61 research outputs found

    Modified recombinant human IgG1-Fc is superior to natural IVIG at inhibiting immune-mediated demyelination.

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    Intravenous immunoglobulin (IVIG) is an established treatment for numerous autoimmune conditions. Although Fc fragments derived from IVIG have shown efficacy in controlling immune thrombocytopenia (ITP) in children, the mechanisms of action are unclear and controversial. The aim of this study is to dissect IVIG effector mechanisms using further adapted Fc fragments on demyelination in an ex vivo model of the central nervous system (CNS)-immune interface. Using organotypic cerebellar slice cultures (OSC) from transgenic mice we induced extensive immune-mediated demyelination and oligodendrocyte loss with an antibody specific for myelin oligodendrocyte glycoprotein (MOG) and complement. Protective effects of adapted Fc fragments were assessed by live imaging of GFP expression, immunohistochemistry and confocal microscopy. Cysteine and glycan adapted Fc fragments protected OSC from demyelination in a dose-dependent manner where equimolar concentrations of either IVIG or control Fc were ineffective. The protective effects of the adapted Fc fragments are partly attributed to interference with complement-mediated oligodendroglia damage. Transcriptome analysis ruled out signatures associated with inflammatory or innate immune responses. Taken together our findings show that recombinant biomimetics can be made that are at least two hundred-fold more effective than IVIG in controlling demyelination by anti-MOG antibodies

    TITLE PAGE Small-Molecule Vasopressin-2 Receptor Antagonist Identified by a G- Protein Coupled Receptor 'Pathway' Screen MOL #34496 2 RUNNING TITLE PAGE Running title: Small-molecule V 2 R antagonist Corresponding author: Number of text pages: 31 Number o

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    Number of references: 40 Number of words in Abstract: 200 Number of words in Introduction: 496 Number of words in Discussion: 1,102 List of abbreviations: GPCR, G-protein coupled receptor; cAMP, cyclic adenosine monophospate; CFTR

    Cross-reactivity of mouse IgG subclasses to human Fc gamma receptors: Antibody deglycosylation only eliminates IgG2b binding

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    Immunoglobulin G (IgG) antibodies are important for protection against pathogens and exert effector functions through binding to IgG-Fc receptors (FcγRs) on myeloid and natural killer cells, resulting in destruction of opsonized target cells. Despite interspecies differences, IgG subclasses and FcγRs show substantial similarities and functional conservation between mammals. Accordingly, binding of human IgG (hIgG) to mouse FcγRs (mFcγRs) has been utilized to study effector functions of hIgG in mice. In other applications, such as immunostaining with mouse IgG monoclonal antibodies (mAbs), these cross-reactivities are undesired and prone to misinterpretation. Despite this drawback, the binding of mouse IgG (mIgG) subclasses to human FcγR (hFcγR) classes has never been fully documented. Here, we report detailed and quantifiable characterization of binding affinities for all mIgG subclasses to hFcγRs, including functional polymorphic variants. mIgG subclasses show the strongest binding to hFcγRIa, with relative affinities mIgG2a = mIgG2c > mIgG3 >> mIgG2b, and no binding by mIgG1. hFcγRIIa/b showed general low reactivities to all mIgG (mIgG1> mIgG2a/c > mIgG2b), with no reactivity to mIgG3. A particularly high affinity was observed for mIgG1 to the hFcγRIIa-R131 polymorphic variant. hFcγRIIIa showed lower binding (mIgG2a/c > mIgG3), slightly favouring binding to the hFcγRIIIa-V158 over the F158 polymorphic variant. No binding was observed of mIgG to hFcγRIIIb. Deglycosylation of mIgG1 did not abrogate binding to hFcγRIIa-R131, nor did deglycosylation of mIgG2a/c and mIgG3 prevent hFcγRIa binding. Importantly, deglycosylation of the least cross-reactive mIgG subclass, mIgG2b, abrogated reactivity to all hFcγRs. Together, these data document for the first time the full spectrum of cross-reactivities of mouse IgG to human FcγRs

    Affinity-matured ‘aquaporumab’ anti-aquaporin-4 antibody for therapy of seropositive neuromyelitis optica spectrum disorders

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    Pathogenesis in seropositive neuromyelitis optica spectrum disorders (herein called NMO) involves binding of IgG1 autoantibodies to aquaporin-4 (AQP4) on astrocytes in the central nervous system, which initiates complement and cellular injury. We previously developed an antibody blocking approach for potential therapy of NMO in which an engineered, monoclonal, anti-AQP4 antibody lacking cytotoxicity effector functions (called aquaporumab) blocked binding of NMO autoantibodies to astrocyte AQP4 (Tradtrantip et al. Ann. Neurol. 71, 314-322, 2012). Here, a high-affinity aquaporumab, which was generated by affinity maturation using saturation mutagenesis, was shown to block cellular injury caused by NMO patient sera. Anti-AQP4 antibody rAb-53, a fully human antibody with effector function neutralizing Fc mutations L234A/L235A and affinity-enhancing Fab mutations Y50R/S56R, called AQmabAM, bound to AQP4 in cell cultures with Kd ~ 18 ng/ml (~0.12 nM), ~8-fold greater affinity than the original antibody. AQmabAM, but without L234A/L235A Fc mutations, produced complement-dependent cytotoxicity (CDC) with EC50 ~ 82 ng/ml. AQmabAM prevented CDC produced by sera from eight NMO patients with IC50 ranging from 40 to 80 ng/ml, and similarly prevented antibody-dependent cellular cytotoxicity (ADCC). Mechanistic studies demonstrated that AQmabAM blocked binding of serum NMO autoantibodies to AQP4. AQmabAM offers a targeted, non-immunosuppressive approach for therapy of seropositive NMO. Autoantibody blocking may be a useful therapeutic strategy for other autoimmune diseases as well

    Designing a broad-spectrum integrative approach for cancer prevention and treatment.

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    Targeted therapies and the consequent adoption of "personalized" oncology have achieved notable successes in some cancers; however, significant problems remain with this approach. Many targeted therapies are highly toxic, costs are extremely high, and most patients experience relapse after a few disease-free months. Relapses arise from genetic heterogeneity in tumors, which harbor therapy-resistant immortalized cells that have adopted alternate and compensatory pathways (i.e., pathways that are not reliant upon the same mechanisms as those which have been targeted). To address these limitations, an international task force of 180 scientists was assembled to explore the concept of a low-toxicity "broad-spectrum" therapeutic approach that could simultaneously target many key pathways and mechanisms. Using cancer hallmark phenotypes and the tumor microenvironment to account for the various aspects of relevant cancer biology, interdisciplinary teams reviewed each hallmark area and nominated a wide range of high-priority targets (74 in total) that could be modified to improve patient outcomes. For these targets, corresponding low-toxicity therapeutic approaches were then suggested, many of which were phytochemicals. Proposed actions on each target and all of the approaches were further reviewed for known effects on other hallmark areas and the tumor microenvironment. Potential contrary or procarcinogenic effects were found for 3.9% of the relationships between targets and hallmarks, and mixed evidence of complementary and contrary relationships was found for 7.1%. Approximately 67% of the relationships revealed potentially complementary effects, and the remainder had no known relationship. Among the approaches, 1.1% had contrary, 2.8% had mixed and 62.1% had complementary relationships. These results suggest that a broad-spectrum approach should be feasible from a safety standpoint. This novel approach has potential to be relatively inexpensive, it should help us address stages and types of cancer that lack conventional treatment, and it may reduce relapse risks. A proposed agenda for future research is offered.Multiple funders. See acknowledgments within article for details.This is the final version of the article. It first appeared from Elsevier via http://dx.doi.org/10.1016/j.semcancer.2015.09.00

    Complete Removal of Extracellular IgG Antibodies in a Randomized Dose-Escalation Phase I Study with the Bacterial Enzyme IdeS--A Novel Therapeutic Opportunity.

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    IdeS is a streptococcal protease that cleaves IgG antibodies into F(ab’)2 and Fc fragments with a unique degree of specificity, thereby providing a novel treatment opportunity of IgG-driven autoimmune conditions and antibody mediated transplant rejection. Here we report the results from a first in man, double blinded and randomized study with single ascending doses of IdeS in healthy, male subjects. Twenty healthy subjects were given intravenous single ascending doses of IdeS. With impressive efficacy IdeS cleaved the entire plasma IgG-pool only minutes after dosing. IgG reached nadir 6-24 hours after dosing and then slowly recovered. The half-life of IdeS was 4.9 (±2.8) hours at 0.24 mg/kg with the main fraction eliminated during 24 hours. Already two hours after IdeS-dosing, the phagocytic capacity of IgG/IgG-fragments was reduced to background levels. Importantly, IdeS has the capacity to inactivate Fc-mediated effector function in vivo, was considered safe with no serious adverse events, and without dose limiting toxicity in this study. The complete, rapid, but temporary removal of IgG provides a new potent therapeutic opportunity in IgG-mediated pathogenic conditions.ClinicalTrials.gov NCT01802697

    Treatment of neuromyelitis optica: state-of-the-art and emerging therapies.

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    Neuromyelitis optica (NMO) is an autoimmune disease of the CNS that is characterized by inflammatory demyelinating lesions in the spinal cord and optic nerve, potentially leading to paralysis and blindness. NMO can usually be distinguished from multiple sclerosis (MS) on the basis of seropositivity for IgG antibodies against the astrocytic water channel aquaporin-4 (AQP4). Differentiation from MS is crucial, because some MS treatments can exacerbate NMO. NMO pathogenesis involves AQP4-IgG antibody binding to astrocytic AQP4, which causes complement-dependent cytotoxicity and secondary inflammation with granulocyte and macrophage infiltration, blood-brain barrier disruption and oligodendrocyte injury. Current NMO treatments include general immunosuppressive agents, B-cell depletion, and plasma exchange. Therapeutic strategies targeting complement proteins, the IL-6 receptor, neutrophils, eosinophils and CD19--all initially developed for other indications--are under clinical evaluation for repurposing for NMO. Therapies in the preclinical phase include AQP4-blocking antibodies and AQP4-IgG enzymatic inactivation. Additional, albeit currently theoretical, treatment options include reduction of AQP4 expression, disruption of AQP4 orthogonal arrays, enhancement of complement inhibitor expression, restoration of the blood-brain barrier, and induction of immune tolerance. Despite the many therapeutic options in NMO, no controlled clinical trials in patients with this condition have been conducted to date
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