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1.
Cell ; 158(3): 481-91, 2014 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-25065977

RESUMO

Development of strategies for induction of HIV-1 broadly neutralizing antibodies (bnAbs) by vaccines is a priority. Determining the steps of bnAb induction in HIV-1-infected individuals who make bnAbs is a key strategy for immunogen design. Here, we study the B cell response in a bnAb-producing individual and report cooperation between two B cell lineages to drive bnAb development. We isolated a virus-neutralizing antibody lineage that targeted an envelope region (loop D) and selected virus escape mutants that resulted in both enhanced bnAb lineage envelope binding and escape mutant neutralization-traits associated with increased B cell antigen drive. Thus, in this individual, two B cell lineages cooperated to induce the development of bnAbs. Design of vaccine immunogens that simultaneously drive both helper and broadly neutralizing B cell lineages may be important for vaccine-induced recapitulation of events that transpire during the maturation of neutralizing antibodies in HIV-1-infected individuals.


Assuntos
Vacinas contra a AIDS/química , Vacinas contra a AIDS/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , HIV-1/fisiologia , Sequência de Aminoácidos , Linfócitos B/imunologia , Evasão da Resposta Imune , Modelos Moleculares , Dados de Sequência Molecular , Mutação , Alinhamento de Sequência , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia
2.
Immunity ; 49(6): 1162-1174.e8, 2018 12 18.
Artigo em Inglês | MEDLINE | ID: mdl-30552024

RESUMO

Elicitation of VRC01-class broadly neutralizing antibodies (bnAbs) is an appealing approach for a preventative HIV-1 vaccine. Despite extensive investigations, strategies to induce VRC01-class bnAbs and overcome the barrier posed by the envelope N276 glycan have not been successful. Here, we inferred a high-probability unmutated common ancestor (UCA) of the VRC01 lineage and reconstructed the stages of lineage maturation. Env immunogens designed on reverted VRC01-class bnAbs bound to VRC01 UCA with affinity sufficient to activate naive B cells. Early mutations defined maturation pathways toward limited or broad neutralization, suggesting that focusing the immune response is likely required to steer B cell maturation toward the development of neutralization breadth. Finally, VRC01 lineage bnAbs with long CDR H3s overcame the HIV-1 N276 glycan barrier without shortening their CDR L1, revealing a solution for broad neutralization in which the heavy chain, not CDR L1, is the determinant to accommodate the N276 glycan.


Assuntos
Vacinas contra a AIDS/imunologia , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Polissacarídeos/imunologia , Vacinas contra a AIDS/administração & dosagem , Vacinas contra a AIDS/genética , Sequência de Aminoácidos , Anticorpos Monoclonais/classificação , Anticorpos Monoclonais/genética , Anticorpos Neutralizantes/classificação , Anticorpos Neutralizantes/genética , Linfócitos B/imunologia , Linfócitos B/metabolismo , Sítios de Ligação/genética , Anticorpos Amplamente Neutralizantes , Antígenos CD4/genética , Antígenos CD4/imunologia , Antígenos CD4/metabolismo , Anticorpos Anti-HIV , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp120 do Envelope de HIV/metabolismo , Infecções por HIV/terapia , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Humanos , Filogenia , Polissacarídeos/metabolismo , Homologia de Sequência de Aminoácidos
3.
J Immunol ; 198(3): 1047-1055, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28011932

RESUMO

In the RV144 gp120 HIV vaccine trial, decreased transmission risk was correlated with Abs that reacted with a linear epitope at a lysine residue at position 169 (K169) in the HIV-1 envelope (Env) V2 region. The K169 V2 response was restricted to Abs bearing Vλ rearrangements that expressed aspartic acid/glutamic acid in CDR L2. The AE.A244 gp120 in AIDSVAX B/E also bound to the unmutated ancestor of a V2-glycan broadly neutralizing Ab, but this Ab type was not induced in the RV144 trial. In this study, we sought to determine whether immunodominance of the V2 linear epitope could be overcome in the absence of human Vλ rearrangements. We immunized IgH- and Igκ-humanized mice with the AE.A244 gp120 Env. In these mice, the V2 Ab response was focused on a linear epitope that did not include K169. V2 Abs were isolated that used the same human VH gene segment as an RV144 V2 Ab but paired with a mouse λ L chain. Structural characterization of one of these V2 Abs revealed how the linear V2 epitope could be engaged, despite the lack of aspartic acid/glutamic acid encoded in the mouse repertoire. Thus, despite the absence of the human Vλ locus in these humanized mice, the dominance of Vλ pairing with human VH for HIV-1 Env V2 recognition resulted in human VH pairing with mouse λ L chains instead of allowing otherwise subdominant V2-glycan broadly neutralizing Abs to develop.


Assuntos
Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , HIV-1/imunologia , Vacinas contra a AIDS/imunologia , Motivos de Aminoácidos , Animais , Anticorpos Neutralizantes/imunologia , Epitopos , Humanos , Cadeias Pesadas de Imunoglobulinas/imunologia , Cadeias lambda de Imunoglobulina/imunologia , Camundongos
4.
Nature ; 496(7446): 469-76, 2013 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-23552890

RESUMO

Current human immunodeficiency virus-1 (HIV-1) vaccines elicit strain-specific neutralizing antibodies. However, cross-reactive neutralizing antibodies arise in approximately 20% of HIV-1-infected individuals, and details of their generation could provide a blueprint for effective vaccination. Here we report the isolation, evolution and structure of a broadly neutralizing antibody from an African donor followed from the time of infection. The mature antibody, CH103, neutralized approximately 55% of HIV-1 isolates, and its co-crystal structure with the HIV-1 envelope protein gp120 revealed a new loop-based mechanism of CD4-binding-site recognition. Virus and antibody gene sequencing revealed concomitant virus evolution and antibody maturation. Notably, the unmutated common ancestor of the CH103 lineage avidly bound the transmitted/founder HIV-1 envelope glycoprotein, and evolution of antibody neutralization breadth was preceded by extensive viral diversification in and near the CH103 epitope. These data determine the viral and antibody evolution leading to induction of a lineage of HIV-1 broadly neutralizing antibodies, and provide insights into strategies to elicit similar antibodies by vaccination.


Assuntos
Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/imunologia , Evolução Molecular , Anticorpos Anti-HIV/química , Anticorpos Anti-HIV/imunologia , HIV-1/química , HIV-1/imunologia , Vacinas contra a AIDS/imunologia , África , Sequência de Aminoácidos , Anticorpos Monoclonais/química , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/genética , Antígenos CD4/química , Antígenos CD4/imunologia , Linhagem da Célula , Células Cultivadas , Células Clonais/citologia , Reações Cruzadas/imunologia , Cristalografia por Raios X , Epitopos/química , Epitopos/imunologia , Anticorpos Anti-HIV/genética , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/classificação , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Mutação , Testes de Neutralização , Filogenia , Estrutura Terciária de Proteína
5.
PLoS Pathog ; 11(8): e1005042, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26237403

RESUMO

HIV-1 mucosal transmission begins with virus or virus-infected cells moving through mucus across mucosal epithelium to infect CD4+ T cells. Although broadly neutralizing antibodies (bnAbs) are the type of HIV-1 antibodies that are most likely protective, they are not induced with current vaccine candidates. In contrast, antibodies that do not neutralize primary HIV-1 strains in the TZM-bl infection assay are readily induced by current vaccine candidates and have also been implicated as secondary correlates of decreased HIV-1 risk in the RV144 vaccine efficacy trial. Here, we have studied the capacity of anti-Env monoclonal antibodies (mAbs) against either the immunodominant region of gp41 (7B2 IgG1), the first constant region of gp120 (A32 IgG1), or the third variable loop (V3) of gp120 (CH22 IgG1) to modulate in vivo rectal mucosal transmission of a high-dose simian-human immunodeficiency virus (SHIV-BaL) in rhesus macaques. 7B2 IgG1 or A32 IgG1, each containing mutations to enhance Fc function, was administered passively to rhesus macaques but afforded no protection against productive clinical infection while the positive control antibody CH22 IgG1 prevented infection in 4 of 6 animals. Enumeration of transmitted/founder (T/F) viruses revealed that passive infusion of each of the three antibodies significantly reduced the number of T/F genomes. Thus, some antibodies that bind HIV-1 Env but fail to neutralize virus in traditional neutralization assays may limit the number of T/F viruses involved in transmission without leading to enhancement of viral infection. For one of these mAbs, gp41 mAb 7B2, we provide the first co-crystal structure in complex with a common cyclical loop motif demonstrated to be critical for infection by other retroviruses.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/prevenção & controle , Vírus da Imunodeficiência Símia/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Imunofluorescência , HIV-1/imunologia , Humanos , Mucosa Intestinal/virologia , Macaca mulatta , Conformação Proteica , Reto , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ressonância de Plasmônio de Superfície , Proteínas do Envelope Viral/química
6.
J Virol ; 88(6): 3329-39, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24390332

RESUMO

UNLABELLED: The development of a vaccine that can induce high titers of functional antibodies against HIV-1 remains a high priority. We have developed an adjuvant based on an oil-in-water emulsion that incorporates Toll-like receptor (TLR) ligands to test whether triggering multiple pathogen-associated molecular pattern receptors could enhance immunogenicity. Compared to single TLR agonists or other pairwise combinations, TLR7/8 and TLR9 agonists combined were able to elicit the highest titers of binding, neutralizing, and antibody-dependent cellular cytotoxicity-mediating antibodies against the protein immunogen, transmitted/founder HIV-1 envelope gp140 (B.63521). We further found that the combination of TLR7/8 and TLR9 agonists was associated with the release of CXCL10 (IP-10), suggesting that this adjuvant formulation may have optimally stimulated innate and adaptive immunity to elicit high titers of antibodies. IMPORTANCE: Combining TLR agonists in an adjuvant formulation resulted in higher antibody levels compared to an adjuvant without TLR agonists. Adjuvants that combine TLR agonists may be useful for enhancing antibody responses to HIV-1 vaccines.


Assuntos
Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Receptor 7 Toll-Like/agonistas , Receptor 8 Toll-Like/agonistas , Receptor Toll-Like 9/agonistas , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Feminino , Infecções por HIV/virologia , HIV-1/genética , Humanos , Imunização , Ligantes , Macaca mulatta , Masculino , Receptor 7 Toll-Like/imunologia , Receptor 8 Toll-Like/imunologia , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/administração & dosagem , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
7.
J Immunol ; 191(3): 1260-75, 2013 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-23825311

RESUMO

Developing an HIV-1 vaccine has been hampered by the inability of immunogens to induce broadly neutralizing Abs (BnAbs) that protect against infection. Previously, we used knockin (KI) mice expressing a prototypical gp41-specific BnAb, 2F5, to demonstrate that immunological tolerance triggered by self-reactivity of the 2F5 H chain impedes BnAb induction. In this study, we generate KI models expressing H chains from two other HIV-1 Abs, 4E10 (another self-/polyreactive, anti-gp41 BnAb) and 48d (an anti-CD4 inducible, nonpolyreactive Ab), and find a similar developmental blockade consistent with central B cell deletion in 4E10, but not in 48d VH KI mice. Furthermore, in KI strains expressing the complete 2F5 and 4E10 Abs as BCRs, we find that residual splenic B cells arrest at distinct developmental stages, yet exhibit uniformly low BCR densities, elevated basal activation, and profoundly muted responses to BCR ligation and, when captured as hybridoma mAb lines, maintain their dual (gp41/lipid) affinities and capacities to neutralize HIV-1, establishing a key role for anergy in suppressing residual 2F5- or 4E10-expressing B cells. Importantly, serum IgGs from naive 2F5 and 4E10 KI strains selectively eliminate gp41 and lipid binding, respectively, suggesting B cells expressing 2F5 or 4E10 as BCRs exhibit specificity for a distinct spectrum of host Ags, including selective interactions by 2F5 BCR(+) B cells (i.e., and not 4E10 BCR(+) B cells) with those mimicked by its gp41 neutralization epitope.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Reações Cruzadas , Anticorpos Anti-HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , Lipídeos/imunologia , Vacinas contra a AIDS/imunologia , Animais , Anticorpos Neutralizantes/biossíntese , Linfócitos B/imunologia , Anticorpos Amplamente Neutralizantes , Diferenciação Celular , Técnicas de Introdução de Genes , Anticorpos Anti-HIV/biossíntese , HIV-1/imunologia , Tolerância Imunológica , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Antígenos de Linfócitos B/metabolismo
8.
J Immunol ; 191(5): 2538-50, 2013 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-23918977

RESUMO

A goal of HIV-1 vaccine development is to elicit broadly neutralizing Abs (BnAbs). Using a knock-in (KI) model of 2F5, a human HIV-1 gp41 membrane proximal external region (MPER)-specific BnAb, we previously demonstrated that a key obstacle to BnAb induction is clonal deletion of BnAb-expressing B cells. In this study of this model, we provide a proof-of-principle that robust serum neutralizing IgG responses can be induced from pre-existing, residual, self-reactive BnAb-expressing B cells in vivo using a structurally compatible gp41 MPER immunogen. Furthermore, in CD40L-deficient 2F5 KI mice, we demonstrate that these BnAb responses are elicited via a type II T-independent pathway, coinciding with expansion and activation of transitional splenic B cells specific for 2F5's nominal gp41 MPER-binding epitope (containing the 2F5 neutralization domain ELDKWA). In contrast, constitutive production of nonneutralizing serum IgGs in 2F5 KI mice is T dependent and originates from a subset of splenic mature B2 cells that have lost their ability to bind 2F5's gp41 MPER epitope. These results suggest that residual, mature B cells expressing autoreactive BnAbs, like 2F5 as BCR, may be limited in their ability to participate in T-dependent responses by purifying selection that selectively eliminates reactivity for neutralization epitope-containing/mimicked host Ags.


Assuntos
Vacinas contra a AIDS/imunologia , Anticorpos Monoclonais/imunologia , Anticorpos Anti-HIV/imunologia , Ativação Linfocitária/imunologia , Linfócitos T/imunologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Antígenos Virais/imunologia , Linfócitos B/imunologia , Anticorpos Amplamente Neutralizantes , Ensaio de Imunoadsorção Enzimática , Técnicas de Introdução de Genes , Anticorpos Anti-HIV/sangue , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , Humanos , Camundongos , Camundongos Knockout , Testes de Neutralização
9.
J Virol ; 87(8): 4185-201, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23365441

RESUMO

Human immunodeficiency virus type 1 (HIV-1) vaccine development requires selection of appropriate envelope (Env) immunogens. Twenty HIV-1 Env glycoproteins were examined for their ability to bind human anti-HIV-1 monoclonal antibodies (MAbs) and then used as immunogens in guinea pigs to identify promising immunogens. These included five Envs derived from chronically infected individuals, each representing one of five common clades and eight consensus Envs based on these five clades, as well as the consensus of the entire HIV-1 M group, and seven transmitted/founder (T/F) Envs from clades B and C. Sera from immunized guinea pigs were tested for neutralizing activity using 36 HIV-1 Env-pseudotyped viruses. All Envs bound to CD4 binding site, membrane-proximal, and V1/V2 MAbs with similar apparent affinities, although the T/F Envs bound with higher affinity to the MAb 17b, a CCR5 coreceptor binding site antibody. However, the various Envs differed in their ability to induce neutralizing antibodies. Consensus Envs elicited the most potent responses, but neutralized only a subset of viruses, including mostly easy-to-neutralize tier 1 and some more-difficult-to-neutralize tier 2 viruses. T/F Envs elicited fewer potent neutralizing antibodies but exhibited greater breadth than chronic or consensus Envs. Finally, chronic Envs elicited the lowest level and most limited breadth of neutralizing antibodies overall. Thus, each group of Env immunogens elicited a different antibody response profile. The complementary benefits of consensus and T/F Env immunogens raise the possibility that vaccines utilizing a combination of consensus and T/F Envs may be able to induce neutralizing responses with greater breadth and potency than single Env immunogens.


Assuntos
Antígenos Virais/imunologia , Glicoproteínas/imunologia , HIV-1/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Animais , Anticorpos Neutralizantes/sangue , Afinidade de Anticorpos , Antígenos Virais/genética , Reações Cruzadas , Glicoproteínas/genética , Cobaias , Anticorpos Anti-HIV/sangue , HIV-1/genética , HIV-1/isolamento & purificação , Humanos , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
10.
Cell Host Microbe ; 32(5): 693-709.e7, 2024 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-38670093

RESUMO

A major goal of HIV-1 vaccine development is the induction of broadly neutralizing antibodies (bnAbs). Although success has been achieved in initiating bnAb B cell lineages, design of boosting immunogens that select for bnAb B cell receptors with improbable mutations required for bnAb affinity maturation remains difficult. Here, we demonstrate a process for designing boosting immunogens for a V3-glycan bnAb B cell lineage. The immunogens induced affinity-matured antibodies by selecting for functional improbable mutations in bnAb precursor knockin mice. Moreover, we show similar success in prime and boosting with nucleoside-modified mRNA-encoded HIV-1 envelope trimer immunogens, with improved selection by mRNA immunogens of improbable mutations required for bnAb binding to key envelope glycans. These results demonstrate the ability of both protein and mRNA prime-boost immunogens for selection of rare B cell lineage intermediates with neutralizing breadth after bnAb precursor expansion, a key proof of concept and milestone toward development of an HIV-1 vaccine.


Assuntos
Vacinas contra a AIDS , Anticorpos Neutralizantes , Linfócitos B , Anticorpos Anti-HIV , HIV-1 , Vacinas contra a AIDS/imunologia , Vacinas contra a AIDS/genética , Animais , Anticorpos Anti-HIV/imunologia , HIV-1/imunologia , HIV-1/genética , Camundongos , Anticorpos Neutralizantes/imunologia , Linfócitos B/imunologia , Humanos , Infecções por HIV/imunologia , Infecções por HIV/prevenção & controle , Anticorpos Amplamente Neutralizantes/imunologia , Mutação , Desenvolvimento de Vacinas , Imunização Secundária , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
11.
J Virol ; 85(3): 1340-7, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21106741

RESUMO

HIV-1 gp41 envelope antibodies, which are frequently induced in HIV-1-infected individuals, are predominantly nonneutralizing. The rare and difficult-to-induce neutralizing antibodies (2F5 and 4E10) that target gp41 membrane-proximal epitopes (MPER) are polyspecific and require lipid binding for HIV-1 neutralization. These results raise the questions of how prevalent polyreactivity is among gp41 antibodies and how the binding properties of gp41-nonneutralizing antibodies differ from those of antibodies that are broadly neutralizing. In this study, we have characterized a panel of human gp41 antibodies with binding specificities within the immunodominant cluster I (gp41 amino acids [aa] 579 to 613) or cluster II (gp41 aa 644 to 667) for reactivity to autoantigens, to the gp140 protein, and with MPER peptide-lipid conjugates. We report that while none of the gp41 cluster I antibodies studied were polyspecific, all three gp41 cluster II antibodies bound either to lipids or autoantigens, thus showing the propensity of cluster II antibodies to manifest polyreactivity. All cluster II gp41 monoclonal antibodies (MAbs), including those that were lipid reactive, failed to bind to gp41 MPER peptide-lipid complexes. Cluster II antibodies bound strongly with nanomolar binding affinity (dissociation constant [K(d)]) to oligomeric gp140 proteins, and thus, they recognize conformational epitopes on gp41 that are distinct from those of neutralizing gp41 antibodies. These results demonstrate that lipid-reactive gp41 cluster II antibodies are nonneutralizing due to their inability to bind to the relevant neutralizing epitopes on gp41.


Assuntos
Anticorpos Monoclonais/imunologia , Autoantígenos/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , Fosfolipídeos/imunologia , Humanos , Cinética , Ligação Proteica
12.
Proc Natl Acad Sci U S A ; 106(48): 20234-9, 2009 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-19906992

RESUMO

Induction of effective antibody responses against HIV-1 infection remains an elusive goal for vaccine development. Progress may require in-depth understanding of the molecular mechanisms of neutralization by monoclonal antibodies. We have analyzed the molecular actions of two rare, broadly neutralizing, human monoclonal antibodies, 4E10 and 2F5, which target the transiently exposed epitopes in the membrane proximal external region (MPER) of HIV-1 gp41 envelope during viral entry. Both have long CDR H3 loops with a hydrophobic surface facing away from the peptide epitope. We find that the hydrophobic residues of 4E10 mediate a reversible attachment to the viral membrane and that they are essential for neutralization, but not for interaction with gp41. We propose that these antibodies associate with the viral membrane in a required first step and are thereby poised to capture the transient gp41 fusion intermediate. These results bear directly on strategies for rational design of HIV-1 envelope immunogens.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Modelos Moleculares , Vacinas Virais/imunologia , Anticorpos Monoclonais/genética , Proteína gp41 do Envelope de HIV/imunologia , Humanos , Membranas Artificiais , Mutação/genética , Testes de Neutralização , Ligação Proteica/genética
13.
Cell Rep ; 38(11): 110514, 2022 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-35294883

RESUMO

The success of nucleoside-modified mRNAs in lipid nanoparticles (mRNA-LNP) as COVID-19 vaccines heralded a new era of vaccine development. For HIV-1, multivalent envelope (Env) trimer protein nanoparticles are superior immunogens compared with trimers alone for priming of broadly neutralizing antibody (bnAb) B cell lineages. The successful expression of complex multivalent nanoparticle immunogens with mRNAs has not been demonstrated. Here, we show that mRNAs can encode antigenic Env trimers on ferritin nanoparticles that initiate bnAb precursor B cell expansion and induce serum autologous tier 2 neutralizing activity in bnAb precursor VH + VL knock-in mice. Next-generation sequencing demonstrates acquisition of critical mutations, and monoclonal antibodies that neutralize heterologous HIV-1 isolates are isolated. Thus, mRNA-LNP can encode complex immunogens and may be of use in design of germline-targeting and sequential boosting immunogens for HIV-1 vaccine development.


Assuntos
Vacinas contra a AIDS , COVID-19 , HIV-1 , Nanopartículas , Animais , Anticorpos Monoclonais , Anticorpos Neutralizantes , Vacinas contra COVID-19 , Epitopos , Ferritinas/genética , Anticorpos Anti-HIV , Humanos , Lipossomos , Camundongos , RNA Mensageiro , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
14.
Cell Rep ; 36(7): 109561, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34407396

RESUMO

Elicitation of broadly neutralizing antibodies (bnAbs) by an HIV vaccine will involve priming the immune system to activate antibody precursors, followed by boosting immunizations to select for antibodies with functional features required for neutralization breadth. The higher the number of acquired mutations necessary for function, the more convoluted are the antibody developmental pathways. HIV bnAbs acquire a large number of somatic mutations, but not all mutations are functionally important. In this study, we identify a minimal subset of mutations sufficient for the function of the naturally occurring V3-glycan bnAb DH270.6. Using antibody library screening, candidate envelope immunogens that interact with DH270.6-like antibodies containing this set of key mutations are identified and selected in vitro. Our results demonstrate that less complex B cell evolutionary pathways than those naturally observed exist for the induction of HIV bnAbs by vaccination, and they establish rational approaches to identify boosting candidate immunogens.


Assuntos
Anticorpos Neutralizantes/imunologia , Linfócitos B/citologia , Linhagem da Célula , Mutação/genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Sequência de Aminoácidos , Linhagem Celular , Humanos , Mutagênese/genética , Ligação Proteica , Vacinação , Produtos do Gene env do Vírus da Imunodeficiência Humana/química
15.
bioRxiv ; 2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34401876

RESUMO

The success of nucleoside-modified mRNAs in lipid nanoparticles (mRNA-LNP) as COVID-19 vaccines heralded a new era of vaccine development. For HIV-1, multivalent envelope (Env) trimer protein nanoparticles are superior immunogens compared to trimers alone for priming of broadly neutralizing antibody (bnAb) B cell lineages. The successful expression of complex multivalent nanoparticle immunogens with mRNAs has not been demonstrated. Here we show that mRNAs can encode antigenic Env trimers on ferritin nanoparticles that initiate bnAb precursor B cell expansion and induce serum autologous tier 2 neutralizing activity in bnAb precursor VH + VL knock-in mice. Next generation sequencing demonstrated acquisition of critical mutations, and monoclonal antibodies that neutralized heterologous HIV-1 isolates were isolated. Thus, mRNA-LNP can encode complex immunogens and are of use in design of germline-targeting and sequential boosting immunogens for HIV-1 vaccine development.

16.
Immunohorizons ; 4(11): 713-728, 2020 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-33172842

RESUMO

Rab11 recycling endosomes are involved in immunological synaptic functions, but the roles of Rab11 family-interacting protein 5 (Rab11Fip5), one of the Rab11 effectors, in the immune system remain obscure. Our previous study demonstrated that RAB11FIP5 transcripts are significantly elevated in PBMCs from HIV-1-infected individuals, making broadly HIV-1-neutralizing Abs compared with those without broadly neutralizing Abs; however, the role of Rab11FiP5 in immune functions remains unclear. In this study, a RAB11FIP5 gene knockout (RAB11FIP5 -/-) mouse model was employed to study the role of Rab11Fip5 in immune responses. RAB11FIP5 -/- mice exhibited no perturbation in lymphoid tissue cell subsets, and Rab11Fip5 was not required for serum Ab induction following HIV-1 envelope immunization, Ab transcytosis to mucosal sites, or survival after influenza challenge. However, differences were observed in multiple transcripts, including cytokine genes, in lymphocyte subsets from envelope-immunized RAB11FIP5 -/- versus control mice. These included alterations in several genes in NK cells that mirrored observations in NKs from HIV-infected humans expressing less RAB11FIP5, although Rab11Fip5 was dispensable for NK cell cytolytic activity. Notably, immunized RAB11FIP5 -/- mice had lower IL4 expression in CD4+ T follicular helper cells and showed lower TNF expression in CD8+ T cells. Likewise, TNF-α production by human CD8+ T cells correlated with PBMC RAB11FIP5 expression. These observations in RAB11FIP5 -/- mice suggest a role for Rab11Fip5 in regulating cytokine responses.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/imunologia , Anticorpos Neutralizantes/imunologia , Infecções por HIV/imunologia , Transcriptoma , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Linfócitos T CD8-Positivos/imunologia , Citocinas/metabolismo , Feminino , Infecções por HIV/fisiopatologia , HIV-1/patogenicidade , Humanos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
17.
J Virol ; 82(1): 115-25, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17942537

RESUMO

Two human monoclonal antibodies (MAbs) (2F5 and 4E10) against the human immunodeficiency virus type 1 (HIV-1) envelope g41 cluster II membrane proximal external region (MPER) broadly neutralize HIV-1 primary isolates. However, these antibody specificities are rare, are not induced by Env immunization or HIV-1 infection, and are polyspecific and also react with lipids such as cardiolipin or phosphatidylserine. To probe MPER anti-gp41 antibodies that are produced in HIV-1 infection, we have made two novel murine MAbs, 5A9 and 13H11, against HIV-1 gp41 envelope that partially cross-blocked 2F5 MAb binding to Env but did not neutralize HIV-1 primary isolates or bind host lipids. Competitive inhibition assays using labeled 13H11 MAb and HIV-1-positive patient plasma samples demonstrated that cluster II 13H11-blocking plasma antibodies were made in 83% of chronically HIV-1 infected patients and were acquired between 5 to 10 weeks after acute HIV-1 infection. Both the mouse 13H11 MAb and the three prototypic cluster II human MAbs (98-6, 126-6, and 167-D) blocked 2F5 binding to gp41 epitopes to variable degrees; the combination of 98-6 and 13H11 completely blocked 2F5 binding. These data provide support for the hypothesis that in some patients, B cells make nonneutralizing cluster II antibodies that may mask or otherwise down-modulate B-cell responses to immunogenic regions of gp41 that could be recognized by B cells capable of producing antibodies like 2F5.


Assuntos
Epitopos/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/isolamento & purificação , Reações Cruzadas , Anticorpos Anti-HIV/sangue , Anticorpos Anti-HIV/metabolismo , Humanos , Cinética , Lipídeos/imunologia , Camundongos , Testes de Neutralização , Ligação Proteica
18.
Nat Commun ; 10(1): 654, 2019 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-30737386

RESUMO

Somatic mutations within antibody variable and framework regions (FWR) can alter thermostability and structural flexibility, but their impact on functional potency is unclear. Here we study thermostability and use molecular dynamics (MD) simulations to assess the role of FWR mutations during maturation of HIV-1 broadly neutralizing antibodies (bnAbs). The tested bnAbs show lower thermostability than their unmutated ancestor antibodies. FWR mutations in the Fab elbow region are frequently observed in HIV-1 bnAbs and MD simulations show that such FWR mutations alter interdomain flexibility in two HIV-1 bnAbs. In a CD4-binding site lineage, reversion mutations result in a loss of neutralization potency in an early intermediate and affinity-matured bnAb against autologous and heterologous Tier-2 viruses, respectively. Elbow region reversion mutations in a glycan-V3 bnAb modestly reduces potency against an autologous virus isolate. Thus, selection of mutations in the Fab elbow region impacts interdomain conformational flexibility and paratope plasticity during bnAb development.


Assuntos
Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/genética , Anticorpos Anti-HIV/imunologia , Mutação/genética , Varredura Diferencial de Calorimetria , Dicroísmo Circular , HIV-1/imunologia , Humanos , Simulação de Dinâmica Molecular , Testes de Neutralização , Ressonância de Plasmônio de Superfície
19.
J Virol Methods ; 143(1): 104-11, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17416428

RESUMO

Functional human immunodeficiency virus type 1 (HIV-1) env genes have been widely used for vaccine design, neutralization assays, and pathogenesis studies. However, obtaining bona fide functional env clones is a time consuming and labor intensive process. A new high throughput method has been developed to characterize HIV-1 env genes. Multiple rev/env gene cassettes were obtained from each of seven HIV-1 strains using single genome amplification (SGA) PCR. The cytomegalovirus (CMV) promoter was amplified separately by PCR. A promoter PCR (pPCR) method was developed to link both PCR products using an overlapping PCR method. Pseudovirions were generated by cotransfection of pPCR products and pSG3 Delta env backbone into 293T cells. After infecting TZM-bl cells, 75 out of 87 (86%) of the rev/env gene cassettes were functional. Pseudoviruses generated with pPCR products or corresponding plasmid DNA showed similar sensitivity to six HIV-1 positive sera and three monoclonal antibodies, suggesting neutralization properties are not altered in pPCR pseudovirions. Furthermore, sufficient amounts of pseudovirions can be obtained for a large number of neutralization assays. The new pPCR method eliminates cloning, transformation, and plasmid DNA preparation steps in the generation of HIV-1 pseudovirions. This allows for quick analysis of multiple env genes from HIV-1 infected individuals.


Assuntos
Produtos do Gene env/isolamento & purificação , Genes env , HIV-1/isolamento & purificação , Reação em Cadeia da Polimerase/métodos , Síndrome da Imunodeficiência Adquirida , Linhagem Celular , Citomegalovirus/genética , HIV-1/genética , Humanos , Regiões Promotoras Genéticas , Zâmbia
20.
J Virol Methods ; 249: 85-93, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28837840

RESUMO

Development of a HIV-1 vaccine is a major global priority. The yellow fever virus (YFV) attenuated vaccine 17D is among the most effective of currently used vaccines. However, the stability of the YFV17D vector when carrying non-flavivirus genes has been problematic. We have constructed and expressed HIV-1 Env in YFV17D with either single transmembrane (STM) or double transmembrane (DTM) YFV E protein domains for the development of anti-HIV antibodies. Here we describe modifications of the YFV17D vector such that HIV-1 Env gp120 is expressed in up to 5 passages in Vero cells. Immunization with recombinant YFV17D vector prime followed by HIV-1 CH505 TF gp120 protein boosts were able to induce neutralizing antibodies for a HIV-1 tier 1 isolate in mice. This modified YFV vector may be a starting point for constructing HIV-1 vaccine candidate priming vectors.


Assuntos
Vacinas contra a AIDS/imunologia , Proteína gp120 do Envelope de HIV/genética , HIV-1/genética , HIV-1/imunologia , Vírus da Febre Amarela/genética , Vacinas contra a AIDS/genética , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Chlorocebus aethiops , Vetores Genéticos , Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , Infecções por HIV/imunologia , Infecções por HIV/prevenção & controle , Imunização Secundária , Camundongos , Testes de Neutralização , Vacinas Atenuadas/imunologia , Vacinas de DNA/imunologia , Células Vero
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