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1.
Cell ; 170(4): 637-648.e10, 2017 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-28757252

RESUMO

Non-neutralizing antibodies (nnAbs) to HIV-1 show little measurable activity in prevention or therapy in animal models yet were the only correlate of protection in the RV144 vaccine trial. To investigate the role of nnAbs on HIV-1 infection in vivo, we devised a replication-competent HIV-1 reporter virus that expresses a heterologous HA-tag on the surface of infected cells and virions. Anti-HA antibodies bind to, but do not neutralize, the reporter virus in vitro. However, anti-HA protects against infection in humanized mice and strongly selects for nnAb-resistant viruses in an entirely Fc-dependent manner. Similar results were also obtained with tier 2 HIV-1 viruses using a human anti-gp41 nnAb, 246D. While nnAbs are demonstrably less effective than broadly neutralizing antibodies (bNAbs) against HIV-1 in vitro and in vivo, the data show that nnAbs can protect against and alter the course of HIV-1 infection in vivo. PAPERCLIP.


Assuntos
Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/fisiologia , Vacinas contra a AIDS/imunologia , Animais , Antígenos CD4/química , Antígenos CD4/metabolismo , Modelos Animais de Doenças , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/genética , Humanos , Camundongos , Mutação , Receptores Fc/imunologia , Linfócitos T/virologia
2.
Mol Ther ; 32(10): 3712-3728, 2024 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-39086132

RESUMO

Targeting multiple viral proteins is pivotal for sustained suppression of highly mutable viruses. In recent years, broadly neutralizing antibodies that target the influenza virus hemagglutinin and neuraminidase glycoproteins have been developed, and antibody monotherapy has been tested in preclinical and clinical studies to treat or prevent influenza virus infection. However, the impact of dual neutralization of the hemagglutinin and neuraminidase on the course of infection, as well as its therapeutic potential, has not been thoroughly tested. For this purpose, we generated a bispecific antibody that neutralizes both the hemagglutinin and the neuraminidase of influenza viruses. We demonstrated that this bispecific antibody has a dual-antiviral activity as it blocks infection and prevents the release of progeny viruses from the infected cells. We show that dual neutralization of the hemagglutinin and the neuraminidase by a bispecific antibody is advantageous over monoclonal antibody combination as it resulted an improved neutralization capacity and augmented the antibody effector functions. Notably, the bispecific antibody showed enhanced antiviral activity in influenza virus-infected mice, reduced mice mortality, and limited the virus mutation profile upon antibody administration. Thus, dual neutralization of the hemagglutinin and neuraminidase could be effective in controlling influenza virus infection.


Assuntos
Anticorpos Biespecíficos , Anticorpos Neutralizantes , Anticorpos Antivirais , Glicoproteínas de Hemaglutininação de Vírus da Influenza , Neuraminidase , Anticorpos Biespecíficos/farmacologia , Anticorpos Biespecíficos/imunologia , Animais , Neuraminidase/antagonistas & inibidores , Neuraminidase/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Camundongos , Humanos , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Anticorpos Antivirais/imunologia , Antivirais/farmacologia , Antivirais/uso terapêutico , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/tratamento farmacológico , Infecções por Orthomyxoviridae/virologia , Testes de Neutralização , Cães , Modelos Animais de Doenças , Células Madin Darby de Rim Canino , Influenza Humana/imunologia , Influenza Humana/virologia , Influenza Humana/tratamento farmacológico , Feminino
3.
PLoS Pathog ; 18(1): e1010242, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-35020754

RESUMO

In-depth analysis of SARS-CoV-2 quasispecies is pivotal for a thorough understating of its evolution during infection. The recent deployment of COVID-19 vaccines, which elicit protective anti-spike neutralizing antibodies, has stressed the importance of uncovering and characterizing SARS-CoV-2 variants with mutated spike proteins. Sequencing databases have allowed to follow the spread of SARS-CoV-2 variants that are circulating in the human population, and several experimental platforms were developed to study these variants. However, less is known about the SARS-CoV-2 variants that are developed in the respiratory system of the infected individual. To gain further insight on SARS-CoV-2 mutagenesis during natural infection, we preformed single-genome sequencing of SARS-CoV-2 isolated from nose-throat swabs of infected individuals. Interestingly, intra-host SARS-CoV-2 variants with mutated S genes or N genes were detected in all individuals who were analyzed. These intra-host variants were present in low frequencies in the swab samples and were rarely documented in current sequencing databases. Further examination of representative spike variants identified by our analysis showed that these variants have impaired infectivity capacity and that the mutated variants showed varied sensitivity to neutralization by convalescent plasma and to plasma from vaccinated individuals. Notably, analysis of the plasma neutralization activity against these variants showed that the L1197I mutation at the S2 subunit of the spike can affect the plasma neutralization activity. Together, these results suggest that SARS-CoV-2 intra-host variants should be further analyzed for a more thorough characterization of potential circulating variants.


Assuntos
Vacina BNT162/administração & dosagem , COVID-19 , Proteínas do Nucleocapsídeo de Coronavírus , Bases de Dados de Ácidos Nucleicos , Genoma Viral , Mutação , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Adulto , Idoso , COVID-19/genética , COVID-19/imunologia , COVID-19/prevenção & controle , Criança , Proteínas do Nucleocapsídeo de Coronavírus/genética , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Feminino , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Fosfoproteínas/genética , Fosfoproteínas/imunologia , SARS-CoV-2/genética , SARS-CoV-2/imunologia , Análise de Sequência de RNA , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia
4.
Immunity ; 42(2): 344-355, 2015 Feb 17.
Artigo em Inglês | MEDLINE | ID: mdl-25680274

RESUMO

Bacteria, such as Fusobacterium nucleatum, are present in the tumor microenvironment. However, the immunological consequences of intra-tumoral bacteria remain unclear. Here, we have shown that natural killer (NK) cell killing of various tumors is inhibited in the presence of various F. nucleatum strains. Our data support that this F. nucleatum-mediated inhibition is mediated by human, but not by mouse TIGIT, an inhibitory receptor present on all human NK cells and on various T cells. Using a library of F. nucleatum mutants, we found that the Fap2 protein of F. nucleatum directly interacted with TIGIT, leading to the inhibition of NK cell cytotoxicity. We have further demonstrated that tumor-infiltrating lymphocytes expressed TIGIT and that T cell activities were also inhibited by F. nucleatum via Fap2. Our results identify a bacterium-dependent, tumor-immune evasion mechanism in which tumors exploit the Fap2 protein of F. nucleatum to inhibit immune cell activity via TIGIT.


Assuntos
Adenocarcinoma/imunologia , Adenocarcinoma/microbiologia , Neoplasias do Colo/imunologia , Neoplasias do Colo/microbiologia , Fusobacterium nucleatum/imunologia , Receptores Imunológicos/imunologia , Evasão Tumoral/imunologia , Microambiente Tumoral/imunologia , Animais , Proteínas da Membrana Bacteriana Externa/imunologia , Linhagem Celular , Proliferação de Células , Humanos , Células Matadoras Naturais/imunologia , Linfócitos do Interstício Tumoral/imunologia , Camundongos , Ligação Proteica
5.
Proc Natl Acad Sci U S A ; 117(31): 18719-18728, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32690692

RESUMO

CD4-based decoy approaches against HIV-1 are attractive options for long-term viral control, but initial designs, including soluble CD4 (sCD4) and CD4-Ig, were ineffective. To evaluate a therapeutic that more accurately mimics HIV-1 target cells compared with monomeric sCD4 and dimeric CD4-Ig, we generated virus-like nanoparticles that present clusters of membrane-associated CD4 (CD4-VLPs) to permit high-avidity binding of trimeric HIV-1 envelope spikes. In neutralization assays, CD4-VLPs were >12,000-fold more potent than sCD4 and CD4-Ig and >100-fold more potent than the broadly neutralizing antibody (bNAb) 3BNC117, with >12,000-fold improvements against strains poorly neutralized by 3BNC117. CD4-VLPs also neutralized patient-derived viral isolates that were resistant to 3BNC117 and other bNAbs. Intraperitoneal injections of CD4-CCR5-VLP produced only subneutralizing plasma concentrations in HIV-1-infected humanized mice but elicited CD4-binding site mutations that reduced viral fitness. All mutant viruses showed reduced sensitivity to sCD4 and CD4-Ig but remained sensitive to neutralization by CD4-VLPs in vitro. In vitro evolution studies demonstrated that CD4-VLPs effectively controlled HIV-1 replication at neutralizing concentrations, and viral escape was not observed. Moreover, CD4-VLPs potently neutralized viral swarms that were completely resistant to CD4-Ig, suggesting that escape pathways that confer resistance against conventional CD4-based inhibitors are ineffective against CD4-VLPs. These findings suggest that therapeutics that mimic HIV-1 target cells could prevent viral escape by exposing a universal vulnerability of HIV-1: the requirement to bind CD4 on a target cell. We propose that therapeutic and delivery strategies that ensure durable bioavailability need to be developed to translate this concept into a clinically feasible functional cure therapy.


Assuntos
Antígenos CD4 , HIV-1 , Nanopartículas , Vírion , Fármacos Anti-HIV , Antígenos CD4/química , Antígenos CD4/metabolismo , Linhagem Celular , HIV-1/química , HIV-1/genética , HIV-1/metabolismo , Humanos , Mimetismo Molecular , Nanomedicina/métodos , Nanopartículas/química , Nanopartículas/metabolismo , Vírion/química , Vírion/metabolismo
6.
Clin Oral Investig ; 27(9): 5499-5508, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37490117

RESUMO

OBJECTIVE: To identify, quantify, and characterize leukocyte populations in PI and periodontitis using flow cytometry. METHODS: Fresh biopsies from human PI and periodontitis lesions were processed to a single-cell suspension. The immune cell types were identified using flow cytometry. RESULTS: Twenty-one biopsies were obtained and analyzed corresponding to fourteen PI and seven periodontitis samples. Participants' average age was 63.95 ± 14.77 years without a significant difference between PI and periodontitis patients, the female/male ratio was 8/12, and mean PD was 8.5 ± 2.17. High similarity was found between periodontitis and PI in the main immune cell types. Out of the leukocytes, the PMN proportion was 40% in PI and 33% in periodontitis. T-cells 22% in PI and 18% in periodontitis. Similar proportions of B-cells 10% and macrophages 6% were found in PI and periodontitis. Dendritic and NK cells were found in low proportions (~ 1%) in PI and periodontitis. T-cell sub-analysis showed that CD4-positive were more prevalent than CD8-positive in both diseases (CD4/CD8 ratio of 1.2). CONCLUSION: With the use of flow cytometry analysis, the leukocyte populations in human peri-implantitis and periodontitis were classified. In PI and periodontitis, we identified similar proportions of specific (CD4/CD8) and innate (dendritic and NK) immune cells. These results corroborate previous histological studies. CLINICAL RELEVANCE: Flow cytometry analysis can be used to identify and quantify immune cells in PI and periodontitis, including sub-classification of T cells (CD4/8) as well as detection of cells that require multiple markers for identification (such as dendritic cells).


Assuntos
Implantes Dentários , Peri-Implantite , Periodontite , Humanos , Masculino , Feminino , Pessoa de Meia-Idade , Idoso , Peri-Implantite/metabolismo , Citometria de Fluxo , Periodontite/metabolismo , Leucócitos
7.
Rheumatology (Oxford) ; 61(8): 3439-3447, 2022 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-34849628

RESUMO

OBJECTIVES: The mRNA-based COVID-19 vaccines are now employed globally and have shown high efficacy in preventing SARS-CoV-2 infection. However, less is known about the vaccine efficacy in immune-suppressed individuals. This study sought to explore whether humoral immunity to the COVID-19 vaccine BNT162b2 is altered in RA patients treated with Janus kinase inhibitors by analysing their antibodies titre, neutralization activity and B cell responses. METHODS: We collected plasma samples from 12 RA patients who were treated with Janus kinase inhibitors and received two doses of the BNT162b2 vaccine, as well as 26 healthy individuals who were vaccinated with the same vaccine. We analysed the quantity of the anti-spike IgG and IgA antibodies that were elicited following the BNT162b2 vaccination, the plasma neutralization capacity and the responsiveness of the B-lymphocytes. We used ELISA to quantify the antibody titres, and a plasma neutralization assay was used to determine the virus neutralization capacity. Alteration in expression of the genes that are associated with B cell activation and the germinal centre response were analysed by quantitative PCR. RESULTS: Reduced levels of anti-spike IgG antibodies and neutralization capacity were seen in the RA patients who were treated with JAK inhibitors in comparison with healthy individuals. Furthermore, B cell responsiveness to the SARS-CoV-2 spike protein was reduced in the RA patients. CONCLUSION: RA patients who are treated with JAK inhibitors show a suppressed humoral response following BNT162b2 vaccination, as revealed by the quantity and quality of the anti-spike antibodies.


Assuntos
Artrite Reumatoide , Vacina BNT162 , COVID-19 , Imunidade Humoral , Inibidores de Janus Quinases , Anticorpos Neutralizantes , Anticorpos Antivirais , Artrite Reumatoide/tratamento farmacológico , Vacina BNT162/imunologia , COVID-19/prevenção & controle , Humanos , Imunoglobulina G , Inibidores de Janus Quinases/uso terapêutico , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Vacinação
8.
Nature ; 535(7613): 556-60, 2016 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-27338952

RESUMO

Interruption of combination antiretroviral therapy in HIV-1-infected individuals leads to rapid viral rebound. Here we report the results of a phase IIa open label clinical trial evaluating 3BNC117,a broad and potent neutralizing antibody against the CD4 binding site of the HIV-1 Env protein, during analytical treatment interruption in 13 HIV-1-infected individuals. Participants with 3BNC117-sensitive virus outgrowth cultures were enrolled. Results show that two or four 30 mg kg(-1) 3BNC117 infusions,separated by 3 or 2 weeks, respectively, are generally well tolerated.Infusions are associated with a delay in viral rebound of 5-9 weeks after two infusions, and up to 19 weeks after four infusions, or an average of 6.7 and 9.9 weeks, respectively, compared with 2.6 weeks for historical controls (P < 0.00001). Rebound viruses arise predominantly from a single provirus. In most individuals,emerging viruses show increased resistance, indicating escape.However, 30% of participants remained suppressed until antibody concentrations waned below 20 µg ml(-1), and the viruses emerging in all but one of these individuals showed no apparent resistance to 3BCN117, suggesting failure to escape over a period of 9-19 weeks.We conclude that the administration of 3BNC117 exerts strong selective pressure on HIV-1 emerging from latent reservoirs during analytical treatment interruption in humans.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/crescimento & desenvolvimento , HIV-1/imunologia , Adolescente , Adulto , Idoso , Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/uso terapêutico , Anticorpos Monoclonais Humanizados , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/uso terapêutico , Sítios de Ligação/efeitos dos fármacos , Sítios de Ligação/imunologia , Anticorpos Amplamente Neutralizantes , Antígenos CD4/metabolismo , Reservatórios de Doenças/virologia , Esquema de Medicação , Feminino , Anticorpos Anti-HIV/administração & dosagem , Anticorpos Anti-HIV/uso terapêutico , Proteína gp160 do Envelope de HIV/antagonistas & inibidores , Proteína gp160 do Envelope de HIV/química , Proteína gp160 do Envelope de HIV/imunologia , Proteína gp160 do Envelope de HIV/metabolismo , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , Estudo Historicamente Controlado , Humanos , Masculino , Pessoa de Meia-Idade , Provírus/efeitos dos fármacos , Provírus/crescimento & desenvolvimento , Provírus/imunologia , Fatores de Tempo , Distribuição Tecidual , Carga Viral/efeitos dos fármacos , Carga Viral/imunologia , Adulto Jovem
9.
J Immunol ; 198(9): 3662-3670, 2017 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-28356383

RESUMO

NK cells are part of the innate immune system, and are able to identify and kill hazardous cells. The discrimination between normal and hazardous cells is possible due to an array of inhibitory and activating receptors. NKG2D is one of the prominent activating receptors expressed by all human NK cells. This receptor binds stress-induced ligands, including human MICA, MICB, and UL16-binding proteins 1-6. The interaction between NKG2D and its ligands facilitates the elimination of cells under cellular stress, such as tumor transformation. However, the mechanisms regulating the expression of these ligands are still not well understood. Under normal conditions, the NKG2D ligands were shown to be posttranscriptionally regulated by cellular microRNAs and RNA-binding proteins (RBPs). Thus far, only the 3' untranslated regions (UTRs) of MICA, MICB, and UL16-binding protein 2 were shown to be regulated by RBPs and microRNAs, usually resulting in their downregulation. In this study we investigated whether MICB expression is controlled by RBPs through its 5'UTR. We used an RNA pull-down assay followed by mass spectrometry and identified vigilin, a ubiquitously expressed multifunctional RNA-binding protein. We demonstrated that vigilin binds and negatively regulates MICB expression through its 5'UTR. Additionally, vigilin downregulation in target cells led to a significant increase in NK cell activation against said target cells. Taken together, we have discovered a novel mode of MICB regulation.


Assuntos
Antígenos de Histocompatibilidade Classe I/metabolismo , Vigilância Imunológica , Células Matadoras Naturais/imunologia , Proteínas de Ligação a RNA/metabolismo , Estresse Fisiológico/imunologia , Regiões 5' não Traduzidas/genética , Linhagem Celular Tumoral , Regulação da Expressão Gênica/genética , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Ligantes , Ativação Linfocitária , Subfamília K de Receptores Semelhantes a Lectina de Células NK/agonistas , Ligação Proteica , RNA Interferente Pequeno/genética , Proteínas de Ligação a RNA/genética
10.
Eur J Immunol ; 47(4): 692-703, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28191644

RESUMO

Natural killer (NK) cells are capable of killing various pathogens upon stimulation of activating receptors. Human metapneumovirus (HMPV) is a respiratory virus, which was discovered in 2001 and is responsible for acute respiratory tract infection in infants and children worldwide. HMPV infection is very common, infecting around 70% of all children under the age of five. Under immune suppressive conditions, HMPV infection can be fatal. Not much is known on how NK cells respond to HMPV. In this study, using reporter assays and NK-cell cytotoxicity assays performed with human and mouse NK cells, we demonstrated that the NKp46-activating receptor and its mouse orthologue Ncr1, both members of the natural cytotoxicity receptor (NCR) family, recognized an unknown ligand expressed by HMPV-infected human cells. We demonstrated that MHC class I is upregulated and MICA is downregulated upon HMPV infection. We also characterized mouse NK-cell phenotype in the blood and the lungs of HMPV-infected mice and found that lung NK cells are more activated and expressing NKG2D, CD43, CD27, KLRG1, and CD69 compared to blood NK cells regardless of HMPV infection. Finally, we demonstrated, using Ncr1-deficient mice, that NCR1 plays a critical role in controlling HMPV infection.


Assuntos
Antígenos Ly/metabolismo , Células Matadoras Naturais/imunologia , Pulmão/imunologia , Metapneumovirus/imunologia , Receptor 1 Desencadeador da Citotoxicidade Natural/metabolismo , Infecções por Paramyxoviridae/imunologia , Animais , Antígenos Ly/genética , Criança , Citotoxicidade Imunológica , Células HEK293 , Humanos , Lactente , Células Matadoras Naturais/virologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Receptor 1 Desencadeador da Citotoxicidade Natural/genética , Carga Viral
11.
J Virol ; 91(19)2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28724773

RESUMO

The recent approval of oncolytic virus for therapy of melanoma patients has increased the need for precise evaluation of the mechanisms by which oncolytic viruses affect tumor growth. Here we show that the human NK cell-activating receptor NKp46 and the orthologous mouse protein NCR1 recognize the reovirus sigma1 protein in a sialic-acid-dependent manner. We identify sites of NKp46/NCR1 binding to sigma1 and show that sigma1 binding by NKp46/NCR1 leads to NK cell activation in vitro Finally, we demonstrate that NCR1 activation is essential for reovirus-based therapy in vivo Collectively, we have identified sigma1 as a novel ligand for NKp46/NCR1 and demonstrated that NKp46/NCR1 is needed both for clearance of reovirus infection and for reovirus-based tumor therapy.IMPORTANCE Reovirus infects much of the population during childhood, causing mild disease, and hence is considered to be efficiently controlled by the immune system. Reovirus also specifically infects tumor cells, leading to tumor death, and is currently being tested in human clinical trials for cancer therapy. The mechanisms by which our immune system controls reovirus infection and tumor killing are not well understood. We report here that natural killer (NK) cells recognize a viral protein named sigma1 through the NK cell-activating receptor NKp46. Using several mouse tumor models, we demonstrate the importance of NK cells in protection from reovirus infection and in reovirus killing of tumors in vivo Collectively, we identify a new ligand for the NKp46 receptor and provide evidence for the importance of NKp46 in the control of reovirus infections and in reovirus-based cancer therapy.


Assuntos
Antígenos Ly/metabolismo , Células Matadoras Naturais/imunologia , Orthoreovirus Mamífero 3/metabolismo , Receptor 1 Desencadeador da Citotoxicidade Natural/metabolismo , Terapia Viral Oncolítica/métodos , Vírus Oncolíticos/metabolismo , Proteínas Virais/metabolismo , Animais , Sítios de Ligação , Chlorocebus aethiops , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Humanos , Ativação Linfocitária/imunologia , Melanoma/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ácido N-Acetilneuramínico/metabolismo , Células Vero , Proteínas Virais/genética
12.
PLoS Pathog ; 9(8): e1003568, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23966863

RESUMO

Natural killer (NK) cells are innate immune cells able to rapidly kill virus-infected and tumor cells. Two NK cell populations are found in the blood; the majority (90%) expresses the CD16 receptor and also express the CD56 protein in intermediate levels (CD56(Dim) CD16(Pos)) while the remaining 10% are CD16 negative and express CD56 in high levels (CD56(Bright) CD16(Neg)). NK cells also reside in some tissues and traffic to various infected organs through the usage of different chemokines and chemokine receptors. Kaposi's sarcoma-associated herpesvirus (KSHV) is a human virus that has developed numerous sophisticated and versatile strategies to escape the attack of immune cells such as NK cells. Here, we investigate whether the KSHV derived cytokine (vIL-6) and chemokines (vMIP-I, vMIP-II, vMIP-III) affect NK cell activity. Using transwell migration assays, KSHV infected cells, as well as fusion and recombinant proteins, we show that out of the four cytokine/chemokines encoded by KSHV, vMIP-II is the only one that binds to the majority of NK cells, affecting their migration. We demonstrate that vMIP-II binds to two different receptors, CX3CR1 and CCR5, expressed by naïve CD56(Dim) CD16(Pos) NK cells and activated NK cells, respectively. Furthermore, we show that the binding of vMIP-II to CX3CR1 and CCR5 blocks the binding of the natural ligands of these receptors, Fractalkine (Fck) and RANTES, respectively. Finally, we show that vMIP-II inhibits the migration of naïve and activated NK cells towards Fck and RANTES. Thus, we present here a novel mechanism in which KSHV uses a unique protein that antagonizes the activity of two distinct chemokine receptors to inhibit the migration of naïve and activated NK cells.


Assuntos
Fármacos Anti-HIV/farmacologia , Antagonistas dos Receptores CCR5 , Movimento Celular/efeitos dos fármacos , Quimiocinas/farmacologia , Herpesvirus Humano 8/química , Células Matadoras Naturais/efeitos dos fármacos , Receptores de Quimiocinas/antagonistas & inibidores , Receptor 1 de Quimiocina CX3C , Células Cultivadas , Quimiocina CCL5/metabolismo , Quimiocina CX3CL1/metabolismo , Citocinas/genética , Citocinas/metabolismo , Humanos , Immunoblotting , Interleucina-6 , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Reação em Cadeia da Polimerase , Receptores CCR5/genética , Receptores CCR5/metabolismo , Receptores de Quimiocinas/genética , Receptores de Quimiocinas/metabolismo
13.
J Infect Dis ; 210(3): 410-8, 2014 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-24532603

RESUMO

Natural Killer (NK) cells play a central role in the defense against viral infections and in the elimination of transformed cells. The recognition of pathogen-infected and tumor cells is controlled by inhibitory and activating receptors. We have previously shown that among the activating (killer) NK cell receptors the natural cytotoxicity receptors, NKp44 and NKp46, interact with the viral hemagglutinin (HA) protein expressed on the cell surface of influenza-virus-infected cells. We further showed that the interaction between NKp44/NKp46 and viral HA is sialic-acid dependent and that the recognition of HA by NKp44 and NKp46 leads to the elimination of the infected cells. Here we demonstrate that the influenza virus developed a counter-attack mechanism and that the virus uses its neuraminidase (NA) protein to prevent the recognition of HA by both the NKp44 and NKp46 receptors, resulting in reduced elimination of the infected cells by NK cells.


Assuntos
Vírus da Influenza A/fisiologia , Neuraminidase/metabolismo , Receptores de Células Matadoras Naturais/metabolismo , Animais , Linhagem Celular Tumoral , Hemaglutininas/metabolismo , Humanos , Camundongos , Neuraminidase/genética
14.
Blood ; 120(19): 3915-24, 2012 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-22983444

RESUMO

Recent evidence suggests that kindlin-3 is a major coactivator, required for most, if not all, integrin activities. Here we studied the function of kindlin-3 in regulating NK cell activation by studying a patient with kindlin-3 deficiency (leukocyte adhesion deficiency-III). We found that kindlin-3 is required for NK cell migration and adhesion under shear force. Surprisingly, we also found that kindlin-3 lowers the threshold for NK cell activation. Loss of kindlin-3 has a pronounced effect on NK cell-mediated cytotoxicity triggered by single activating receptors. In contrast, for activation through multiple receptors, kindlin-3 deficiency is overcome and target cells killed. The realization that NK cell activity is impaired, but not absent in leukocyte adhesion deficiency, may lead to the development of more efficient therapy for this rare disease.


Assuntos
Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Síndrome da Aderência Leucocítica Deficitária/imunologia , Síndrome da Aderência Leucocítica Deficitária/metabolismo , Proteínas de Membrana/deficiência , Proteínas de Neoplasias/deficiência , Actinas/química , Actinas/metabolismo , Adesão Celular/genética , Adesão Celular/imunologia , Movimento Celular/genética , Movimento Celular/imunologia , Células Cultivadas , Códon de Terminação , Citotoxicidade Imunológica , Genótipo , Humanos , Síndrome da Aderência Leucocítica Deficitária/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Linhagem , Multimerização Proteica , Transporte Proteico , Receptores de IgG/imunologia , Receptores de IgG/metabolismo , Receptores de Células Matadoras Naturais/imunologia , Receptores de Células Matadoras Naturais/metabolismo , Resistência ao Cisalhamento
15.
Front Immunol ; 14: 1231782, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37753084

RESUMO

In recent years, reoviruses have been of major interest in immunotherapy because of their oncolytic properties. Preclinical and clinical trials, in which reovirus was used for the treatment of melanoma and glioblastoma, have paved the way for future clinical use of reovirus. However, little is known about how reovirus infection affects the tumor microenvironment and immune response towards infected tumor cells. Studies have shown that reovirus can directly stimulate natural killer (NK) cells, but how reovirus affects cellular ligands on tumor cells, which are ultimately key to tumor recognition and elimination by NK cells, has not been investigated. We tested how reovirus infection affects the binding of the NK Group-2 member D (NKG2D) receptor, which is a dominant mediator of NK cell anti-tumor activity. Using models of human-derived melanoma and glioblastoma tumors, we demonstrated that NKG2D ligands are downregulated in tumor cells post-reovirus-infection due to the impaired translation of these ligands in reovirus-infected cells. Moreover, we showed that downregulation of NKG2D ligands significantly impaired the binding of NKG2D to infected tumor cells. We further demonstrated that reduced recognition of NKG2D ligands significantly alters NK cell anti-tumor cytotoxicity in human primary NK cells and in the NK cell line NK-92. Thus, this study provides novel insights into reovirus-host interactions and could lead to the development of novel reovirus-based therapeutics that enhance the anti-tumor immune response.


Assuntos
Glioblastoma , Melanoma , Orthoreovirus , Infecções por Reoviridae , Reoviridae , Humanos , Anticorpos Antivirais , Glioblastoma/terapia , Ligantes , Melanoma/terapia , Subfamília K de Receptores Semelhantes a Lectina de Células NK , Microambiente Tumoral
16.
J Cell Biol ; 222(9)2023 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-37389656

RESUMO

The GPI-anchoring pathway plays important roles in normal development and immune modulation. MHC Class I Polypeptide-related Sequence A (MICA) is a stress-induced ligand, downregulated by human cytomegalovirus (HCMV) to escape immune recognition. Its most prevalent allele, MICA*008, is GPI-anchored via an uncharacterized pathway. Here, we identify cleft lip and palate transmembrane protein 1-like protein (CLPTM1L) as a GPI-anchoring pathway component and show that during infection, the HCMV protein US9 downregulates MICA*008 via CLPTM1L. We show that the expression of some GPI-anchored proteins (CD109, CD59, and MELTF)-but not others (ULBP2, ULBP3)-is CLPTM1L-dependent, and further show that like MICA*008, MELTF is downregulated by US9 via CLPTM1L during infection. Mechanistically, we suggest that CLPTM1L's function depends on its interaction with a free form of PIG-T, normally a part of the GPI transamidase complex. We suggest that US9 inhibits this interaction and thereby downregulates the expression of CLPTM1L-dependent proteins. Altogether, we report on a new GPI-anchoring pathway component that is targeted by HCMV.


Assuntos
Infecções por Citomegalovirus , Proteínas de Membrana , Humanos , Alelos , Citomegalovirus , Proteínas de Membrana/genética , Proteínas de Neoplasias , Fatores de Transcrição , Infecções por Citomegalovirus/metabolismo
17.
Cell Rep ; 40(10): 111311, 2022 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-36070690

RESUMO

Antiretroviral therapy controls, but does not cure, HIV-1 infection due to a reservoir of rare CD4+ T cells harboring latent proviruses. Little is known about the transcriptional program of latent cells. Here, we report a strategy to enrich clones of latent cells carrying intact, replication-competent HIV-1 proviruses from blood based on their expression of unique T cell receptors. Latent cell enrichment enabled single-cell transcriptomic analysis of 1,050 CD4+ T cells belonging to expanded clones harboring intact HIV-1 proviruses from 6 different individuals. The analysis reveals that most of these cells are T effector memory cells that are enriched for expression of HLA-DR, HLA-DP, CD74, CCL5, granzymes A and K, cystatin F, LYAR, and DUSP2. We conclude that expanded clones of latent cells carrying intact HIV-1 proviruses persist preferentially in a distinct CD4+ T cell population, opening possibilities for eradication.


Assuntos
Infecções por HIV , Soropositividade para HIV , HIV-1 , Linfócitos T CD4-Positivos/metabolismo , Células Clonais , Proteínas de Ligação a DNA/metabolismo , Expressão Gênica , HIV-1/genética , HIV-1/metabolismo , Humanos , Proteínas Nucleares/metabolismo , Provírus/genética , Provírus/metabolismo , Latência Viral/genética
18.
Cell Mol Immunol ; 19(2): 234-244, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34992275

RESUMO

Global pandemics caused by influenza or coronaviruses cause severe disruptions to public health and lead to high morbidity and mortality. There remains a medical need for vaccines against these pathogens. CMV (cytomegalovirus) is a ß-herpesvirus that induces uniquely robust immune responses in which remarkably large populations of antigen-specific CD8+ T cells are maintained for a lifetime. Hence, CMV has been proposed and investigated as a novel vaccine vector for expressing antigenic peptides or proteins to elicit protective cellular immune responses against numerous pathogens. We generated two recombinant murine CMV (MCMV) vaccine vectors expressing hemagglutinin (HA) of influenza A virus (MCMVHA) or the spike protein of severe acute respiratory syndrome coronavirus 2 (MCMVS). A single injection of MCMVs expressing either viral protein induced potent neutralizing antibody responses, which strengthened over time. Importantly, MCMVHA-vaccinated mice were protected from illness following challenge with the influenza virus, and we excluded that this protection was due to the effects of memory T cells. Conclusively, we show here that MCMV vectors induce not only long-term cellular immunity but also humoral responses that provide long-term immune protection against clinically relevant respiratory pathogens.


Assuntos
Vacinas contra COVID-19/imunologia , COVID-19/prevenção & controle , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Imunidade Humoral , Vírus da Influenza A/imunologia , Vacinas contra Influenza/imunologia , Infecções por Orthomyxoviridae/prevenção & controle , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinação/métodos , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , COVID-19/virologia , Chlorocebus aethiops , Citomegalovirus/imunologia , Cães , Feminino , Células HEK293 , Humanos , Imunidade Celular , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Infecções por Orthomyxoviridae/virologia , Células Vero
19.
J Virol ; 84(8): 3993-4001, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20130050

RESUMO

Today, global attention is focused on two influenza virus strains: the current pandemic strain, swine origin influenza virus (H1N1-2009), and the highly pathogenic avian influenza virus, H5N1. At present, the infection caused by the H1N1-2009 is moderate, with mortality rates of less <1%. In contrast, infection with the H5N1 virus resulted in high mortality rates, and ca. 60% of the infected patients succumb to the infection. Thus, one of the world greatest concerns is that the H5N1 virus will evolve to allow an efficient human infection and human-to-human transmission. Natural killer (NK) cells are one of the innate immune components playing an important role in fighting against influenza viruses. One of the major NK activating receptors involved in NK cell cytotoxicity is NKp46. We previously demonstrated that NKp46 recognizes the hemagglutinin proteins of B and A influenza virus strains. Whether NKp46 could also interact with H1N1-2009 virus or with the avian influenza virus is still unknown. We analyzed the immunological properties of both the avian and the H1N1-2009 influenza viruses. We show that NKp46 recognizes the hemagglutinins of H1N1-2009 and H5 and that this recognition leads to virus killing both in vitro and in vivo. However, importantly, while the swine H1-NKp46 interactions lead to the direct killing of the infected cells, the H5-NKp46 interactions were unable to elicit direct killing, probably because the NKp46 binding sites for these two viruses are different.


Assuntos
Vírus da Influenza A Subtipo H1N1/imunologia , Virus da Influenza A Subtipo H5N1/imunologia , Células Matadoras Naturais/imunologia , Sequência de Aminoácidos , Animais , Linhagem Celular , Sobrevivência Celular , Células Cultivadas , Citotoxicidade Imunológica , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Receptor 1 Desencadeador da Citotoxicidade Natural/metabolismo , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/virologia , Ligação Proteica
20.
Mol Ther Oncol ; 32(3): 200865, 2024 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-39297072
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