Agglutinating mouse IgG3 compares favourably with IgMs in typing of the blood group B antigen: Functionality and stability studies

Klaus, Tomasz; Bzowska, Monika; Kulesza, Małgorzata; Kabat, Agnieszka M.; Jemioła-Rzemińska, Małgorzata; Czaplicki, Dominik; Makuch, Krzysztof; Jucha, Jarosław; Karabasz, Alicja; Bereta, Joanna

doi:10.1038/srep30938

Cited by 11 publications

(38 citation statements)

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“…Next, we aimed at elucidating the mechanism of truncation of the IgM heavy chain. We assumed that the cleavage is catalyzed by a serum protease 7 and, to determine its specificity, we identified the residues within P6-P4′ sequence essential for trimming (Fig. 2a ).…”

Section: Resultsmentioning

confidence: 99%

“…High Mw, heavy glycosylation, dozens of disulfide bonds and a complex oligomeric structure make IgMs very difficult to express, purify and formulate. IgMs activity is rapidly lost through aggregation that is predominantly disulfide driven 7 . In the case of mouse IgMs we observed additional phenomenon adversely affecting their activity: the heavy chain N-terminus trimming, which yields 55 kDa μ’ chain 7 .…”

Section: Introductionmentioning

confidence: 99%

“…We have previously reported that the quantity of μ’ chain increases during storage of IgM-based blood typing reagents 7 . Our results indicated that μ’ chain is generated extracellularly from full-length IgM in the presence of serum or in the presence of bovine serum albumin (BSA) preparation, which, apart from albumin, contains numerous other proteins.…”

Section: Introductionmentioning

confidence: 99%

“…We have observed that the accumulation of μ’ chain adversely affected the activity of the IgM reagents. Substantial amount of μ’ chain was detected also in normal mouse serum 7 , 8 , but its influence on immune response and IgM activity in vivo is unknown.…”

Section: Introductionmentioning

confidence: 99%

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Mouse Antibody of IgM Class is Prone to Non-Enzymatic Cleavage between CH1 and CH2 Domains

Klaus

Stalińska

Czaplicki

et al. 2018

Sci Rep

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IgM is a multivalent antibody which evolved as a first line defense of adaptive immunity. It consists of heavy and light chains assembled into a complex oligomer. In mouse serum there are two forms of IgM, a full-length and a truncated one. The latter contains μ’ chain, which lacks a variable region. Although μ’ chain was discovered many years ago, its origin has not yet been elucidated. Our results indicate that μ’ chain is generated from a full-length heavy chain by non-enzymatic cleavage of the protein backbone. The cleavage occurred specifically after Asn209 and is prevented by mutating this residue into any other amino acid. The process requires the presence of other proteins, preferentially with an acidic isoelectric point, and is facilitated by neutral or alkaline pH. This unique characteristic of the investigated phenomenon distinguishes it from other, already described, Asn-dependent protein reactions. A single IgM molecule is able to bind up to 12 epitopes via its antigen binding fragments (Fabs). The cleavage at Asn209 generates truncated IgM molecules and free Fabs, resulting in a reduced IgM valence and probably affecting IgM functionality in vivo.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mouse Antibody of IgM Class is Prone to Non-Enzymatic Cleavage between CH1 and CH2 Domains

Klaus

Stalińska

Czaplicki

et al. 2018

Sci Rep

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“…(35,41). This difference may be due to greater contribution of the mIgG 3 hinge region or disulfide bonds to K. pneumoniae CPS epitopes (42), and further digestion of the antibody into Fab fragments or replacement of heavy chain domains may shed more light on these interactions (8). Nonetheless, aggregation and cooperative binding appear to be important in defending against encapsulated organisms, since in addition to mIgG 3 and cold agglutinin IgM, the T-independent subclass hIgG 2 has also demonstrated the ability to self-aggregate (44).…”

Section: Discussionmentioning

confidence: 99%

The Role of IgG Subclass in Antibody-Mediated Protection against Carbapenem-ResistantKlebsiella pneumoniae

Motley

Diago-Navarro

Banerjee

et al. 2020

Preprint

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Monoclonal antibodies (Abs) have the potential to assist in the battle against multidrug-resistant bacteria such as Carbapenem-Resistant Klebsiella pneumoniae (CR-Kp). However, the characteristics by which these Abs function, such as the role of antibody subclass, must be determined before such modalities can be carried from the bench to the bedside. We performed a subclass switch on anti-capsular monoclonal murine IgG3 (mIgG3) hybridomas and identified and purified a murine IgG1 (mIgG1) hybridoma line through sib selection. We then compared the ability of the mIgG1 and mIgG3 antibodies to control CR-Kp ST258 infection both in vitro and in vivo. We found by ELISA and flow cytometry that mIgG3 has superior binding to CR-Kp CPS and superior agglutinating ability compared to mIgG1. The mIgG3 also predictably had better complement-mediated serum bactericidal activity than the mIgG1 and also promoted neutrophil-mediated killing at concentrations lower than the mIgG1. In contrast, the mIgG1 had marginally better activity in improving macrophage-mediated phagocytosis. Comparing their activities in a pulmonary infection model with wild type as well as neutropenic mice, both antibodies reduced organ burden in a non-lethal challenge, regardless of neutrophil status, with mIgG1 having the highest overall burden reduction in both scenarios. However, at a lethal inoculum, both antibodies showed reduced efficacy in neutropenic mice, with mIgG3 retaining the most activity. These findings suggest the viability of monoclonal Ab adjunctive therapy in neutropenic patients that cannot mount their own immune response, while also providing some insight into the relative contributions of immune mediators in CR-Kp protection.ImportanceCarbapenem-resistant Klebsiella pneumoniae is an urgent public health threat that causes life-threatening infections in immunocompromised hosts. Its resistance to nearly all antibiotics necessitates novel strategies to treat it, including the use of monoclonal antibodies. Monoclonal antibodies are emerging as important adjuncts to traditional pharmaceuticals, and studying how they protect against specific bacteria such as Klebsiella pneumoniae is crucial to their development as effective therapies. Antibody subclass is often overlooked but is a major factor in how an antibody interacts with other mediators of immunity. This paper is the first to examine how the subclass of anti-capsular monoclonal antibodies can affect efficacy against CR-Kp. Additionally, this work sheds light on the viability of monoclonal antibody therapy in neutropenic patients, who are most vulnerable to CR-Kp infection.

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Targeting BMI‐1 to deplete antibody‐secreting cells in autoimmunity

Polmear,

Hailes,

Olshansky

et al. 2023

Clin & Trans Imm

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ObjectivesB cells drive the production of autoreactive antibody‐secreting cells (ASCs) in autoimmune diseases such as Systemic Lupus Erythematosus (SLE) and Sjögren's syndrome, causing long‐term organ damage. Current treatments for antibody‐mediated autoimmune diseases target B cells or broadly suppress the immune system. However, pre‐existing long‐lived ASCs are often refractory to treatment, leaving a reservoir of autoreactive cells that continue to produce antibodies. Therefore, the development of novel treatment methods targeting ASCs is vital to improve patient outcomes. Our objective was to test whether targeting the epigenetic regulator BMI‐1 could deplete ASCs in autoimmune conditions in vivo and in vitro.MethodsUse of a BMI‐1 inhibitor in both mouse and human autoimmune settings was investigated. Lyn−/− mice, a model of SLE, were treated with the BMI‐1 small molecule inhibitor PTC‐028, before assessment of ASCs, serum antibody and immune complexes. To examine human ASC survival, a novel human fibroblast‐based assay was established, and the impact of PTC‐028 on ASCs derived from Sjögren's syndrome patients was evaluated.ResultsBMI‐1 inhibition significantly decreased splenic and bone marrow ASCs in Lyn−/− mice. The decline in ASCs was linked to aberrant cell cycle gene expression and led to a significant decrease in serum IgG3, immune complexes and anti‐DNA IgG. PTC‐028 was also efficacious in reducing ex vivo plasma cell survival from both Sjögren's syndrome patients and age‐matched healthy donors.ConclusionThese data provide evidence that inhibiting BMI‐1 can deplete ASC in a variety of contexts and thus BMI‐1 is a viable therapeutic target for antibody‐mediated autoimmune diseases.

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Agglutinating mouse IgG3 compares favourably with IgMs in typing of the blood group B antigen: Functionality and stability studies

Cited by 11 publications

References 65 publications

Mouse Antibody of IgM Class is Prone to Non-Enzymatic Cleavage between CH1 and CH2 Domains

Mouse Antibody of IgM Class is Prone to Non-Enzymatic Cleavage between CH1 and CH2 Domains

The Role of IgG Subclass in Antibody-Mediated Protection against Carbapenem-ResistantKlebsiella pneumoniae

Targeting BMI‐1 to deplete antibody‐secreting cells in autoimmunity

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