Structural basis for the inhibition of βFXIIa by garadacimab.

Activated FXII (FXIIa) is the principal initiator of the plasma contact system and can activate both procoagulant and proinflammatory pathways. Its activity is important in the pathophysiology of hereditary angioedema (HAE). Here, we describe a high-resolution cryoelectron microscopy (cryo-EM) structure of the beta-chain from FXIIa (βFXIIa) complexed with the Fab fragment of garadacimab.

Inhibition of VEGF-B signaling prevents non-alcoholic fatty liver disease development by targeting lipolysis in the white adipose tissue.

Background & Aims: Hepatic steatosis is a hallmark of non-alcoholic fatty liver disease (NAFLD), a common comorbidity in type 2 diabetes mellitus (T2DM). The pathogenesis of NAFLD is complex and involves the crosstalk between the liver and the white adipose tissue (WAT). Vascular endothelial growth factor B (VEGF-B) has been shown to control tissue lipid accumulation by regulating the transport properties of the vasculature.

HDX-MS study on garadacimab binding to activated FXII reveals potential binding interfaces through differential solvent exposure.

Hageman factor (FXII) is an essential component in the intrinsic coagulation cascade and a therapeutic target for the prophylactic treatment of hereditary angioedema (HAE). CSL312 (garadacimab) is a novel high-affinity human antibody capable of blocking activated FXII activity that is currently undergoing Phase 3 clinical trials in HAE. Structural studies using hydrogen/deuterium exchange coupled to mass spectrometry revealed evidence of interaction between the antibody and regions surrounding the S1 specificity pocket of FXII, including the 99-loop, 140-loop, 180-loop, and neighboring regions.

Butyrophilin 2A1 is essential for phosphoantigen reactivity by γδ T cells.

Gamma delta (γδ) T cells are essential to protective immunity. In humans, most γδ T cells express Vγ9Vδ2 T cell receptors (TCRs) that respond to phosphoantigens (pAgs) produced by cellular pathogens and overexpressed by cancers. However, the molecular targets recognized by these γδTCRs are unknown.

A potential association between IL-3 and type I and III interferons in systemic lupus erythematosus.

Objectives: Plasmacytoid dendritic cells (pDCs), through the production of type 1 interferons (IFNs) and other cytokines, are major contributors to systemic lupus erythematosus (SLE) pathogenesis. IL-3 promotes pDC survival, but its role in SLE is not well characterised. This study investigated serum IL-3 and IFN levels, and a whole blood 'IL-3 gene signature', in human SLE.

Lack of Fractalkine Receptor on Macrophages Impairs Spontaneous Recovery of Ribbon Synapses After Moderate Noise Trauma in C57BL/6 Mice.

Noise trauma causes loss of synaptic connections between cochlear inner hair cells (IHCs) and the spiral ganglion neurons (SGNs). Such synaptic loss can trigger slow and progressive degeneration of SGNs. Macrophage fractalkine signaling is critical for neuron survival in the injured cochlea, but its role in cochlear synaptopathy is unknown.

A therapeutic gingipain vaccine induces neutralising IgG1 antibodies that protect against experimental periodontitis.

infected mice with an established -specific inflammatory immune response were protected from developing alveolar bone resorption by therapeutic vaccination with a chimera (KAS2-A1) immunogen targeting the major virulence factors of the bacterium, the gingipain proteinases. Protection was characterised by an antigen-specific IgG1 isotype antibody and Th2 cell response. Adoptive transfer of KAS2-A1-specific IgG1 or IgG2 expressing B cells confirmed that IgG1-mediated protection.

Role of the β Common (βc) Family of Cytokines in Health and Disease.

The β common ([βc]/CD131) family of cytokines comprises granulocyte macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-3, and IL-5, all of which use βc as their key signaling receptor subunit. This is a prototypic signaling subunit-sharing cytokine family that has unveiled many biological paradigms and structural principles applicable to the IL-2, IL-4, and IL-6 receptor families, all of which also share one or more signaling subunits. Originally identified for their functions in the hematopoietic system, the βc cytokines are now known to be truly pleiotropic, impacting on multiple cell types, organs, and biological systems, and thereby controlling the balance between health and disease.

An Anti-VEGF-B Antibody Fragment Induces Regression of Pre-Existing Blood Vessels in the Rat Cornea.

Purpose: We tested the ability of an antibody fragment with specificity for vascular endothelial growth factor-B (VEGF-B) to regress nascent and established corneal blood vessels in the rat.

Methods: A single chain variable antibody fragment (scFv) with specificity for VEGF-B was engineered from the 2H10 hybridoma. Binding to rat, mouse, and human VEGF-B was confirmed by surface plasmon resonance.

A neutralizing anti-G-CSFR antibody blocks G-CSF-induced neutrophilia without inducing neutropenia in nonhuman primates.

Neutrophils are the most abundant WBCs and have an essential role in the clearance of pathogens. Tight regulation of neutrophil numbers and their recruitment to sites of inflammation is critical in maintaining a balanced immune response. In various inflammatory conditions, such as rheumatoid arthritis, vasculitis, cystic fibrosis, and inflammatory bowel disease, increased serum G-CSF correlates with neutrophilia and enhanced neutrophil infiltration into inflamed tissues.

Reducing VEGF-B Signaling Ameliorates Renal Lipotoxicity and Protects against Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is the most common cause of severe renal disease, and few treatment options are available today that prevent the progressive loss of renal function. DKD is characterized by altered glomerular filtration and proteinuria. A common observation in DKD is the presence of renal steatosis, but the mechanism(s) underlying this observation and to what extent they contribute to disease progression are unknown.

Therapeutic Targeting of the G-CSF Receptor Reduces Neutrophil Trafficking and Joint Inflammation in Antibody-Mediated Inflammatory Arthritis.

G-CSF is a hemopoietic growth factor that has a role in steady state granulopoiesis, as well as in mature neutrophil activation and function. G-CSF- and G-CSF receptor-deficient mice are profoundly protected in several models of rheumatoid arthritis, and Ab blockade of G-CSF also protects against disease. To further investigate the actions of blocking G-CSF/G-CSF receptor signaling in inflammatory disease, and as a prelude to human studies of the same approach, we developed a neutralizing mAb to the murine G-CSF receptor, which potently antagonizes binding of murine G-CSF and thereby inhibits STAT3 phosphorylation and G-CSF receptor signaling.

A cytotoxic anti-IL-3Rα antibody targets key cells and cytokines implicated in systemic lupus erythematosus.

To date, the major target of biologic therapeutics in systemic lupus erythematosus (SLE) has been the B cell, which produces pathogenic autoantibodies. Recently, targeting type I IFN, which is elaborated by plasmacytoid dendritic cells (pDCs) in response to endosomal TLR7 and TLR9 stimulation by SLE immune complexes, has shown promising results. pDCs express high levels of the IL-3Rα chain (CD123), suggesting an alternative potential targeting strategy.

Species Cross-Reactivity of Antibodies Used to Treat Ophthalmic Conditions.

Purpose: The species cross-reactivity of the monoclonal antibodies infliximab, bevacizumab, and an anti-VEGF-B antibody, 2H10, in humans and rodents was determined.

Methods: The binding of infliximab to human, mouse, and rat TNF-α, of bevacizumab to human, mouse, and rat VEGF-A, and of the 2H10 antibody to human, mouse, and rat VEGF-B was evaluated by ELISA. The sequence of human, mouse, and rat TNF-α and VEGF-A at the binding sites for infliximab and bevacizumab were compared.

CSL311, a novel, potent, therapeutic monoclonal antibody for the treatment of diseases mediated by the common β chain of the IL-3, GM-CSF and IL-5 receptors.

The β common-signaling cytokines interleukin (IL)-3, granulocyte-macrophage colony stimulating factor (GM-CSF) and IL-5 stimulate pro-inflammatory activities of haematopoietic cells via a receptor complex incorporating cytokine-specific α and shared β common (βc, CD131) receptor. Evidence from animal models and recent clinical trials demonstrate that these cytokines are critical mediators of the pathogenesis of inflammatory airway disease such as asthma. However, no therapeutic agents, other than steroids, that specifically and effectively target inflammation mediated by all 3 of these cytokines exist.

Prospects for treatment of Porphyromonas gingivalis-mediated disease - immune-based therapy.

Chronic periodontitis is an inflammatory disease of the supporting tissues of the teeth associated with a polymicrobial biofilm (subgingival plaque) accreted to the tooth which results in destruction of the tooth's supporting tissues. A characteristic feature of the disease-associated plaque is the emergence of proteolytic species. One of these species, Porphyromonas gingivalis has recently been described as a keystone pathogen as it dysregulates the host immune response to favour the polymicrobial biofilm disrupting homeostasis to cause dysbiosis and disease.

Dual mechanism of interleukin-3 receptor blockade by an anti-cancer antibody.

Interleukin-3 (IL-3) is an activated T cell product that bridges innate and adaptive immunity and contributes to several immunopathologies. Here, we report the crystal structure of the IL-3 receptor α chain (IL3Rα) in complex with the anti-leukemia antibody CSL362 that reveals the N-terminal domain (NTD), a domain also present in the granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-5, and IL-13 receptors, adopting unique "open" and classical "closed" conformations. Although extensive mutational analyses of the NTD epitope of CSL362 show minor overlap with the IL-3 binding site, CSL362 only inhibits IL-3 binding to the closed conformation, indicating alternative mechanisms for blocking IL-3 signaling.

Crystallization and preliminary X-ray diffraction analysis of the interleukin-3 alpha receptor bound to the Fab fragment of antibody CSL362.

Interleukin-3 (IL-3) is a member of the beta common family of cytokines that regulate multiple functions of myeloid cells. The IL-3 receptor-specific alpha subunit (IL3Rα) is overexpressed on stem cells/progenitor cells of patients with acute myeloid leukaemia, where elevated receptor expression correlates clinically with a reduced patient survival rate. The monoclonal antibody (MAb) CSL362 is a humanized MAb derived from the murine MAb 7G3, originally identified for its ability to specifically recognize the human IL-3 receptor and for blocking the signalling of IL-3 in myeloid and endothelial cells.

A factor XIIa inhibitory antibody provides thromboprotection in extracorporeal circulation without increasing bleeding risk.

Currently used anticoagulants prevent thrombosis but increase bleeding. We show an anticoagulation therapy without bleeding risk based on a plasma protease factor XII function-neutralizing antibody. We screened for antibodies against activated factor XII (FXIIa) using phage display and demonstrated that recombinant fully human antibody 3F7 binds into the FXIIa enzymatic pocket.

Interleukin-11 is the dominant IL-6 family cytokine during gastrointestinal tumorigenesis and can be targeted therapeutically.

Among the cytokines linked to inflammation-associated cancer, interleukin (IL)-6 drives many of the cancer "hallmarks" through downstream activation of the gp130/STAT3 signaling pathway. However, we show that the related cytokine IL-11 has a stronger correlation with elevated STAT3 activation in human gastrointestinal cancers. Using genetic mouse models, we reveal that IL-11 has a more prominent role compared to IL-6 during the progression of sporadic and inflammation-associated colon and gastric cancers.

Production of a human neutralizing monoclonal antibody and its crystal structure in complex with ectodomain 3 of the interleukin-13 receptor α1.

Gene deletion studies in mice have revealed critical roles for IL (interleukin)-4 and -13 in asthma development, with the latter controlling lung airways resistance and mucus secretion. We have now developed human neutralizing monoclonal antibodies against human IL-13Rα1 (IL-13 receptor α1) subunit that prevent activation of the receptor complex by both IL-4 and IL-13. We describe the crystal structures of the Fab fragment of antibody 10G5H6 alone and in complex with D3 (ectodomain 3) of IL-13Rα1.

Targeting VEGF-B as a novel treatment for insulin resistance and type 2 diabetes.

The prevalence of type 2 diabetes is rapidly increasing, with severe socioeconomic impacts. Excess lipid deposition in peripheral tissues impairs insulin sensitivity and glucose uptake, and has been proposed to contribute to the pathology of type 2 diabetes. However, few treatment options exist that directly target ectopic lipid accumulation.

VEGF-B is dispensable for blood vessel growth but critical for their survival, and VEGF-B targeting inhibits pathological angiogenesis.

VEGF-B, a homolog of VEGF discovered a long time ago, has not been considered an important target in antiangiogenic therapy. Instead, it has received little attention from the field. In this study, using different animal models and multiple types of vascular cells, we revealed that although VEGF-B is dispensable for blood vessel growth, it is critical for their survival.