ABSTRACT
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.
ABSTRACT
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.
Subject(s)
Glioblastoma , Melanoma , Orthoreovirus , Reoviridae Infections , Reoviridae , Humans , Antibodies, Viral , Glioblastoma/therapy , Ligands , Melanoma/therapy , NK Cell Lectin-Like Receptor Subfamily K , Tumor MicroenvironmentABSTRACT
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).
Subject(s)
Dental Implants , Peri-Implantitis , Periodontitis , Humans , Male , Female , Middle Aged , Aged , Peri-Implantitis/metabolism , Flow Cytometry , Periodontitis/metabolism , LeukocytesABSTRACT
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.
Subject(s)
Cytomegalovirus Infections , Membrane Proteins , Humans , Alleles , Cytomegalovirus , Membrane Proteins/genetics , Neoplasm Proteins , Transcription Factors , Cytomegalovirus Infections/metabolismABSTRACT
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.
Subject(s)
HIV Infections , HIV Seropositivity , HIV-1 , CD4-Positive T-Lymphocytes/metabolism , Clone Cells , DNA-Binding Proteins/metabolism , Gene Expression , HIV-1/genetics , HIV-1/metabolism , Humans , Nuclear Proteins/metabolism , Proviruses/genetics , Proviruses/metabolism , Virus Latency/geneticsABSTRACT
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.
Subject(s)
BNT162 Vaccine/administration & dosage , COVID-19 , Coronavirus Nucleocapsid Proteins , Databases, Nucleic Acid , Genome, Viral , Mutation , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Adult , Aged , COVID-19/genetics , COVID-19/immunology , COVID-19/prevention & control , Child , Coronavirus Nucleocapsid Proteins/genetics , Coronavirus Nucleocapsid Proteins/immunology , Female , HEK293 Cells , Humans , Male , Middle Aged , Phosphoproteins/genetics , Phosphoproteins/immunology , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Sequence Analysis, RNA , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunologyABSTRACT
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.
Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , Hemagglutinin Glycoproteins, Influenza Virus/immunology , Immunity, Humoral , Influenza A virus/immunology , Influenza Vaccines/immunology , Orthomyxoviridae Infections/prevention & control , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Vaccination/methods , Animals , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , COVID-19/virology , Chlorocebus aethiops , Cytomegalovirus/immunology , Dogs , Female , HEK293 Cells , Humans , Immunity, Cellular , Madin Darby Canine Kidney Cells , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Orthomyxoviridae Infections/virology , Vero CellsABSTRACT
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.
Subject(s)
Arthritis, Rheumatoid , BNT162 Vaccine , COVID-19 , Immunity, Humoral , Janus Kinase Inhibitors , Antibodies, Neutralizing , Antibodies, Viral , Arthritis, Rheumatoid/drug therapy , BNT162 Vaccine/immunology , COVID-19/prevention & control , Humans , Immunoglobulin G , Janus Kinase Inhibitors/therapeutic use , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , VaccinationABSTRACT
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.
Subject(s)
CD4 Antigens , HIV-1 , Nanoparticles , Virion , Anti-HIV Agents , CD4 Antigens/chemistry , CD4 Antigens/metabolism , Cell Line , HIV-1/chemistry , HIV-1/genetics , HIV-1/metabolism , Humans , Molecular Mimicry , Nanomedicine/methods , Nanoparticles/chemistry , Nanoparticles/metabolism , Virion/chemistry , Virion/metabolismABSTRACT
Monotherapy of HIV-1 infection with single antiretroviral agents is ineffective because error-prone HIV-1 replication leads to the production of drug-resistant viral variants1,2. Combinations of drugs can establish long-term control, however, antiretroviral therapy (ART) requires daily dosing, can cause side effects and does not eradicate the infection3,4. Although anti-HIV-1 antibodies constitute a potential alternative to ART5,6, treatment of viremic individuals with a single antibody also results in emergence of resistant viral variants7-9. Moreover, combinations of first-generation anti-HIV-1 broadly neutralizing antibodies (bNAbs) had little measurable effect on the infection10-12. Here we report on a phase 1b clinical trial ( NCT02825797 ) in which two potent bNAbs, 3BNC11713 and 10-107414, were administered in combination to seven HIV-1 viremic individuals. Infusions of 30 mg kg-1 of each of the antibodies were well-tolerated. In the four individuals with dual antibody-sensitive viruses, immunotherapy resulted in an average reduction in HIV-1 viral load of 2.05 log10 copies per ml that remained significantly reduced for three months following the first of up to three infusions. In addition, none of these individuals developed resistance to both antibodies. Larger studies will be necessary to confirm the efficacy of antibody combinations in reducing HIV-1 viremia and limiting the emergence of resistant viral variants.
Subject(s)
Antibodies, Neutralizing/administration & dosage , HIV Infections/drug therapy , Immunotherapy , Viremia/drug therapy , Adolescent , Adult , Aged , Antibodies, Neutralizing/adverse effects , Antiretroviral Therapy, Highly Active , Female , HIV Infections/virology , HIV Seropositivity , HIV-1/pathogenicity , Humans , Male , Middle Aged , Viral Load/drug effects , Viremia/virology , Young AdultABSTRACT
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.
Subject(s)
HIV Antibodies/immunology , HIV Infections/immunology , HIV-1/physiology , AIDS Vaccines/immunology , Animals , CD4 Antigens/chemistry , CD4 Antigens/metabolism , Disease Models, Animal , HIV Envelope Protein gp120/chemistry , HIV Envelope Protein gp120/metabolism , HIV-1/genetics , Humans , Mice , Mutation , Receptors, Fc/immunology , T-Lymphocytes/virologyABSTRACT
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.
Subject(s)
Antigens, Ly/metabolism , Killer Cells, Natural/immunology , Mammalian orthoreovirus 3/metabolism , Natural Cytotoxicity Triggering Receptor 1/metabolism , Oncolytic Virotherapy/methods , Oncolytic Viruses/metabolism , Viral Proteins/metabolism , Animals , Binding Sites , Chlorocebus aethiops , Disease Models, Animal , Enzyme-Linked Immunosorbent Assay , Humans , Lymphocyte Activation/immunology , Melanoma/therapy , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , N-Acetylneuraminic Acid/metabolism , Vero Cells , Viral Proteins/geneticsABSTRACT
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.
Subject(s)
Histocompatibility Antigens Class I/metabolism , Immunologic Surveillance , Killer Cells, Natural/immunology , RNA-Binding Proteins/metabolism , Stress, Physiological/immunology , 5' Untranslated Regions/genetics , Cell Line, Tumor , Gene Expression Regulation/genetics , Histocompatibility Antigens Class I/genetics , Humans , Ligands , Lymphocyte Activation , NK Cell Lectin-Like Receptor Subfamily K/agonists , Protein Binding , RNA, Small Interfering/genetics , RNA-Binding Proteins/geneticsABSTRACT
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.
Subject(s)
Antigens, Ly/metabolism , Killer Cells, Natural/immunology , Lung/immunology , Metapneumovirus/immunology , Natural Cytotoxicity Triggering Receptor 1/metabolism , Paramyxoviridae Infections/immunology , Animals , Antigens, Ly/genetics , Child , Cytotoxicity, Immunologic , HEK293 Cells , Humans , Infant , Killer Cells, Natural/virology , Lymphocyte Activation , Mice , Mice, Inbred BALB C , Mice, Knockout , Natural Cytotoxicity Triggering Receptor 1/genetics , Viral LoadABSTRACT
Some HIV-1-infected patients develop broad and potent HIV-1 neutralizing antibodies (bNAbs) that when passively transferred to mice or macaques can treat or prevent infection. However, bNAbs typically fail to neutralize coexisting autologous viruses due to antibody-mediated selection against sensitive viral strains. We describe an HIV-1 controller expressing HLA-B57*01 and HLA-B27*05 who maintained low viral loads for 30 years after infection and developed broad and potent serologic activity against HIV-1. Neutralization was attributed to three different bNAbs targeting nonoverlapping sites on the HIV-1 envelope trimer (Env). One of the three, BG18, an antibody directed against the glycan-V3 portion of Env, is the most potent member of this class reported to date and, as revealed by crystallography and electron microscopy, recognizes HIV-1 Env in a manner that is distinct from other bNAbs in this class. Single-genome sequencing of HIV-1 from serum samples obtained over a period of 9 years showed a diverse group of circulating viruses, 88.5% (31 of 35) of which remained sensitive to at least one of the temporally coincident autologous bNAbs and the individual's serum. Thus, bNAb-sensitive strains of HIV-1 coexist with potent neutralizing antibodies that target the virus and may contribute to control in this individual. When administered as a mix, the three bNAbs controlled viremia in HIV-1YU2-infected humanized mice. Our finding suggests that combinations of bNAbs may contribute to control of HIV-1 infection.
Subject(s)
Antibodies, Neutralizing/immunology , HIV Antibodies/immunology , HIV-1/immunology , HLA-B Antigens/immunology , HLA-B27 Antigen/immunology , Animals , B-Lymphocytes/metabolism , Cohort Studies , Crystallography, X-Ray , Epitopes/immunology , HEK293 Cells , HIV Infections/immunology , Humans , Mice , Mice, Transgenic , Neutralization Tests , Viral Load , Viremia , env Gene Products, Human Immunodeficiency Virus/immunologyABSTRACT
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.
Subject(s)
Anti-HIV Agents/administration & dosage , Antibodies, Neutralizing/immunology , HIV Antibodies/immunology , HIV Infections/drug therapy , HIV Infections/virology , HIV-1/growth & development , HIV-1/immunology , Adolescent , Adult , Aged , Anti-HIV Agents/pharmacology , Anti-HIV Agents/therapeutic use , Antibodies, Monoclonal, Humanized , Antibodies, Neutralizing/administration & dosage , Antibodies, Neutralizing/therapeutic use , Binding Sites/drug effects , Binding Sites/immunology , Broadly Neutralizing Antibodies , CD4 Antigens/metabolism , Disease Reservoirs/virology , Drug Administration Schedule , Female , HIV Antibodies/administration & dosage , HIV Antibodies/therapeutic use , HIV Envelope Protein gp160/antagonists & inhibitors , HIV Envelope Protein gp160/chemistry , HIV Envelope Protein gp160/immunology , HIV Envelope Protein gp160/metabolism , HIV Infections/immunology , HIV-1/drug effects , Historically Controlled Study , Humans , Male , Middle Aged , Proviruses/drug effects , Proviruses/growth & development , Proviruses/immunology , Time Factors , Tissue Distribution , Viral Load/drug effects , Viral Load/immunology , Young AdultABSTRACT
Cells in our body can induce hundreds of antiviral genes following virus sensing, many of which remain largely uncharacterized. CEACAM1 has been previously shown to be induced by various innate systems; however, the reason for such tight integration to innate sensing systems was not apparent. Here, we show that CEACAM1 is induced following detection of HCMV and influenza viruses by their respective DNA and RNA innate sensors, IFI16 and RIG-I. This induction is mediated by IRF3, which bound to an ISRE element present in the human, but not mouse, CEACAM1 promoter. Furthermore, we demonstrate that, upon induction, CEACAM1 suppresses both HCMV and influenza viruses in an SHP2-dependent process and achieves this broad antiviral efficacy by suppressing mTOR-mediated protein biosynthesis. Finally, we show that CEACAM1 also inhibits viral spread in ex vivo human decidua organ culture.
Subject(s)
Antigens, CD/metabolism , Cell Adhesion Molecules/metabolism , Cytomegalovirus/physiology , Orthomyxoviridae/physiology , Animals , Cytomegalovirus Infections/metabolism , Cytomegalovirus Infections/virology , DEAD Box Protein 58/metabolism , DNA, Viral/metabolism , Humans , Influenza, Human/metabolism , Influenza, Human/virology , Interferon Regulatory Factor-3/metabolism , Mice , Organ Culture Techniques , Protein Biosynthesis , Receptors, Immunologic , TOR Serine-Threonine Kinases/metabolism , Virus ReplicationABSTRACT
Natural Killer (NK) cells are critical in the defense against viruses in general and against influenza in particular. We previously demonstrated that the activating NK cell receptor NKp46 is involved in the killing of influenza-virus infected cells through its interaction with viral hemagglutinin (HA). Furthermore, the recognition by NKp46 and consequent elimination of influenza infected cells were determined to be sialic-acid dependent. Here, we show that the human co-activating receptors 2B4 and NTB-A directly recognize the viral HA protein and co-stimulate killing by NK cells. We demonstrate that the 2B4/NTB-A-HA interactions require the sialylation of these receptors, and we identified the binding sites mediating these interactions. We also show that the virus counters these interactions through its neuraminidase (NA) protein. These results emphasize the critical role played by NK cells in eliminating influenza, a significant cause of worldwide morbidity and mortality.
Subject(s)
Hemagglutinin Glycoproteins, Influenza Virus/immunology , Influenza, Human/immunology , Killer Cells, Natural/immunology , Signaling Lymphocytic Activation Molecule Family/immunology , Cell Line, Tumor , Cytotoxicity, Immunologic/immunology , Humans , Influenza A Virus, H1N1 SubtypeABSTRACT
Genetic deficiencies provide insights into gene function in humans. Here we describe a patient with a very rare genetic deficiency of ADAM17. We show that the patient's PBMCs had impaired cytokine secretion in response to LPS stimulation, correlating with the clinical picture of severe bacteremia from which the patient suffered. ADAM17 was shown to cleave CD16, a major NK killer receptor. Functional analysis of patient's NK cells demonstrated that his NK cells express normal levels of activating receptors and maintain high surface levels of CD16 following mAb stimulation. Activation of individual NK cell receptors showed that the patient's NK cells are more potent when activated directly by CD16, albeit no difference was observed in Antibody Depedent Cytotoxicity (ADCC) assays. Our data suggest that ADAM17 inhibitors currently considered for clinical use to boost CD16 activity should be cautiously applied, as they might have severe side effects resulting from impaired cytokine secretion.
Subject(s)
ADAM Proteins/deficiency , Cytokines/metabolism , Immunologic Deficiency Syndromes/enzymology , Killer Cells, Natural/enzymology , Leukocytes, Mononuclear/enzymology , Lymphocyte Activation , ADAM Proteins/genetics , ADAM Proteins/immunology , ADAM17 Protein , Antibody-Dependent Cell Cytotoxicity , Cell Line, Tumor , Child, Preschool , Cytokines/immunology , Fatal Outcome , GPI-Linked Proteins/immunology , GPI-Linked Proteins/metabolism , Genetic Predisposition to Disease , Humans , Immunity, Innate , Immunologic Deficiency Syndromes/diagnosis , Immunologic Deficiency Syndromes/genetics , Immunologic Deficiency Syndromes/immunology , Killer Cells, Natural/immunology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Lipopolysaccharides/pharmacology , Male , Phenotype , Receptors, IgG/immunology , Receptors, IgG/metabolismABSTRACT
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.