africa, abolished the neutralizing activity of these public antibodies. The discovery of public antibodies and shared escape mutation highlight the intricate relationship between antibody response and SARS-CoV-2, and provide critical reference for the development of antibody and vaccine strategies to overcome the antigenic variation of SARS-CoV-2.
Assuntos
Enzima de Conversão de Angiotensina 2/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Receptores Virais/imunologia , SARS-CoV-2/imunologia , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , Sítios de Ligação/imunologia , COVID-19/imunologia , Cricetinae , Modelos Animais de Doenças , Epitopos/imunologia , Feminino , Humanos , Masculino , Testes de Neutralização , Receptores de Antígenos de Linfócitos B/imunologia , Glicoproteína da Espícula de Coronavírus/imunologiaRESUMO
Neutralizing monoclonal antibodies (nAbs) to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) represent promising candidates for clinical intervention against coronavirus disease 2019 (COVID-19). We isolated a large number of nAbs from SARS-CoV-2-infected individuals capable of disrupting proper interaction between the receptor binding domain (RBD) of the viral spike (S) protein and the receptor angiotensin converting enzyme 2 (ACE2). However, the structural basis for their potent neutralizing activity remains unclear. Here, we report cryo-EM structures of the ten most potent nAbs in their native full-length IgG-form or in both IgG-form and Fab-form bound to the trimeric S protein of SARS-CoV-2. The bivalent binding of the full-length IgG is found to associate with more RBDs in the "up" conformation than the monovalent binding of Fab, perhaps contributing to the enhanced neutralizing activity of IgG and triggering more shedding of the S1 subunit from the S protein. Comparison of a large number of nAbs identified common and unique structural features associated with their potent neutralizing activities. This work provides a structural basis for further understanding the mechanism of nAbs, especially through revealing the bivalent binding and its correlation with more potent neutralization and the shedding of S1 subunit.
Assuntos
Anticorpos Neutralizantes/imunologia , COVID-19/imunologia , Imunoglobulina G/imunologia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Enzima de Conversão de Angiotensina 2/imunologia , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/ultraestrutura , Anticorpos Antivirais/química , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/ultraestrutura , Interações Hospedeiro-Patógeno , Humanos , Imunoglobulina G/química , Imunoglobulina G/ultraestrutura , Modelos Moleculares , Conformação Proteica , Multimerização Proteica , SARS-CoV-2/fisiologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/ultraestruturaRESUMO
The development of a safe and effective vaccine against SARS-CoV-2, the causative agent of pandemic coronavirus disease-2019 (COVID-19), is a global priority. Here, we aim to develop novel SARS-CoV-2 vaccines based on a derivative of less commonly used rare adenovirus serotype AdC68 vector. Three vaccine candidates were constructed expressing either the full-length spike (AdC68-19S) or receptor-binding domain (RBD) with two different signal sequences (AdC68-19RBD and AdC68-19RBDs). Single-dose intramuscular immunization induced robust and sustained binding and neutralizing antibody responses in BALB/c mice up to 40 weeks after immunization, with AdC68-19S being superior to AdC68-19RBD and AdC68-19RBDs. Importantly, immunization with AdC68-19S induced protective immunity against high-dose challenge with live SARS-CoV-2 in a golden Syrian hamster model of SARS-CoV-2 infection. Vaccinated animals demonstrated dramatic decreases in viral RNA copies and infectious virus in the lungs, as well as reduced lung pathology compared to the control animals. Similar protective effects were also found in rhesus macaques. Taken together, these results confirm that AdC68-19S can induce protective immune responses in experimental animals, meriting further development toward a human vaccine against SARS-CoV-2.
Assuntos
Vacinas contra Adenovirus/administração & dosagem , Vacinas contra COVID-19/administração & dosagem , COVID-19/prevenção & controle , Esquemas de Imunização , Imunogenicidade da Vacina , SARS-CoV-2/imunologia , Vacinação/métodos , Vacinas contra Adenovirus/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/virologia , Vacinas contra COVID-19/imunologia , Cricetinae , Modelos Animais de Doenças , Feminino , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pan troglodytes , RNA Viral/sangue , Glicoproteína da Espícula de Coronavírus/imunologia , Transfecção , Resultado do TratamentoRESUMO
Zika virus (ZIKV) specific neutralizing antibodies hold great promise for antibody-based interventions and vaccine design against ZIKV infection. However, their development in infected patients remains unclear. Here, we applied next-generation sequencing (NGS) to probe the dynamic development of a potent and protective ZIKV E DIII-specific antibody ZK2B10 isolated from a ZIKV convalescent individual. The unbiased repertoire analysis showed dramatic changes in the usage of antibody variable region germline genes. However, lineage tracing of ZK2B10 revealed limited somatic hypermutation and transient expansion during the 12 months following the onset of symptoms. The NGS-derived, germline-like ZK2B10 somatic variants neutralized ZIKV potently and protected mice from lethal challenge of ZIKV without detectable cross-reactivity with Dengue virus (DENV). Site-directed mutagenesis identified two residues within the λ chain, N31 and S91, that are essential to the functional maturation of ZK2B10. The repertoire and lineage features unveiled here will help elucidate the developmental process and protective potential of E DIII-directed antibodies against ZIKV infection.
Assuntos
Anticorpos Antivirais/imunologia , Células Germinativas/imunologia , Células Germinativas/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Infecção por Zika virus/genética , Infecção por Zika virus/imunologia , Zika virus/imunologia , Animais , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/química , Formação de Anticorpos/genética , Formação de Anticorpos/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biologia Computacional/métodos , Convalescença , Reações Cruzadas , Variação Genética , Sequenciamento de Nucleotídeos em Larga Escala , Interações Hospedeiro-Patógeno/genética , Humanos , Camundongos , Testes de Neutralização , Filogenia , Relação Estrutura-Atividade , Infecção por Zika virus/virologiaRESUMO
We previously reported a human monoclonal antibody, ZK2B10, capable of protection against Zika virus (ZIKV) infection and microcephaly in developing mouse embryos. Here, we report the structural features and mechanism of action of ZK2B10. The crystal structure at a resolution of 2.32 Å revealed that the epitope is located on the lateral ridge of DIII of the envelope glycoprotein. Cryo-EM structure with mature ZIKV showed that the antibody binds to DIIIs around the icosahedral 2-fold, 3-fold, and 5-fold axes, a distinct feature compared to those reported for DIII-specific antibodies. The binding of ZK2B10 to ZIKV has no detectable effect on viral attachment to target cells or on conformational changes of the E glycoprotein in the acidic environment, suggesting that ZK2B10 functions at steps between the formation of the fusion intermediate and membrane fusion. These results provide structural and mechanistic insights into how ZK2B10 mediates protection against ZIKV infection.
Assuntos
Anticorpos Neutralizantes/química , Anticorpos Antivirais/química , Proteínas do Envelope Viral/química , Zika virus/química , Animais , Chlorocebus aethiops , Cristalografia por Raios X , Células HEK293 , Humanos , Células VeroRESUMO
Zika virus (ZIKV) is a mosquito-transmitted flavivirus that is generally benign in humans. However, an emergent strain of ZIKV has become widespread, causing severe pre- and post-natal neurological defects. There is now an urgent need for prophylactic and therapeutic agents. To address this, we investigated six human monoclonal antibodies with ZIKV epitope specificity and neutralizing activity in mouse models of ZIKV infection and microcephaly. A single intraperitoneal injection of these antibodies conveyed distinct levels of adult and in utero protection from ZIKV infection, which closely mirrored their respective in vitro neutralizing activities. One antibody, ZK2B10, showed the most potent neutralization activity, completely protected uninfected mice, and markedly reduced tissue pathology in infected mice. Thus, ZK2B10 is a promising candidate for the development of antibody-based interventions and informs the rational design of ZIKV vaccine.
Assuntos
Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/farmacologia , Anticorpos Antivirais/farmacologia , Embrião de Mamíferos/embriologia , Microcefalia/tratamento farmacológico , Complicações Infecciosas na Gravidez/tratamento farmacológico , Infecção por Zika virus/tratamento farmacológico , Zika virus/metabolismo , Animais , Modelos Animais de Doenças , Embrião de Mamíferos/patologia , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Humanos , Camundongos , Camundongos Endogâmicos ICR , Microcefalia/metabolismo , Microcefalia/patologia , Gravidez , Complicações Infecciosas na Gravidez/metabolismo , Complicações Infecciosas na Gravidez/patologia , Infecção por Zika virus/metabolismo , Infecção por Zika virus/patologiaRESUMO
The causal link between Zika virus (ZIKV) infection and microcephaly has raised alarm worldwide. Microglial hyperplasia, reactive gliosis, and myelination delay have been reported in ZIKV-infected microcephalic fetuses. However, whether and how ZIKV infection affects glial cell development remain unclear. Here we show that ZIKV infection of embryos at the later stage of development causes severe microcephaly after birth. ZIKV infects the glial progenitors during brain development. Specifically, ZIKV infection disturbs the proliferation and differentiation of the oligodendrocyte progenitor cells and leads to the abolishment of oligodendrocyte development. More importantly, a single intraperitoneal injection of pregnant mice with a human monoclonal neutralizing antibody provides full protection against ZIKV infection and its associated damages in the developing fetuses. Our results not only provide more insights into the pathogenesis of ZIKV infection, but also present a new model for the preclinical test of prophylactic and therapeutic agents against ZIKV infection.