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Here we report the characterization of 17T2, a SARS-CoV-2 pan-neutralizing human monoclonal antibody isolated from a COVID-19 convalescent individual infected during the first pandemic wave. 17T2 is a class 1 VH1-58/κ3-20 antibody, derived from a receptor binding domain (RBD)-specific IgA+ memory B cell, with a broad neutralizing activity against former and new SARS-CoV-2 variants, including XBB.1.16 and BA.2.86 Omicron subvariants. Consistently, 17T2 demonstrates in vivo prophylactic and therapeutic activity against Omicron BA.1.1 infection in K18-hACE2 mice. Cryo-electron microscopy reconstruction shows that 17T2 binds the BA.1 spike with the RBD in "up" position and blocks the receptor binding motif, as other structurally similar antibodies do, including S2E12. Yet, unlike S2E12, 17T2 retains its neutralizing activity against all variants tested, probably due to a larger RBD contact area. These results highlight the impact of small structural antibody changes on neutralizing performance and identify 17T2 as a potential candidate for future clinical interventions.
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Anticuerpos Monoclonales , COVID-19 , Humanos , Animales , Ratones , SARS-CoV-2 , Microscopía por Crioelectrón , Anticuerpos Monoclonales Humanizados , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Glicoproteína de la Espiga del Coronavirus/genéticaRESUMEN
BACKGROUND: The vacuoles, E1-enzyme, X linked, autoinflammatory and somatic (VEXAS) syndrome is an adult-onset autoinflammatory disease (AID) due to postzygotic UBA1 variants. OBJECTIVES: To investigate the presence of VEXAS syndrome among patients with adult-onset undiagnosed AID. Additional studies evaluated the mosaicism distribution and the circulating cytokines. METHODS: Gene analyses were performed by both Sanger and amplicon-based deep sequencing. Patients' data were collected from their medical charts. Cytokines were quantified by Luminex. RESULTS: Genetic analyses of enrolled patients (n=42) identified 30 patients carrying UBA1 pathogenic variants, with frequencies compatible for postzygotic variants. All patients were male individuals who presented with a late-onset disease (mean 67.5 years; median 67.0 years) characterised by cutaneous lesions (90%), fever (66.7%), pulmonary manifestations (66.7%) and arthritis (53.3%). Macrocytic anaemia and increased erythrocyte sedimentation rate and ferritin were the most relevant analytical abnormalities. Glucocorticoids ameliorated the inflammatory manifestations, but most patients became glucocorticoid-dependent. Positive responses were obtained when targeting the haematopoietic component of the disease with either decitabine or allogeneic haematopoietic stem cell transplantation. Additional analyses detected the UBA1 variants in both haematopoietic and non-haematopoietic tissues. Finally, analysis of circulating cytokines did not identify inflammatory mediators of the disease. CONCLUSION: Thirty patients with adult-onset AID were definitively diagnosed with VEXAS syndrome through genetic analyses. Despite minor interindividual differences, their main characteristics were in concordance with previous reports. We detected for the first time the UBA1 mosaicism in non-haematopoietic tissue, which questions the previous concept of myeloid-restricted mosaicism and may have conceptual consequences for the disease mechanisms.
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Artritis , Mosaicismo , Adulto , Humanos , Masculino , Femenino , Citocinas/genética , Ferritinas , Glucocorticoides , MutaciónRESUMEN
Somatic hypermutation (SHM) is an important diversification mechanism that plays a part in the creation of immune memory. Immunoglobulin (Ig) variable region gene lineage trees were used over the last four decades to model SHM and the selection mechanisms operating on B cell clones. We hereby present IgTreeZ (Immunoglobulin Tree analyZer), a python-based tool that analyses many aspects of Ig gene lineage trees and their repertoires. Using simulations, we show that IgTreeZ can be reliably used for mutation and selection analyses. We used IgTreeZ on empirical data, found evidence for different mutation patterns in different B cell subpopulations, and gained insights into antigen-driven selection in corona virus disease 19 (COVID-19) patients. Most importantly, we show that including the CDR3 regions in selection analyses - which is only possible if these analyses are lineage tree-based - is crucial for obtaining correct results. Overall, we present a comprehensive lineage tree analysis tool that can reveal new biological insights into B cell repertoire dynamics.
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COVID-19 , Genes de Inmunoglobulinas , Humanos , Región Variable de Inmunoglobulina/genética , Linfocitos B , Células ClonalesRESUMEN
The massive COVID-19 vaccine purchases made by high-income countries have resulted in important sample losses, mainly due to the complexity of their handling. Here, we evaluated the possibility of preserving the immunogenicity of COVID-19 mRNA vaccines after re-freezing vials, following the extraction of the maximum possible number of samples, as an alternative approach to minimizing their wastage. Thus, we exposed the vaccine vials to different re-freezing conditions and evaluated mRNA integrity and the effects in mice after in vivo administration. We reveal that the mRNA integrity of Comirnaty® and Spikevax® vaccines remained unaffected after re-freezing during 1 month at -20 °C or -80 °C. The immunological responses also remained unchanged in mice after these re-freezing conditions and no apparent side effects were revealed. The preservation of mRNA integrity and immunogenicity under these handling conditions opens the possibility of re-freezing the mRNA COVID-19 vaccine vials to limit their wastage and to facilitate vaccination processes.
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B cells, which are critical for intestinal homeostasis, remain understudied in ulcerative colitis (UC). In this study, we recruited three cohorts of patients with UC (primary cohort, n = 145; validation cohort 1, n = 664; and validation cohort 2, n = 143) to comprehensively define the landscape of B cells during UC-associated intestinal inflammation. Using single-cell RNA sequencing, single-cell IgH gene sequencing and protein-level validation, we mapped the compositional, transcriptional and clonotypic landscape of mucosal and circulating B cells. We found major perturbations within the mucosal B cell compartment, including an expansion of naive B cells and IgG+ plasma cells with curtailed diversity and maturation. Furthermore, we isolated an auto-reactive plasma cell clone targeting integrin αvß6 from inflamed UC intestines. We also identified a subset of intestinal CXCL13-expressing TFH-like T peripheral helper cells that were associated with the pathogenic B cell response. Finally, across all three cohorts, we confirmed that changes in intestinal humoral immunity are reflected in circulation by the expansion of gut-homing plasmablasts that correlates with disease activity and predicts disease complications. Our data demonstrate a highly dysregulated B cell response in UC and highlight a potential role of B cells in disease pathogenesis.
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Colitis Ulcerosa , Células Plasmáticas , Linfocitos B , Colitis Ulcerosa/genética , Humanos , Mucosa Intestinal/patología , Recuento de Linfocitos , Linfocitos T Colaboradores-InductoresRESUMEN
Immunotherapy is a mainstay of non-small cell lung cancer (NSCLC) management. While tumor mutational burden (TMB) correlates with response to immunotherapy, little is known about the relationship between the baseline immune response and tumor genotype. Using single-cell RNA sequencing, we profiled 361,929 cells from 35 early-stage NSCLC lesions. We identified a cellular module consisting of PDCD1+CXCL13+ activated T cells, IgG+ plasma cells, and SPP1+ macrophages, referred to as the lung cancer activation module (LCAMhi). We confirmed LCAMhi enrichment in multiple NSCLC cohorts, and paired CITE-seq established an antibody panel to identify LCAMhi lesions. LCAM presence was found to be independent of overall immune cell content and correlated with TMB, cancer testis antigens, and TP53 mutations. High baseline LCAM scores correlated with enhanced NSCLC response to immunotherapy even in patients with above median TMB, suggesting that immune cell composition, while correlated with TMB, may be a nonredundant biomarker of response to immunotherapy.
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Carcinoma de Pulmón de Células no Pequeñas/inmunología , Inmunoterapia/métodos , Neoplasias Pulmonares/inmunología , Análisis de la Célula Individual/métodos , HumanosRESUMEN
mRNA-based vaccines effectively induce protective neutralizing antibodies against SARS-CoV-2, the etiological agent of COVID-19. Yet, the kinetics and compositional patterns of vaccine-induced antibody responses to the original strain and emerging variants of concern remain largely unknown. Here we characterized serum antibody classes and subclasses targeting the spike receptor-binding domain of SARS-CoV-2 wild type and α, ß, γ and δ variants in a longitudinal cohort of SARS-CoV-2 naïve and COVID-19 recovered individuals receiving the mRNA-1273 vaccine. We found that mRNA-1273 vaccine recipients developed a SARS-CoV-2-specific antibody response with a subclass profile comparable to that induced by natural infection. Importantly, these antibody responses targeted both wild type SARS-CoV-2 as well as its α, ß, γ and δ variants. Following primary vaccination, individuals with pre-existing immunity showed higher induction of all antibodies but IgG3 compared to SARS-CoV-2-naïve subjects. Unlike naïve individuals, COVID-19 recovered subjects did not mount a recall antibody response upon the second vaccine dose. In these individuals, secondary immunization resulted in a slight reduction of IgG1 against the receptor-binding domain of ß and γ variants. Despite the lack of recall humoral response, vaccinees with pre-existing immunity still showed higher titers of IgG1 and IgA to all variants analyzed compared to fully vaccinated naïve individuals. Our findings indicate that mRNA-1273 vaccine triggered cross-variant antibody responses with distinct profiles in vaccinees with or without pre-existing immunity and suggest that individuals with prior history of SARS-CoV-2 infection may not benefit from the second mRNA vaccine dose with the current standard regimen.
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Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , Vacunas contra la COVID-19/inmunología , COVID-19/inmunología , SARS-CoV-2/inmunología , Vacuna nCoV-2019 mRNA-1273 , Adulto , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , COVID-19/prevención & control , Convalecencia , Femenino , Humanos , Inmunoglobulina A/sangre , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Inmunoglobulina M/sangre , Estudios Longitudinales , Masculino , España , Glicoproteína de la Espiga del Coronavirus/inmunología , VacunaciónRESUMEN
OBJECTIVES: SARS-CoV-2 infection induces virus-reactive memory B cells expressing unmutated antibodies, which hints at their emergence from naïve B cells. Yet, the dynamics of virus-specific naïve B cells and their impact on immunity and immunopathology remain unclear. METHODS: We longitudinally profiled SARS-CoV-2-specific B-cell responses in 25 moderate-to-severe COVID-19 patients by high-dimensional flow cytometry and isotyping and subtyping ELISA. We also explored the relationship of B-cell responses to SARS-CoV-2 with the activation of effector and regulatory cells from the innate or adaptive immune system. RESULTS: We found a virus-specific antibody response with a broad spectrum of classes and subclasses during acute infection, which evolved into an IgG1-dominated response during convalescence. Acute infection was associated with increased mature B-cell progenitors in the circulation and the unexpected expansion of virus-targeting naïve-like B cells. The latter further augmented during convalescence together with virus-specific memory B cells. In addition to a transitory increase in tissue-homing CXCR3+ plasmablasts and extrafollicular memory B cells, most COVID-19 patients showed persistent activation of CD4+ and CD8+ T cells along with transient or long-lasting changes of key innate immune cells. Remarkably, virus-specific antibodies and the frequency of naïve B cells were among the major variables defining distinct immune signatures associated with disease severity and inflammation. CONCLUSION: Aside from providing new insights into the complexity of the immune response to SARS-CoV-2, our findings indicate that the de novo recruitment of mature B-cell precursors into the periphery may be central to the induction of antiviral immunity.
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Stromal-derived follicular dendritic cells (FDCs) are essential for germinal centers (GCs), the site where B cells maturate their antibodies. FDCs present native antigen to B cells and maintain a CXCL13 gradient to form the B cell follicle. Yet despite their essential role, the transcriptome of human FDCs remains undefined. Using single-cell RNA sequencing and microarray, we provided the transcriptome of these enigmatic cells as a comprehensive resource. Key genes were validated by flow cytometry and microscopy. Surprisingly, marginal reticular cells (MRCs) rather than FDCs expressed B cell activating factor (BAFF). Furthermore, we found that human FDCs expressed TLR4 and can alter antigen availability in response to pathogen-associated molecular patterns (PAMPs). High expression of PD-L1 and PD-L2 on FDCs activated PD1 on T cells. In addition, we found expression of genes related to T cell regulation, such as HLA-DRA, CD40, and others. These data suggest intimate contact between human FDCs and T cells.
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Presentación de Antígeno , Linfocitos B/inmunología , Células Dendríticas Foliculares/fisiología , Inmunidad Adaptativa , Células Presentadoras de Antígenos/inmunología , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Cadenas alfa de HLA-DR/genética , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Células Jurkat , Proteína 2 Ligando de Muerte Celular Programada 1/genética , Proteína 2 Ligando de Muerte Celular Programada 1/metabolismo , Receptor de Muerte Celular Programada 1/metabolismo , Receptores de Factores de Crecimiento/genética , Receptores de Factores de Crecimiento/metabolismo , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismoRESUMEN
In this issue of Cell Host & Microbe, Jia et al. used a vesicular stomatitis virus-based probe to isolate B cells expressing broadly neutralizing HIV-1 antibodies. Besides identifying neutralizing epitopes, this study highlights potential protection afforded by IgA arising from either direct IgM-to-IgA or sequential IgM-to-IgG-to-IgA class switching.
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VIH-1/inmunología , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo , Anticuerpos de Dominio Único , Betacoronavirus , COVID-19 , Infecciones por Coronavirus , Anticuerpos Anti-VIH , Humanos , Inmunoglobulina A , Inmunoglobulina G , Pandemias , Neumonía Viral , SARS-CoV-2RESUMEN
Humoral immune responses at mucosal surfaces have historically focused on IgA. Growing evidence highlights the complexity of IgA-inducing pathways and the functional impact of IgA on mucosal commensal bacteria. In the gut, IgA contributes to the establishment of a mutualistic host-microbiota relationship that is required to maintain homeostasis and prevent disease. This Review discusses how mucosal IgA responses occur in an increasingly complex humoral defence network that also encompasses IgM, IgG and IgD. Aside from integrating the protective functions of IgA, these hitherto neglected mucosal antibodies may strengthen the communication between mucosal and systemic immune compartments.
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Anticuerpos Antibacterianos/inmunología , Bacterias/inmunología , Inmunidad Mucosa/inmunología , Mucosa Intestinal/inmunología , Animales , Bacterias/clasificación , Humanos , Inmunoglobulina A/inmunología , Inmunoglobulina D/inmunología , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Mucosa Intestinal/microbiologíaRESUMEN
Mechanistic target of rapamycin (mTOR) enhances immunity in addition to orchestrating metabolism. Here we show that mTOR coordinates immunometabolic reconfiguration of marginal zone (MZ) B cells, a pre-activated lymphocyte subset that mounts antibody responses to T-cell-independent antigens through a Toll-like receptor (TLR)-amplified pathway involving transmembrane activator and CAML interactor (TACI). This receptor interacts with mTOR via the TLR adapter MyD88. The resulting mTOR activation instigates MZ B-cell proliferation, immunoglobulin G (IgG) class switching, and plasmablast differentiation through a rapamycin-sensitive pathway that integrates metabolic and antibody-inducing transcription programs, including NF-κB. Disruption of TACI-mTOR interaction by rapamycin, truncation of the MyD88-binding domain of TACI, or B-cell-conditional mTOR deficiency interrupts TACI signaling via NF-κB and cooperation with TLRs, thereby hampering IgG production to T-cell-independent antigens but not B-cell survival. Thus, mTOR drives innate-like antibody responses by linking proximal TACI signaling events with distal immunometabolic transcription programs.
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Linfocitos B/inmunología , Inmunoglobulina G/inmunología , Diana Mecanicista del Complejo 1 de la Rapamicina/inmunología , Factor 88 de Diferenciación Mieloide/metabolismo , Serina-Treonina Quinasas TOR/inmunología , Proteína Activadora Transmembrana y Interactiva del CAML/inmunología , Animales , Línea Celular , Proliferación Celular , Activación Enzimática , Perfilación de la Expresión Génica , Células HEK293 , Humanos , Cambio de Clase de Inmunoglobulina/genética , Cambio de Clase de Inmunoglobulina/inmunología , Inmunoglobulina G/biosíntesis , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , FN-kappa B/metabolismo , Transducción de Señal/inmunología , Sirolimus/farmacologíaRESUMEN
Secretory immunoglobulin A (SIgA) enhances host-microbiota symbiosis, whereas SIgM remains poorly understood. We found that gut IgM+ plasma cells (PCs) were more abundant in humans than mice and clonally related to a large repertoire of memory IgM+ B cells disseminated throughout the intestine but rare in systemic lymphoid organs. In addition to sharing a gut-specific gene signature with memory IgA+ B cells, memory IgM+ B cells were related to some IgA+ clonotypes and switched to IgA in response to T cell-independent or T cell-dependent signals. These signals induced abundant IgM which, together with SIgM from clonally affiliated PCs, recognized mucus-embedded commensals. Bacteria recognized by human SIgM were dually coated by SIgA and showed increased richness and diversity compared to IgA-only-coated or uncoated bacteria. Thus, SIgM may emerge from pre-existing memory rather than newly activated naive IgM+ B cells and could help SIgA to anchor highly diverse commensal communities to mucus.
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Angiodisplasia/inmunología , Linfocitos B/inmunología , Neoplasias del Colon/inmunología , Pólipos del Colon/inmunología , Inmunoglobulina M/metabolismo , Intestinos/inmunología , Células Plasmáticas/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Células Clonales , Femenino , Microbioma Gastrointestinal/inmunología , Humanos , Inmunidad Mucosa , Inmunoglobulina A/metabolismo , Cambio de Clase de Inmunoglobulina , Memoria Inmunológica , Intestinos/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , SimbiosisRESUMEN
In a recent issue of Nature, Gordon and colleagues show that, during the first 2 years life, the assembly of the gut microbiota follows predictable architectural patterns that correlate with the development of commensal-specific immunoglobulin A responses.
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Bacterias/inmunología , Microbioma Gastrointestinal/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Mucosa Intestinal/microbiología , Adulto , Interacciones Huésped-Patógeno , Humanos , Inmunoglobulina A/metabolismo , Lactante , Recién Nacido , Enfermedades Inflamatorias del Intestino/microbiología , Mucosa Intestinal/inmunología , Riesgo , SimbiosisRESUMEN
OBJECTIVE: Gain-of-function NLRP3 mutations cause cryopyrin-associated periodic syndrome (CAPS), with gene mosaicism playing a relevant role in the pathogenesis. This study was undertaken to characterize the genetic cause underlying late-onset but otherwise typical CAPS. METHODS: We studied a 64-year-old patient who presented with recurrent episodes of urticaria-like rash, fever, conjunctivitis, and oligoarthritis at age 56 years. DNA was extracted from both unfractionated blood and isolated leukocyte and CD34+ subpopulations. Genetic studies were performed using both the Sanger method of DNA sequencing and next-generation sequencing (NGS) methods. In vitro and ex vivo analyses were performed to determine the consequences that the presence of the variant have in the normal structure or function of the protein of the detected variant. RESULTS: NGS analyses revealed the novel p.Gln636Glu NLRP3 variant in unfractionated blood, with an allele frequency (18.4%) compatible with gene mosaicism. Sanger sequence chromatograms revealed a small peak corresponding to the variant allele. Amplicon-based deep sequencing revealed somatic NLRP3 mosaicism restricted to myeloid cells (31.8% in monocytes, 24.6% in neutrophils, and 11.2% in circulating CD34+ common myeloid progenitor cells) and its complete absence in lymphoid cells. Functional analyses confirmed the gain-of-function behavior of the gene variant and hyperactivity of the NLRP3 inflammasome in the patient. Treatment with anakinra resulted in good control of the disease. CONCLUSION: We identified the novel gain-of-function p.Gln636Glu NLRP3 mutation, which was detected as a somatic mutation restricted to myeloid cells, as the cause of late-onset but otherwise typical CAPS. Our results expand the diversity of CAPS toward milder phenotypes than previously reported, including those starting during adulthood.
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Síndromes Periódicos Asociados a Criopirina/genética , Enfermedades de Inicio Tardío/genética , Mosaicismo , Células Mieloides/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Alelos , Antirreumáticos/uso terapéutico , Simulación por Computador , Síndromes Periódicos Asociados a Criopirina/tratamiento farmacológico , Humanos , Proteína Antagonista del Receptor de Interleucina 1/uso terapéutico , Enfermedades de Inicio Tardío/tratamiento farmacológico , Masculino , Persona de Mediana Edad , Monocitos/metabolismo , Neutrófilos/metabolismo , Análisis de Secuencia de ADNRESUMEN
The mechanisms whereby the gut mucosa tolerates trillions of commensal bacteria without developing inflammation remain poorly understood. A recent Science article reveals that gut innate lymphoid cells constrain inflammatory T cell responses to commensal bacteria by adopting a strategy usually deployed by thymic epithelial cells to negatively select self-reactive T cells.
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Bacterias/inmunología , Linfocitos T CD4-Positivos/inmunología , Colon/microbiología , Antígenos de Histocompatibilidad Clase II/inmunología , Inmunidad Innata , Enfermedades Inflamatorias del Intestino/microbiología , Animales , Femenino , Humanos , MasculinoAsunto(s)
Enfermedades de los Nervios Craneales/genética , Mosaicismo , Mutación , Proteína Adaptadora de Señalización NOD2/genética , Sinovitis/genética , Uveítis/genética , Adolescente , Artritis , Enfermedades de los Nervios Craneales/diagnóstico , Enfermedades de los Nervios Craneales/patología , Exones , Femenino , Expresión Génica , Humanos , Linaje , Sarcoidosis , Sinovitis/diagnóstico , Sinovitis/patología , Uveítis/diagnóstico , Uveítis/patologíaRESUMEN
Innate lymphoid cells (ILCs) constitute a heterogeneous family of effector lymphocytes of the innate immune system that mediate lymphoid organogenesis, tissue repair, immunity and inflammation. The initial view that ILCs exert their protective functions solely during the innate phase of an immune response has been recently challenged by evidence indicating that ILCs shape adaptive immunity by establishing both contact-dependent and contact-independent interactions with multiple hematopoietic and non-hematopoietic cells, including B cells. Some of these interactions enhance antibody responses both systemically and at mucosal sites of entry.
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Formación de Anticuerpos , Inmunidad Innata , Subgrupos Linfocitarios/inmunología , Subgrupos Linfocitarios/metabolismo , Animales , Homeostasis , Humanos , Inmunidad , Tejido Linfoide/embriología , Tejido Linfoide/metabolismo , Membrana Mucosa/inmunología , Membrana Mucosa/metabolismo , Organogénesis , FenotipoRESUMEN
Complex and diverse communities of bacteria establish mutualistic and symbiotic relationships with the gut after birth. The intestinal immune system responds to bacterial colonization by acquiring a state of hypo-responsiveness against commensals and active readiness against pathogens. The resulting homeostatic balance involves a continuous dialog between the microbiota and lymphocytes with the intermediation of epithelial and dendritic cells. This dialog causes massive production of immunoglobulin A (IgA), a non-inflammatory antibody specialized in mucosal protection. Here, we discuss recent advances on the regulation of intestinal IgA responses and their role in host-microbe interaction.