Early Antibody-Mediated Immunity to Tuberculosis in Mice Requires NLRP3.

While antibodies have emerged as potential mediators of protective immunity against (Mtb), their mechanisms of action remain incompletely understood. Here, we demonstrate that immune complexes of Mtb and monoclonal antibodies targeting the Mtb phosphate transporter subunit PstS1 robustly activate the NLRP3 inflammasome in human and murine macrophages, leading to enhanced interleukin-1β secretion. Surprisingly, antibody-mediated inflammasome activation occurred independently of cell-surface Fcγ receptors, as confirmed using Fc-domain glycosylation mutant mAbs and macrophages from Fcγ receptor-deficient mice.

Phagosomal RNA sensing through TLR8 controls susceptibility to tuberculosis.

Genetic determinants of susceptibility to Mycobacterium tuberculosis (Mtb) remain poorly understood but could provide insights into critical pathways involved in infection, informing host-directed therapies and enabling risk stratification at individual and population levels. Through a genome-wide forward genetic screen, we identify Toll-like receptor 8 (TLR8) as a key regulator of intracellular killing of Mtb. Pharmacological TLR8 activation enhances the killing of phylogenetically diverse clinical isolates of drug-susceptible and multidrug-resistant Mtb by macrophages and during in vivo infection in mice.

Characterization of group I introns in generating circular RNAs as vaccines.

Circular RNAs are an increasingly important class of RNA molecules that can be engineered as RNA vaccines and therapeutics. Here, we screened eight different group I introns for their ability to circularize and delineated different features that are important for their function. First, we identified the Scytalidium dimidiatum group I intron as causing minimal innate immune activation inside cells, underscoring its potential to serve as an effective RNA vaccine without triggering unwanted reactogenicity.

Occlusion of TCR binding to HLA-A*11:01 by a non-pathogenic human alloantibody.

Over the last decades, organ transplantation has made rapid progress as a curative therapy for organ failure. However, the adaptive immune system-alloreactive T cells and antibodies targeting human leukocyte antigens (HLA)-is the leading cause of graft rejection. The presence of anti-donor HLA antibodies is considered a risk factor that disqualifies a particular donor-recipient pair.

Antigen-antibody complex density and antibody-induced HLA protein unfolding influence Fc-mediated antibody effector function.

Donor-specific antibodies (DSAs) targeting mismatched human leukocyte antigen (HLA) molecules are one of the principal threats to long-term graft survival in solid organ transplantation. However, many patients with long-term circulating DSAs do not manifest rejection responses, suggesting a degree of heterogeneity in their pathogenicity and related functional activity. Immunologic risk stratification of transplant recipients is complicated by challenges intrinsic to defining alloantibody responses that are potentially pathogenic versus those that are not.

Hybrid immunity augments cross-variant protection against COVID-19 among immunocompromised individuals.

Background: Immunity to SARS-CoV-2 vaccination and infection differs considerably among individuals. We investigate the critical pathways that influence vaccine-induced cross-variant serological immunity among individuals at high-risk of COVID-19 complications.

Methods: Neutralizing antibodies to the wild-type SARS-CoV-2 virus and its variants (Beta, Gamma, Delta and Omicron) were analyzed in patients with autoimmune diseases, chronic comorbidities (multimorbidity), and healthy controls.

Single-shot dendritic cell targeting SARS-CoV-2 vaccine candidate induces broad, durable and protective systemic and mucosal immunity in mice.

Current coronavirus disease 2019 vaccines face limitations including waning immunity, immune escape by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, limited cellular response, and poor mucosal immunity. We engineered a Clec9A-receptor binding domain (RBD) antibody construct that delivers the SARS-CoV-2 RBD to conventional type 1 dendritic cells. Compared with non-targeting approaches, single dose immunization in mice with Clec9A-RBD induced far higher RBD-specific antibody titers that were sustained for up to 21 months after vaccination.

Protocol for structural modeling of antibody to human leukocyte antigen interaction using discovery and targeted cross-linking mass spectrometry.

Cross-linking mass spectrometry (XL-MS) provides low-resolution structural information to model protein structures. Here, we present a protocol to identify cross-links of purified antibody binding to purified human leukocyte antigen (HLA). We describe steps for using a discovery-based XL-MS approach followed by a targeted XL-MS approach.

Type I Interferon Autoantibodies Correlate With Cellular Immune Alterations in Severe COVID-19.

Background: Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can lead to severe disease with increased morbidity and mortality among certain risk groups. The presence of autoantibodies against type I interferons (aIFN-Abs) is one mechanism that contributes to severe coronavirus disease 2019 (COVID-19).

Methods: This study aimed to investigate the presence of aIFN-Abs in relation to the soluble proteome, circulating immune cell numbers, and cellular phenotypes, as well as development of adaptive immunity.

Durable cross-protective neutralizing antibody responses elicited by lipid nanoparticle-formulated SARS-CoV-2 mRNA vaccines.

The advent of SARS-CoV-2 variants with defined mutations that augment pathogenicity and/or increase immune evasiveness continues to stimulate global efforts to improve vaccine formulation and efficacy. The extraordinary advantages of lipid nanoparticles (LNPs), including versatile design, scalability, and reproducibility, make them ideal candidates for developing next-generation mRNA vaccines against circulating SARS-CoV-2 variants. Here, we assess the efficacy of LNP-encapsulated mRNA booster vaccines encoding the spike protein of SARS-CoV-2 for variants of concern (Delta, Omicron) and using a predecessor (YN2016C isolated from bats) strain spike protein to elicit durable cross-protective neutralizing antibody responses.

Relative deficiency in interferon-γ-secreting CD4+ T cells is strongly associated with poorer COVID-19 vaccination responses in older adults.

Although the two-dose mRNA vaccination regime provides protection against SARS-CoV-2, older adults have been shown to exhibit poorer vaccination responses. In addition, the role of vaccine-induced T-cell responses is not well characterised. We aim to assess the impact of age on immune responses after two doses of the BNT162b2 mRNA vaccine, focussing on antigen-specific T-cells.

Employment of a high throughput functional assay to define the critical factors that influence vaccine induced cross-variant neutralizing antibodies for SARS-CoV-2.

The scale and duration of neutralizing antibody responses targeting SARS-CoV-2 viral variants represents a critically important serological parameter that predicts protective immunity for COVID-19. In this study, we describe the development and employment of a new functional assay that measures neutralizing antibodies for SARS-CoV-2 and present longitudinal data illustrating the impact of age, sex and comorbidities on the kinetics and strength of vaccine-induced antibody responses for key variants in an Asian volunteer cohort. We also present an accurate quantitation of serological responses for SARS-CoV-2 that exploits a unique set of in-house, recombinant human monoclonal antibodies targeting the viral Spike and nucleocapsid proteins and demonstrate a reduction in neutralizing antibody titres across all groups 6 months post-vaccination.

Defining neutralization and allostery by antibodies against COVID-19 variants.

The changing landscape of SARS-CoV-2 Spike protein is linked to the emergence of variants, immune-escape and reduced efficacy of the existing repertoire of anti-viral antibodies. The functional activity of neutralizing antibodies is linked to their quaternary changes occurring as a result of antibody-Spike trimer interactions. Here, we reveal the conformational dynamics and allosteric perturbations linked to binding of novel human antibodies and the viral Spike protein.

Analyzing COVID-19 Vaccine Responses in Transplant Recipients.

COVID-19 vaccination has significantly impacted the global pandemic by reducing the severity of infection, lowering rates of hospitalization, and reducing morbidity/mortality in healthy individuals. However, the degree of vaccine-induced protection afforded to renal transplant recipients who receive forms of maintenance immunosuppression remains poorly defined. This is particularly important when we factor in the emergence of SARS-CoV-2 variants of concern (VOCs) that have defined mutations that reduce the effectiveness of Ab responses targeting the Spike Ags from the ancestral Wuhan-Hu-1 variants employed in the most widely used vaccine formats.

Hybrid structural modeling of alloantibody binding to human leukocyte antigen with rapid and reproducible cross-linking mass spectrometry.

Alloantibody recognition of donor human leukocyte antigen (HLA) is associated with poor clinical transplantation outcomes. However, the molecular and structural basis for the alloantibody-HLA interaction is not well understood. Here, we used a hybrid structural modeling approach on a previously studied alloantibody-HLA interacting pair with inputs from ab initio, in silico, and in vitro data.

Platelet TLR7 is essential for the formation of platelet-neutrophil complexes and low-density neutrophils in lupus nephritis.

Objectives: Platelets and low-density neutrophils (LDNs) are major players in the immunopathogenesis of SLE. Despite evidence showing the importance of platelet-neutrophil complexes (PNCs) in inflammation, little is known about the relationship between LDNs and platelets in SLE. We sought to characterize the role of LDNs and Toll-like receptor 7 (TLR7) in clinical disease.

CD28-CAR-T cell activation through FYN kinase signaling rather than LCK enhances therapeutic performance.

Signal transduction induced by chimeric antigen receptors (CARs) is generally believed to rely on the activity of the SRC family kinase (SFK) LCK, as is the case with T cell receptor (TCR) signaling. Here, we show that CAR signaling occurs in the absence of LCK. This LCK-independent signaling requires the related SFK FYN and a CD28 intracellular domain within the CAR.

Switching Heavy Chain Constant Domains Denatures the Paratope 3D Architecture of Influenza Monoclonal Antibodies.

Several human monoclonal Abs for treating Influenza have been evaluated in clinical trials with limited success despite demonstrating superiority in preclinical animal models including mice. To conduct efficacy studies in mice, human monoclonal Abs are genetically engineered to contain mouse heavy chain constant domain to facilitate the engagement of Fc-receptors on mouse immune effector cells. Although studies have consistently reported discrepancies in Ab effectiveness following genetic engineering, the structural and mechanistic basis for these inconsistencies remain uncharacterized.

SARS-CoV-2 Antibody Effectiveness Is Influenced by Non-Epitope Mutation/Binding-Induced Denaturation of the Epitope 3D Architecture.

The public health threat from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to intensify with emerging variants of concern (VOC) aiming to render COVID-19 vaccines/infection-induced antibodies redundant. The SARS-CoV-2 spike protein is responsible for receptor binding and infection of host cells making it a legitimate antibody target. Antibodies mostly target epitopes in the receptor binding domain (RBD).

Efficient recall of SARS-CoV-2 variant-reactive B cells and T responses in the elderly upon heterologous mRNA vaccines as boosters.

Waning antibody levels against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the emergence of variants of concern highlight the need for booster vaccinations. This is particularly important for the elderly population, who are at a higher risk of developing severe coronavirus disease 2019 (COVID-19) disease. While studies have shown increased antibody responses following booster vaccination, understanding the changes in T and B cell compartments induced by a third vaccine dose remains limited.

Defining the specificity and function of a human neutralizing antibody for Hepatitis B virus.

Hepatitis B Virus (HBV) is a hepadnavirus that is the principal pathogen underlying viral liver disease in human populations. In this study, we describe the isolation and characterization of a fully human monoclonal antibody for HBV. This HuMab was isolated by a combinatorial screen of the memory B-cell repertoire from an acute/recovered HBV-infected patient.