RESUMO
The development of an effective human immunodeficiency virus (HIV-1) vaccine is a high global health priority. Soluble native-like HIV-1 envelope glycoprotein trimers (Env), including those based on the SOSIP design, have shown promise as vaccine candidates by inducing neutralizing antibody responses against the autologous virus in animal models. However, to overcome HIV-1's extreme diversity a vaccine needs to induce broadly neutralizing antibodies (bNAbs). Such bNAbs can protect non-human primates (NHPs) and humans from infection. The prototypic BG505 SOSIP.664 immunogen is based on the BG505 env sequence isolated from an HIV-1-infected infant from Kenya who developed a bNAb response. Studying bNAb development during natural HIV-1 infection can inform vaccine design, however, it is unclear to what extent vaccine-induced antibody responses to Env are comparable to those induced by natural infection. Here, we compared Env antibody responses in BG505 SOSIP-immunized NHPs with those in BG505 SHIV-infected NHPs, by analyzing monoclonal antibodies (mAbs). We observed three major differences between BG505 SOSIP immunization and BG505 SHIV infection. First, SHIV infection resulted in more clonal expansion and less antibody diversity compared to SOSIP immunization, likely because of higher and/or prolonged antigenic stimulation and increased antigen diversity during infection. Second, while we retrieved comparatively fewer neutralizing mAbs (NAbs) from SOSIP-immunized animals, these NAbs targeted more diverse epitopes compared to NAbs from SHIV-infected animals. However, none of the NAbs, either elicited by vaccination or infection, showed any breadth. Finally, SOSIP immunization elicited antibodies against the base of the trimer, while infection did not, consistent with the base being placed onto the virus membrane in the latter setting. Together these data provide new insights into the antibody response against BG505 Env during infection and immunization and limitations that need to be overcome to induce better responses after vaccination.
Assuntos
Vacinas contra a AIDS/administração & dosagem , Anticorpos Neutralizantes/imunologia , Epitopos/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Vacinas contra a AIDS/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Formação de Anticorpos , Antígenos Virais/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/virologia , Humanos , Imunização , Lactente , Quênia , Primatas , Multimerização Proteica , Síndrome de Imunodeficiência Adquirida dos Símios/prevenção & controle , Vírus da Imunodeficiência Símia/imunologia , VacinaçãoRESUMO
Previous studies in our laboratory characterized a panel of highly mutated HIV-specific conformational epitope-targeting antibodies (Abs) from a panel of HIV-infected long-term nonprogressors (LTNPs). Despite binding HIV envelope protein and having a high number of somatic amino acid mutations, these Abs had poor neutralizing activity. Because of the evidence of antigen-driven selection and the long CDR3 region (21 amino acids [aa]), we further characterized the epitope targeting of monoclonal Ab (MAb) 76-Q3-2C6 (2C6). We confirmed that 2C6 binds preferentially to trimeric envelope and recognizes the clades A, B, and C SOSIP trimers. 2C6 binds gp140 constructs of clades A, B, C, and D, suggesting a conserved binding site that we localized to the ectodomain of gp41. Ab competition with MAb 50-69 suggested this epitope localizes near aa 579 to 613 (referenced to HXB2 gp160). Peptide library scanning showed consistent binding in this region but to only a single peptide. Lack of overlapping peptide binding supported a nonlinear epitope structure. The significance of this site is supported by 2C6 having Ab-dependent cell cytotoxicity (ADCC) against envelope proteins from two clades. Using 2C6 and variants, alanine scanning mutagenesis identified three amino acids (aa 592, 595, and 596) in the overlapping region of the previously identified peptide. Additional amino acids at sites 524 and 579 were also identified, helping explain its conformational requirement. The fact that different amino acids were included in the epitope depending on the targeted protein supports the conclusion that 2C6 targets a native conformational epitope. When we mapped these amino acids on the trimerized structure, they spanned across oligomers, supporting the notion that the epitope targeted by 2C6 lies in a recessed pocket between two gp41 oligomers. A complete understanding of the epitope specificity of ADCC-mediating Abs is essential for developing effective immunization strategies that optimize protection by these Abs.IMPORTANCE This paper further defines the function and area of the HIV trimeric envelope protein targeted by the monoclonal antibody 2C6. 2C6 binding is influenced by amino acid mutations across two separate gp41 sections of the envelope trimer. This epitope is recognized on multiple clades (variant groups of circulating viruses) of gp41, gp140 trimers, and SOSIP trimers. For the clades tested, 2C6 has robust ADCC. As the target of 2C6 is available in the major clades of HIV and has robust ADCC activity, further definition and appreciation of targeting of antibodies similar to 2C6 during vaccine development should be considered.
Assuntos
Anticorpos Monoclonais/farmacologia , Epitopos/imunologia , Proteína gp41 do Envelope de HIV/química , Infecções por HIV/imunologia , HIV-1/imunologia , Motivos de Aminoácidos , Anticorpos Neutralizantes/farmacologia , Citotoxicidade Celular Dependente de Anticorpos , Epitopos/química , Epitopos/genética , Células HEK293 , Anticorpos Anti-HIV/farmacologia , Proteína gp41 do Envelope de HIV/genética , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Humanos , Mutação , Conformação Proteica/efeitos dos fármacosRESUMO
The prefusion conformation of human metapneumovirus fusion protein (hMPV Pre-F) is critical for eliciting the most potent neutralizing antibodies and is the preferred immunogen for an efficacious vaccine against hMPV respiratory infections. Here we show that an additional cleavage event in the F protein allows closure and correct folding of the trimer. We therefore engineered the F protein to undergo double cleavage, which enabled screening for Pre-F stabilizing substitutions at the natively folded protomer interfaces. To identify these substitutions, we developed an AI convolutional classifier that successfully predicts complex polar interactions often overlooked by physics-based methods and visual inspection. The combination of additional processing, stabilization of interface regions and stabilization of the membrane-proximal stem, resulted in a Pre-F protein vaccine candidate without the need for a heterologous trimerization domain that exhibited high expression yields and thermostability. Cryo-EM analysis shows the complete ectodomain structure, including the stem, and a specific interaction of the newly identified cleaved C-terminus with the adjacent protomer. Importantly, the protein induces high and cross-neutralizing antibody responses resulting in near complete protection against hMPV challenge in cotton rats, making the highly stable, double-cleaved hMPV Pre-F trimer an attractive vaccine candidate.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Metapneumovirus , Proteínas Virais de Fusão , Vacinas Virais , Metapneumovirus/imunologia , Metapneumovirus/genética , Animais , Anticorpos Neutralizantes/imunologia , Humanos , Anticorpos Antivirais/imunologia , Proteínas Virais de Fusão/imunologia , Proteínas Virais de Fusão/química , Proteínas Virais de Fusão/genética , Vacinas Virais/imunologia , Infecções por Paramyxoviridae/prevenção & controle , Infecções por Paramyxoviridae/imunologia , Microscopia Crioeletrônica , Engenharia de Proteínas/métodos , Sigmodontinae , Feminino , Multimerização Proteica , Modelos MolecularesRESUMO
Several studies have shown that the first encounter with influenza virus shapes the immune response to future infections or vaccinations. However, a detailed analysis of the primary antibody response is lacking as this is difficult to study in humans. It is therefore not known what the frequency and dynamics of the strain-specific hemagglutinin (HA) head- and stem-directed antibody responses are directly after primary influenza virus infection. Here, sera of twelve H1N1pdm2009 influenza virus-infected cynomolgus macaques were evaluated for HA-head and HA-stem domain antibody responses. We observed an early induction of HA-stem antibody responses, which was already decreased by day 56. In contrast, responses against the HA-head domain were low early after infection and increased at later timepoint. The HA-specific B cell repertoires in each animal showed diverse VH-gene usage with preferred VH-gene and JH-gene family usage for HA-head or HA-stem B cells but a highly diverse allelic variation within the VH-usage. HA-head B cells had shorter CDRH3s and higher VH-gene somatic hyper mutation levels relative to HA-stem B cells. In conclusion, our data suggest that HA-stem antibodies are the first to react to the infection while HA-head antibodies show a delayed response, but a greater propensity to enter the germinal center and undergo affinity maturation.
Assuntos
Vírus da Influenza A Subtipo H1N1 , Influenza Humana , Infecções por Orthomyxoviridae , Humanos , Animais , Formação de Anticorpos , Hemaglutininas , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Anticorpos AntiviraisRESUMO
[This corrects the article DOI: 10.3389/fimmu.2022.893648.].
RESUMO
The most effective treatment for HIV-1, antiretroviral therapy, suppresses viral replication and averts the disease from progression. Nonetheless, there is a need for alternative treatments as it requires daily administration with the possibility of side effects and occurrence of drug resistance. Broadly neutralizing antibodies or nanobodies targeting the HIV-1 envelope glycoprotein are explored as alternative treatment, since they mediate viral suppression and contribute to the elimination of virus-infected cells. Besides neutralization potency and breadth, Fc-mediated effector functions of bNAbs also contribute to the in vivo efficacy. In this study multivalent J3, 2E7 and 1F10 anti-HIV-1 broadly neutralizing nanobodies were generated to improve neutralization potency and IgG1 Fc fusion was utilized to gain Fc-mediated effector functions. Bivalent and trivalent nanobodies, coupled using long glycine-serine linkers, showed increased binding to the HIV-1 Env and enhanced neutralization potency compared to the monovalent variant. Fusion of an IgG1 Fc domain to J3 improved neutralization potency compared to the J3-bihead and restored Fc-mediated effector functions such as antibody-dependent cellular phagocytosis and trogocytosis, and natural killer cell activation. Due to their neutralization breadth and potency and their ability to induce effector functions these nanobody-IgG1 constructs may prove to be valuable towards alternative HIV-1 therapies.
Assuntos
Soropositividade para HIV , HIV-1 , Anticorpos de Domínio Único , Anticorpos Neutralizantes/farmacologia , Anticorpos Amplamente Neutralizantes , Anticorpos Anti-HIV , Humanos , Imunoglobulina G , Anticorpos de Domínio Único/farmacologiaRESUMO
The Lassa virus is endemic in parts of West Africa, and it causes hemorrhagic fever with high mortality. The development of a recombinant protein vaccine has been hampered by the instability of soluble Lassa virus glycoprotein complex (GPC) trimers, which disassemble into monomeric subunits after expression. Here, we use two-component protein nanoparticles consisting of trimeric and pentameric subunits to stabilize GPC in a trimeric conformation. These GPC nanoparticles present twenty prefusion GPC trimers on the surface of an icosahedral particle. Cryo-EM studies of GPC nanoparticles demonstrated a well-ordered structure and yielded a high-resolution structure of an unliganded GPC. These nanoparticles induced potent humoral immune responses in rabbits and protective immunity against the lethal Lassa virus challenge in guinea pigs. Additionally, we isolated a neutralizing antibody that mapped to the putative receptor-binding site, revealing a previously undefined site of vulnerability. Collectively, these findings offer potential approaches to vaccine and therapeutic design for the Lassa virus.
Assuntos
Febre Lassa , Nanopartículas , Cobaias , Coelhos , Animais , Vírus Lassa/química , Anticorpos Neutralizantes , Febre Lassa/prevenção & controle , Glicoproteínas , Vacinas SintéticasRESUMO
The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has had a large impact on global health, travel, and economy. Therefore, preventative and therapeutic measures are urgently needed. Here, we isolated monoclonal antibodies from three convalescent coronavirus disease 2019 (COVID-19) patients using a SARS-CoV-2 stabilized prefusion spike protein. These antibodies had low levels of somatic hypermutation and showed a strong enrichment in VH1-69, VH3-30-3, and VH1-24 gene usage. A subset of the antibodies was able to potently inhibit authentic SARS-CoV-2 infection at a concentration as low as 0.007 micrograms per milliliter. Competition and electron microscopy studies illustrate that the SARS-CoV-2 spike protein contains multiple distinct antigenic sites, including several receptor-binding domain (RBD) epitopes as well as non-RBD epitopes. In addition to providing guidance for vaccine design, the antibodies described here are promising candidates for COVID-19 treatment and prevention.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Pneumonia Viral/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto , Idoso , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Afinidade de Anticorpos , Antígenos Virais/imunologia , Subpopulações de Linfócitos B/imunologia , Anticorpos Amplamente Neutralizantes/imunologia , COVID-19 , Linhagem Celular Tumoral , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/terapia , Epitopos/imunologia , Feminino , Humanos , Memória Imunológica , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Pneumonia Viral/terapia , Domínios Proteicos , Domínios e Motivos de Interação entre Proteínas/imunologia , Receptores de Coronavírus , Receptores Virais/metabolismo , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/químicaRESUMO
So far, the development of a human immunodeficiency virus (HIV) vaccine has been unsuccessful. However, recent progress in the field of broadly neutralizing antibodies (bNAbs) has reinvigorated the search for an HIV vaccine. bNAbs develop in a minority of HIV infected individuals and passive transfer of these bNAbs to non-human primates provides protection from HIV infection. Studies in a number of HIV infected individuals on bNAb maturation alongside viral evolution and escape have shed light on the features important for bNAb elicitation. Here we review the observations from these studies, and how they influence the rational design of HIV vaccines.