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1.
Nat Immunol ; 23(11): 1600-1613, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36271148

RESUMO

Naïve CD8+ T cells can differentiate into effector (Teff), memory (Tmem) or exhausted (Tex) T cells. These developmental pathways are associated with distinct transcriptional and epigenetic changes that endow cells with different functional capacities and therefore therapeutic potential. The molecular circuitry underlying these developmental trajectories and the extent of heterogeneity within Teff, Tmem and Tex populations remain poorly understood. Here, we used the lymphocytic choriomeningitis virus model of acute-resolving and chronic infection to address these gaps by applying longitudinal single-cell RNA-sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) analyses. These analyses uncovered new subsets, including a subpopulation of Tex cells expressing natural killer cell-associated genes that is dependent on the transcription factor Zeb2, as well as multiple distinct TCF-1+ stem/progenitor-like subsets in acute and chronic infection. These data also revealed insights into the reshaping of Tex subsets following programmed death 1 (PD-1) pathway blockade and identified a key role for the cell stress regulator, Btg1, in establishing the Tex population. Finally, these results highlighted how the same biological circuits such as cytotoxicity or stem/progenitor pathways can be used by CD8+ T cell subsets with highly divergent underlying chromatin landscapes generated during different infections.


Assuntos
Linfócitos T CD8-Positivos , Coriomeningite Linfocítica , Humanos , Linfócitos T CD8-Positivos/metabolismo , Transcriptoma , Vírus da Coriomeningite Linfocítica , Epigênese Genética , Cromatina/genética , Cromatina/metabolismo , Coriomeningite Linfocítica/metabolismo
2.
Nat Immunol ; 23(8): 1183-1192, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35902637

RESUMO

Anti-programmed death-1 (anti-PD-1) immunotherapy reinvigorates CD8 T cell responses in patients with cancer but PD-1 is also expressed by other immune cells, including follicular helper CD4 T cells (Tfh) which are involved in germinal centre responses. Little is known, however, about the effects of anti-PD-1 immunotherapy on noncancer immune responses in humans. To investigate this question, we examined the impact of anti-PD-1 immunotherapy on the Tfh-B cell axis responding to unrelated viral antigens. Following influenza vaccination, a subset of adults receiving anti-PD-1 had more robust circulating Tfh responses than adults not receiving immunotherapy. PD-1 pathway blockade resulted in transcriptional signatures of increased cellular proliferation in circulating Tfh and responding B cells compared with controls. These latter observations suggest an underlying change in the Tfh-B cell and germinal centre axis in a subset of immunotherapy patients. Together, these results demonstrate dynamic effects of anti-PD-1 therapy on influenza vaccine responses and highlight analytical vaccination as an approach that may reveal underlying immune predisposition to adverse events.


Assuntos
Vacinas contra Influenza , Adulto , Humanos , Imunidade Humoral , Estações do Ano , Linfócitos T Auxiliares-Indutores , Vacinação
3.
Immunity ; 51(5): 840-855.e5, 2019 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-31606264

RESUMO

TCF-1 is a key transcription factor in progenitor exhausted CD8 T cells (Tex). Moreover, this Tex cell subset mediates responses to PD-1 checkpoint pathway blockade. However, the role of the transcription factor TCF-1 in early fate decisions and initial generation of Tex cells is unclear. Single-cell RNA sequencing (scRNA-seq) and lineage tracing identified a TCF-1+Ly108+PD-1+ CD8 T cell population that seeds development of mature Tex cells early during chronic infection. TCF-1 mediated the bifurcation between divergent fates, repressing development of terminal KLRG1Hi effectors while fostering KLRG1Lo Tex precursor cells, and PD-1 stabilized this TCF-1+ Tex precursor cell pool. TCF-1 mediated a T-bet-to-Eomes transcription factor transition in Tex precursors by promoting Eomes expression and drove c-Myb expression that controlled Bcl-2 and survival. These data define a role for TCF-1 in early-fate-bifurcation-driving Tex precursor cells and also identify PD-1 as a protector of this early TCF-1 subset.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Redes Reguladoras de Genes , Fator 1 de Transcrição de Linfócitos T/metabolismo , Transcrição Gênica , Animais , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Doença Crônica , Perfilação da Expressão Gênica , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Camundongos , Receptor de Morte Celular Programada 1/metabolismo , Fator 1 de Transcrição de Linfócitos T/genética , Viroses/genética , Viroses/imunologia , Viroses/virologia
4.
Nature ; 571(7764): 211-218, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31207603

RESUMO

Exhausted CD8+ T (Tex) cells in chronic infections and cancer have limited effector function, high co-expression of inhibitory receptors and extensive transcriptional changes compared with effector (Teff) or memory (Tmem) CD8+ T cells. Tex cells are important clinical targets of checkpoint blockade and other immunotherapies. Epigenetically, Tex cells are a distinct immune subset, with a unique chromatin landscape compared with Teff and Tmem cells. However, the mechanisms that govern the transcriptional and epigenetic development of Tex cells remain unknown. Here we identify the HMG-box transcription factor TOX as a central regulator of Tex cells in mice. TOX is largely dispensable for the formation of Teff and Tmem cells, but it is critical for exhaustion: in the absence of TOX, Tex cells do not form. TOX is induced by calcineurin and NFAT2, and operates in a feed-forward loop in which it becomes calcineurin-independent and sustained in Tex cells. Robust expression of TOX therefore results in commitment to Tex cells by translating persistent stimulation into a distinct Tex cell transcriptional and epigenetic developmental program.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Epistasia Genética , Proteínas de Homeodomínio/metabolismo , Transcrição Gênica , Animais , Calcineurina/metabolismo , Sinalização do Cálcio , Retroalimentação Fisiológica , Feminino , Regulação da Expressão Gênica/imunologia , Genótipo , Memória Imunológica , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/metabolismo , Evasão Tumoral
5.
Nature ; 545(7652): 60-65, 2017 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-28397821

RESUMO

Despite the success of monotherapies based on blockade of programmed cell death 1 (PD-1) in human melanoma, most patients do not experience durable clinical benefit. Pre-existing T-cell infiltration and/or the presence of PD-L1 in tumours may be used as indicators of clinical response; however, blood-based profiling to understand the mechanisms of PD-1 blockade has not been widely explored. Here we use immune profiling of peripheral blood from patients with stage IV melanoma before and after treatment with the PD-1-targeting antibody pembrolizumab and identify pharmacodynamic changes in circulating exhausted-phenotype CD8 T cells (Tex cells). Most of the patients demonstrated an immunological response to pembrolizumab. Clinical failure in many patients was not solely due to an inability to induce immune reinvigoration, but rather resulted from an imbalance between T-cell reinvigoration and tumour burden. The magnitude of reinvigoration of circulating Tex cells determined in relation to pretreatment tumour burden correlated with clinical response. By focused profiling of a mechanistically relevant circulating T-cell subpopulation calibrated to pretreatment disease burden, we identify a clinically accessible potential on-treatment predictor of response to PD-1 blockade.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Melanoma/tratamento farmacológico , Melanoma/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Carga Tumoral/imunologia , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/imunologia , Anticorpos Monoclonais Humanizados/farmacocinética , Anticorpos Monoclonais Humanizados/uso terapêutico , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/metabolismo , Feminino , Humanos , Antígeno Ki-67/imunologia , Antígeno Ki-67/metabolismo , Masculino , Melanoma/irrigação sanguínea , Melanoma/patologia , Estadiamento de Neoplasias , Fenótipo , Resultado do Tratamento
6.
Nature ; 509(7498): 55-62, 2014 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-24590074

RESUMO

Antibodies capable of neutralizing HIV-1 often target variable regions 1 and 2 (V1V2) of the HIV-1 envelope, but the mechanism of their elicitation has been unclear. Here we define the developmental pathway by which such antibodies are generated and acquire the requisite molecular characteristics for neutralization. Twelve somatically related neutralizing antibodies (CAP256-VRC26.01-12) were isolated from donor CAP256 (from the Centre for the AIDS Programme of Research in South Africa (CAPRISA)); each antibody contained the protruding tyrosine-sulphated, anionic antigen-binding loop (complementarity-determining region (CDR) H3) characteristic of this category of antibodies. Their unmutated ancestor emerged between weeks 30-38 post-infection with a 35-residue CDR H3, and neutralized the virus that superinfected this individual 15 weeks after initial infection. Improved neutralization breadth and potency occurred by week 59 with modest affinity maturation, and was preceded by extensive diversification of the virus population. HIV-1 V1V2-directed neutralizing antibodies can thus develop relatively rapidly through initial selection of B cells with a long CDR H3, and limited subsequent somatic hypermutation. These data provide important insights relevant to HIV-1 vaccine development.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp160 do Envelope de HIV/química , Proteína gp160 do Envelope de HIV/imunologia , Vacinas contra a AIDS/química , Vacinas contra a AIDS/imunologia , Sequência de Aminoácidos , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/isolamento & purificação , Afinidade de Anticorpos/genética , Afinidade de Anticorpos/imunologia , Linfócitos B/citologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Sítios de Ligação/imunologia , Antígenos CD4/imunologia , Antígenos CD4/metabolismo , Linhagem da Célula , Regiões Determinantes de Complementaridade/química , Regiões Determinantes de Complementaridade/genética , Regiões Determinantes de Complementaridade/imunologia , Mapeamento de Epitopos , Epitopos de Linfócito B/química , Epitopos de Linfócito B/imunologia , Evolução Molecular , Anticorpos Anti-HIV/química , Anticorpos Anti-HIV/genética , Anticorpos Anti-HIV/isolamento & purificação , Infecções por HIV/imunologia , HIV-1/química , HIV-1/imunologia , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Testes de Neutralização , Estrutura Terciária de Proteína , Hipermutação Somática de Imunoglobulina/genética
7.
J Immunol ; 197(4): 1017-22, 2016 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-27430722

RESUMO

The role of Ab and B cells in preventing infection is established. In contrast, the role of B cell responses in containing chronic infections remains poorly understood. IgG2a (IgG1 in humans) can prevent acute infections, and T-bet promotes IgG2a isotype switching. However, whether IgG2a and B cell-expressed T-bet influence the host-pathogen balance during persisting infections is unclear. We demonstrate that B cell-specific loss of T-bet prevents control of persisting viral infection. T-bet in B cells controlled IgG2a production, as well as mucosal localization, proliferation, glycosylation, and a broad transcriptional program. T-bet controlled a broad antiviral program in addition to IgG2a because T-bet in B cells was important, even in the presence of virus-specific IgG2a. Our data support a model in which T-bet is a universal controller of antiviral immunity across multiple immune lineages.


Assuntos
Subpopulações de Linfócitos B/imunologia , Linfócitos B/imunologia , Coriomeningite Linfocítica/imunologia , Proteínas com Domínio T/imunologia , Animais , Separação Celular , Doença Crônica , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Imunoglobulina G/imunologia , Vírus da Coriomeningite Linfocítica , Camundongos , Camundongos Transgênicos , Análise de Sequência com Séries de Oligonucleotídeos
8.
J Virol ; 90(1): 76-91, 2016 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-26468542

RESUMO

UNLABELLED: The epitopes defined by HIV-1 broadly neutralizing antibodies (bNAbs) are valuable templates for vaccine design, and studies of the immunological development of these antibodies are providing insights for vaccination strategies. In addition, the most potent and broadly reactive of these bNAbs have potential for clinical use. We previously described a family of 12 V1V2-directed neutralizing antibodies, CAP256-VRC26, isolated from an HIV-1 clade C-infected donor at years 1, 2, and 4 of infection (N. A. Doria-Rose et al., Nature 509:55-62, 2014, http://dx.doi.org/10.1038/nature13036). Here, we report on the isolation and characterization of new members of the family mostly obtained at time points of peak serum neutralization breadth and potency. Thirteen antibodies were isolated from B cell culture, and eight were isolated using trimeric envelope probes for differential single B cell sorting. One of the new antibodies displayed a 10-fold greater neutralization potency than previously published lineage members. This antibody, CAP256-VRC26.25, neutralized 57% of diverse clade viral isolates and 70% of clade C isolates with remarkable potency. Among the viruses neutralized, the median 50% inhibitory concentration was 0.001 µg/ml. All 33 lineage members targeted a quaternary epitope focused on V2. While all known bNAbs targeting the V1V2 region interact with the N160 glycan, the CAP256-VRC26 antibodies showed an inverse correlation of neutralization potency with dependence on this glycan. Overall, our results highlight the ongoing evolution within a single antibody lineage and describe more potent and broadly neutralizing members with potential clinical utility, particularly in areas where clade C is prevalent. IMPORTANCE: Studies of HIV-1 broadly neutralizing antibodies (bNAbs) provide valuable information for vaccine design, and the most potent and broadly reactive of these bNAbs have potential for clinical use. We previously described a family of V1V2-directed neutralizing antibodies from an HIV-1 clade C-infected donor. Here, we report on the isolation and characterization of new members of the family mostly obtained at time points of peak serum neutralization breadth and potency. One of the new antibodies, CAP256-VRC26.25, displayed a 10-fold greater neutralization potency than previously described lineage members. It neutralized 57% of diverse clade viral isolates and 70% of clade C isolates with remarkable potency: the median 50% inhibitory concentration was 0.001 µg/ml. Our results highlight the ongoing evolution within a single antibody lineage and describe more potent and broadly neutralizing members with potential clinical utility, particularly in areas where clade C is prevalent.


Assuntos
Anticorpos Neutralizantes/imunologia , Reações Cruzadas , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/imunologia , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/isolamento & purificação , Mapeamento de Epitopos , Feminino , Anticorpos Anti-HIV/genética , Anticorpos Anti-HIV/isolamento & purificação , Proteína gp120 do Envelope de HIV/imunologia , Humanos , Concentração Inibidora 50 , Dados de Sequência Molecular , Análise de Sequência de DNA
9.
J Virol ; 86(15): 8319-23, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22623764

RESUMO

Antibody PG9 is a prototypical member of a class of V1/V2-directed antibodies that effectively neutralizes diverse strains of HIV-1. We analyzed strain-specific resistance to PG9 using sequence and structural information. For multiply resistant strains, mutations in a short segment of V1/V2 resulted in gain of sensitivity to PG9 and related V1/V2 neutralizing antibodies, suggesting both a common mechanism of HIV-1 resistance to and a common mode of recognition by this class of antibodies.


Assuntos
Anticorpos Neutralizantes/química , Anticorpos Anti-HIV/química , Proteína gp120 do Envelope de HIV/química , HIV-1/química , Sequência de Aminoácidos , Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , HIV-1/imunologia , Humanos , Relação Estrutura-Atividade
10.
Sci Immunol ; 8(86): eade3369, 2023 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-37595022

RESUMO

Identifying molecular mechanisms of exhausted CD8 T cells (Tex) is a key goal of improving immunotherapy of cancer and other diseases. However, high-throughput interrogation of in vivo Tex can be costly and inefficient. In vitro models of Tex are easily customizable and quickly generate high cellular yield, enabling CRISPR screening and other high-throughput assays. We established an in vitro model of chronic stimulation and benchmarked key phenotypic, functional, transcriptional, and epigenetic features against bona fide in vivo Tex. We leveraged this model of in vitro chronic stimulation in combination with CRISPR screening to identify transcriptional regulators of T cell exhaustion. This approach identified several transcription factors, including BHLHE40. In vitro and in vivo validation defined a role for BHLHE40 in regulating a key differentiation checkpoint between progenitor and intermediate Tex subsets. By developing and benchmarking an in vitro model of Tex, then applying high-throughput CRISPR screening, we demonstrate the utility of mechanistically annotated in vitro models of Tex.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Exaustão das Células T , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Linfócitos T CD8-Positivos , Diferenciação Celular , Epigenômica
11.
bioRxiv ; 2023 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-37131713

RESUMO

Identifying novel molecular mechanisms of exhausted CD8 T cells (T ex ) is a key goal of improving immunotherapy of cancer and other diseases. However, high-throughput interrogation of in vivo T ex can be costly and inefficient. In vitro models of T ex are easily customizable and quickly generate high cellular yield, offering an opportunity to perform CRISPR screening and other high-throughput assays. We established an in vitro model of chronic stimulation and benchmarked key phenotypic, functional, transcriptional, and epigenetic features against bona fide in vivo T ex . We leveraged this model of in vitro chronic stimulation in combination with pooled CRISPR screening to uncover transcriptional regulators of T cell exhaustion. This approach identified several transcription factors, including BHLHE40. In vitro and in vivo validation defined a role for BHLHE40 in regulating a key differentiation checkpoint between progenitor and intermediate subsets of T ex . By developing and benchmarking an in vitro model of T ex , we demonstrate the utility of mechanistically annotated in vitro models of T ex , in combination with high-throughput approaches, as a discovery pipeline to uncover novel T ex biology.

12.
Commun Biol ; 5(1): 1399, 2022 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-36543997

RESUMO

In response to infection or immunization, antibodies are produced that provide protection against re-exposure with the same pathogen. These antibodies can persist at high titers for decades and are maintained by bone marrow-resident long-lived plasma cells (LLPC). However, the durability of antibody responses to immunization varies amongst vaccines. It is unknown what factors contribute to the differential longevity of serum antibody responses and whether heterogeneity in LLPC contributes to this phenomenon. While LLPC differentiation has been studied extensively in mice, little is known about this population in humans or non-human primates (NHP). Here, we use multi-omic single-cell profiling to identify and characterize the LLPC compartment in NHP. We identify LLPC biomarkers including the marker CD102 and show that CD102 in combination with CD31 identifies LLPC in NHP bone marrow. Additionally, we find that CD102 is expressed by LLPC in mouse and humans. These results further our understanding of the LLPC compartment in NHP, identify biomarkers of LLPC, and provide tissue-specific single cell references for future studies.


Assuntos
Multiômica , Plasmócitos , Camundongos , Animais , Formação de Anticorpos , Anticorpos , Primatas , Biomarcadores
13.
Cell Rep ; 35(6): 109120, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33979613

RESUMO

The transcription factors T-bet and Eomesodermin (Eomes) regulate CD8 T cell exhaustion through undefined mechanisms. Here, we show that the subcellular localization of T-bet and Eomes dictate their regulatory activity in exhausted T cells (TEXs). TEXs had a higher ratio of nuclear Eomes:T-bet than memory T cells (TMEMs) during chronic lymphocytic choriomeningitis virus (LCMV) infection in preclinical cancer models and in human tumors. Biochemically, T-bet and Eomes compete for the same DNA sequences, including the Pdcd1 T-box. High nuclear T-bet strongly represses Pdcd1 transcription in TMEM, whereas low nuclear T-bet in TEX leads to a dominant effect of Eomes that acts as a weaker repressor of Pdcd1. Blocking PD-1 signaling in TEXs increases nuclear T-bet, restoring stronger repression of Pdcd1, and driving T-bet-associated gene expression programs of chemotaxis, homing, and activation. These data identify a mechanism whereby the T-bet-Eomes axis regulates exhaustion through their nuclear localization, providing insights into how these transcription factors regulate TEX biology.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Proteínas com Domínio T/metabolismo , Animais , Diferenciação Celular , Humanos , Camundongos , Transdução de Sinais
14.
Science ; 351(6279): 1339-42, 2016 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-26917593

RESUMO

Ebola virus disease in humans is highly lethal, with case fatality rates ranging from 25 to 90%. There is no licensed treatment or vaccine against the virus, underscoring the need for efficacious countermeasures. We ascertained that a human survivor of the 1995 Kikwit Ebola virus disease outbreak maintained circulating antibodies against the Ebola virus surface glycoprotein for more than a decade after infection. From this survivor we isolated monoclonal antibodies (mAbs) that neutralize recent and previous outbreak variants of Ebola virus and mediate antibody-dependent cell-mediated cytotoxicity in vitro. Strikingly, monotherapy with mAb114 protected macaques when given as late as 5 days after challenge. Treatment with a single human mAb suggests that a simplified therapeutic strategy for human Ebola infection may be possible.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antivirais/administração & dosagem , Ebolavirus/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Adulto , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/isolamento & purificação , Ensaios Clínicos como Assunto , Surtos de Doenças , Feminino , Doença pelo Vírus Ebola/epidemiologia , Humanos , Macaca , Masculino , Dados de Sequência Molecular , Sobreviventes
15.
Science ; 340(6133): 751-6, 2013 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-23661761

RESUMO

Serum characterization and antibody isolation are transforming our understanding of the humoral immune response to viral infection. Here, we show that epitope specificities of HIV-1-neutralizing antibodies in serum can be elucidated from the serum pattern of neutralization against a diverse panel of HIV-1 isolates. We determined "neutralization fingerprints" for 30 neutralizing antibodies on a panel of 34 diverse HIV-1 strains and showed that similarity in neutralization fingerprint correlated with similarity in epitope. We used these fingerprints to delineate specificities of polyclonal sera from 24 HIV-1-infected donors and a chimeric siman-human immunodeficiency virus-infected macaque. Delineated specificities matched published specificities and were further confirmed by antibody isolation for two sera. Patterns of virus-isolate neutralization can thus afford a detailed epitope-specific understanding of neutralizing-antibody responses to viral infection.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Animais , Anticorpos Neutralizantes/sangue , Mapeamento de Epitopos , Anticorpos Anti-HIV/sangue , Infecções por HIV/sangue , HIV-1/isolamento & purificação , Humanos , Epitopos Imunodominantes/química , Epitopos Imunodominantes/imunologia , Macaca , Testes de Neutralização , Conformação Proteica , Soro/imunologia
16.
Nat Struct Mol Biol ; 20(7): 804-13, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23708607

RESUMO

HIV-1 uses a diverse N-linked-glycan shield to evade recognition by antibody. Select human antibodies, such as the clonally related PG9 and PG16, recognize glycopeptide epitopes in the HIV-1 V1-V2 region and penetrate this shield, but their ability to accommodate diverse glycans is unclear. Here we report the structure of antibody PG16 bound to a scaffolded V1-V2, showing an epitope comprising both high mannose-type and complex-type N-linked glycans. We combined structure, NMR and mutagenesis analyses to characterize glycan recognition by PG9 and PG16. Three PG16-specific residues, arginine, serine and histidine (RSH), were critical for binding sialic acid on complex-type glycans, and introduction of these residues into PG9 produced a chimeric antibody with enhanced HIV-1 neutralization. Although HIV-1-glycan diversity facilitates evasion, antibody somatic diversity can overcome this and can provide clues to guide the design of modified antibodies with enhanced neutralization.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , Fragmentos de Peptídeos/imunologia , Polissacarídeos/imunologia , Motivos de Aminoácidos , Sequência de Aminoácidos , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/metabolismo , Especificidade de Anticorpos , Reações Antígeno-Anticorpo , Sítios de Ligação de Anticorpos , Configuração de Carboidratos , Sequência de Carboidratos , Cristalografia por Raios X , Epitopos/química , Epitopos/imunologia , Epitopos/metabolismo , Glicosilação/efeitos dos fármacos , Células HEK293 , Anticorpos Anti-HIV/química , Anticorpos Anti-HIV/metabolismo , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Polissacarídeos/química , Polissacarídeos/metabolismo , Conformação Proteica , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Relação Estrutura-Atividade , Swainsonina/farmacologia
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