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1.
Immunity ; 57(3): 559-573.e6, 2024 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-38479361

RESUMO

Epstein-Barr virus (EBV) causes infectious mononucleosis and is associated with B cell lymphomas. EBV glycoprotein 42 (gp42) binds HLA class II and activates membrane fusion with B cells. We isolated gp42-specific monoclonal antibodies (mAbs), A10 and 4C12, which use distinct mechanisms to neutralize virus infection. mAb A10 was more potent than the only known neutralizing gp42 mAb, F-2-1, in neutralizing EBV infection and blocking binding to HLA class II. mAb 4C12 was similar to mAb A10 in inhibiting glycoprotein-mediated B cell fusion but did not block receptor binding, and it was less effective in neutralizing infection. Crystallographic structures of gH/gL/gp42/A10 and gp42/4C12 complexes revealed two distinct sites of vulnerability on gp42 for receptor binding and B cell fusion. Passive transfer of mAb A10 into humanized mice conferred nearly 100% protection from viremia and EBV lymphomas after EBV challenge. These findings identify vulnerable sites on EBV that may facilitate therapeutics and vaccines.


Assuntos
Benzenoacetamidas , Infecções por Vírus Epstein-Barr , Herpesvirus Humano 4 , Piperidonas , Animais , Camundongos , Proteínas Virais/metabolismo , Glicoproteínas/metabolismo , Anticorpos Antivirais
2.
Nat Immunol ; 25(3): 462-470, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38278966

RESUMO

The persistence of CD4+ T cells carrying latent human immunodeficiency virus-1 (HIV-1) proviruses is the main barrier to a cure. New therapeutics to enhance HIV-1-specific immune responses and clear infected cells will probably be necessary to achieve reduction of the latent reservoir. In the present study, we report two single-chain diabodies (scDbs) that target the HIV-1 envelope protein (Env) and the human type III Fcγ receptor (CD16). We show that the scDbs promoted robust and HIV-1-specific natural killer (NK) cell activation and NK cell-mediated lysis of infected cells. Cocultures of CD4+ T cells from people with HIV-1 on antiretroviral therapy (ART) with autologous NK cells and the scDbs resulted in marked elimination of reservoir cells that was dependent on latency reversal. Treatment of human interleukin-15 transgenic NSG mice with one of the scDbs after ART initiation enhanced NK cell activity and reduced reservoir size. Thus, HIV-1-specific scDbs merit further evaluation as potential therapeutics for clearance of the latent reservoir.


Assuntos
Anticorpos Biespecíficos , HIV-1 , Animais , Camundongos , Humanos , Células Matadoras Naturais , Citotoxicidade Imunológica , Morte Celular , Camundongos Transgênicos
3.
J Exp Med ; 220(7)2023 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-37058141

RESUMO

Distinct CD4+ T cell epitopes have been associated with spontaneous control of HIV-1 replication, but analysis of antigen-dependent factors that influence epitope selection is lacking. To examine these factors, we used a cell-free antigen processing system that incorporates soluble HLA-DR (DR1), HLA-DM (DM), cathepsins, and full-length protein antigens for epitope identification by LC-MS/MS. HIV-1 Gag, Pol, Env, Vif, Tat, Rev, and Nef were examined using this system. We identified 35 novel epitopes, including glycopeptides. Epitopes from smaller HIV-1 proteins mapped to regions of low protein stability and higher solvent accessibility. HIV-1 antigens associated with limited CD4+ T cell responses were processed efficiently, while some protective epitopes were inefficiently processed. 55% of epitopes obtained from cell-free processing induced memory CD4+ T cell responses in HIV-1+ donors, including eight of 19 novel epitopes tested. Thus, an in vitro processing system utilizing the components of Class II processing reveals factors influencing epitope selection of HIV-1 and represents an approach to understanding epitope selection from non-HIV-1 antigens.


Assuntos
Infecções por HIV , Vacinas , Humanos , Apresentação de Antígeno , Cromatografia Líquida , Espectrometria de Massas em Tandem , Epitopos de Linfócito T , Antígenos Virais
4.
Immunity ; 55(11): 2135-2148.e6, 2022 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-36306784

RESUMO

Epstein-Barr virus (EBV) is nearly ubiquitous in adults. EBV causes infectious mononucleosis and is associated with B cell lymphomas, epithelial cell malignancies, and multiple sclerosis. The EBV gH/gL glycoprotein complex facilitates fusion of virus membrane with host cells and is a target of neutralizing antibodies. Here, we examined the sites of vulnerability for virus neutralization and fusion inhibition within EBV gH/gL. We developed a panel of human monoclonal antibodies (mAbs) that targeted five distinct antigenic sites on EBV gH/gL and prevented infection of epithelial and B cells. Structural analyses using X-ray crystallography and electron microscopy revealed multiple sites of vulnerability and defined the antigenic landscape of EBV gH/gL. One mAb provided near-complete protection against viremia and lymphoma in a humanized mouse EBV challenge model. Our findings provide structural and antigenic knowledge of the viral fusion machinery, yield a potential therapeutic antibody to prevent EBV disease, and emphasize gH/gL as a target for herpesvirus vaccines and therapeutics.


Assuntos
Infecções por Vírus Epstein-Barr , Herpesvirus Humano 4 , Cricetinae , Camundongos , Animais , Humanos , Proteínas do Envelope Viral , Cricetulus , Glicoproteínas de Membrana , Células CHO
5.
Proc Natl Acad Sci U S A ; 119(15): e2123406119, 2022 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-35394875

RESUMO

HIV-1 infection is incurable due to the persistence of the virus in a latent reservoir of resting memory CD4+ T cells. "Shock-and-kill" approaches that seek to induce HIV-1 gene expression, protein production, and subsequent targeting by the host immune system have been unsuccessful due to a lack of effective latency-reversing agents (LRAs) and kill strategies. In an effort to develop reagents that could be used to promote killing of infected cells, we constructed T cell receptor (TCR)-mimic antibodies to HIV-1 peptide-major histocompatibility complexes (pMHC). Using phage display, we panned for phages expressing antibody-like variable sequences that bound HIV-1 pMHC generated using the common HLA-A*02:01 allele. We targeted three epitopes in Gag and reverse transcriptase identified and quantified via Poisson detection mass spectrometry from cells infected in vitro with a pseudotyped HIV-1 reporter virus (NL4.3 dEnv). Sequences isolated from phages that bound these pMHC were cloned into a single-chain diabody backbone (scDb) sequence, such that one fragment is specific for an HIV-1 pMHC and the other fragment binds to CD3ε, an essential signal transduction subunit of the TCR. Thus, these antibodies utilize the sensitivity of T cell signaling as readouts for antigen processing and as agents to promote killing of infected cells. Notably, these scDbs are exquisitely sensitive and specific for the peptide portion of the pMHC. Most importantly, one scDb caused killing of infected cells presenting a naturally processed target pMHC. This work lays the foundation for a novel therapeutic killing strategy toward elimination of the HIV-1 reservoir.


Assuntos
Anticorpos Biespecíficos , Infecções por HIV , Soropositividade para HIV , HIV-1 , Linfócitos T CD4-Positivos , Humanos , Mimetismo Molecular , Receptores de Antígenos de Linfócitos T , Latência Viral
6.
PLoS Pathog ; 18(1): e1010179, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34990486

RESUMO

Antibodies targeting the hepatitis C virus (HCV) envelope glycoprotein E2 are associated with delayed disease progression, and these antibodies can also facilitate spontaneous clearance of infection in some individuals. However, many infected people demonstrate low titer and delayed anti-E2 antibody responses. Since a goal of HCV vaccine development is induction of high titers of anti-E2 antibodies, it is important to define the mechanisms underlying these suboptimal antibody responses. By staining lymphocytes with a cocktail of soluble E2 (sE2) glycoproteins, we detected HCV E2-specific (sE2+) B cells directly ex vivo at multiple acute infection timepoints in 29 HCV-infected subjects with a wide range of anti-E2 IgG titers, including 17 persistently infected subjects and 12 subjects with spontaneous clearance of infection. We performed multi-dimensional flow cytometric analysis of sE2+ and E2-nonspecific (sE2-) class-switched B cells (csBC). In sE2+ csBC from both persistence and clearance subjects, frequencies of resting memory B cells (rMBC) were reduced, frequencies of activated MBC (actMBC) and tissue-like MBC (tlMBC) were increased, and expression of FCRL5, an IgG receptor, was significantly upregulated. Across all subjects, plasma anti-E2 IgG levels were positively correlated with frequencies of sE2+ rMBC and sE2+ actMBC, while anti-E2 IgG levels were negatively correlated with levels of FCRL5 expression on sE2+ rMBC and PD-1 expression on sE2+ actMBC. Upregulation of FCRL5 on sE2+ rMBC and upregulation of PD-1 on sE2+ actMBC may limit anti-E2 antibody production in vivo. Strategies that limit upregulation of these molecules could potentially generate higher titers of protective antibodies against HCV or other pathogens.


Assuntos
Linfócitos B/imunologia , Anticorpos Anti-Hepatite C/imunologia , Hepatite C/imunologia , Receptor de Morte Celular Programada 1/imunologia , Receptores Fc/imunologia , Hepacivirus/imunologia , Humanos , Proteínas do Envelope Viral/imunologia
7.
Nat Rev Immunol ; 22(8): 499-512, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-34824401

RESUMO

Combination antiretroviral therapy (ART) can block multiple stages of the HIV-1 life cycle to prevent progression to AIDS in people living with HIV-1. However, owing to the persistence of a reservoir of latently infected CD4+ T cells, life-long ART is necessary to prevent viral rebound. One strategy currently under consideration for curing HIV-1 infection is known as 'shock and kill'. This strategy uses latency-reversing agents to induce expression of HIV-1 genes, allowing for infected cells to be cleared by cytolytic immune cells. The role of innate immunity in HIV-1 pathogenesis is best understood in the context of acute infection. Here, we suggest that innate immunity can also be used to improve the efficacy of HIV-1 cure strategies, with a particular focus on dendritic cells (DCs) and natural killer cells. We discuss novel latency-reversing agents targeting DCs as well as DC-based strategies to enhance the clearance of infected cells by CD8+ T cells and strategies to improve the killing activity of natural killer cells.


Assuntos
Infecções por HIV , HIV-1 , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Infecções por HIV/terapia , Humanos , Imunidade Inata , Latência Viral
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