Effect of Individual Fc Methionine Oxidation on FcRn Binding: Met252 Oxidation Impairs FcRn Binding More Profoundly than Met428 Oxidation

Gao, Xuan; Ji, Junyan A.; Veeravalli, Karthik; Wang, Y. John; Zhang, Taylor; Mcgreevy, William; Zheng, Kai; Kelley, Robert F.; Laird, Michael W.; Li, Jun; Cromwell, Mary

doi:10.1002/jps.24136

Cited by 91 publications

(76 citation statements)

References 30 publications

(90 reference statements)

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“…Although a Met amino acid substitution at residue 311 was found to be beneficial, it was not included in any final designs due to the possibility of in vivo oxidation. 35,36 Residues M252, I253 and S254, which are involved in FcRn binding, are part of the 250-helix which exhibits conformational flexibility at various pHs. We hypothesized that reduced conformational flexibility would lead to better exposure of these residues and improve the shape complementarity between Fc and FcRn.…”

Section: Resultsmentioning

confidence: 99%

Extending human IgG half-life using structure-guided design

Booth

Ramakrishnan

Narayan

et al. 2018

mAbs

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Engineering of antibodies for improved pharmacokinetics through enhanced binding to the neonatal Fc receptor (FcRn) has been demonstrated in transgenic mice, non-human primates and humans. Traditionally, such approaches have largely relied on random mutagenesis and display formats, which fail to address related critical attributes of the antibody, such as effector functions or biophysical stability. We have developed a structure- and network-based framework to interrogate the engagement of IgG with multiple Fc receptors (FcRn, C1q, TRIM21, FcγRI, FcγRIIa/b, FcγRIIIa) simultaneously. Using this framework, we identified features that govern Fc-FcRn interactions and identified multiple distinct pathways for enhancing FcRn binding in a pH-specific manner. Network analysis provided a novel lens to study the allosteric impact of half-life-enhancing Fc mutations on FcγR engagement, which occurs distal to the FcRn binding site. Applying these principles, we engineered a panel of unique Fc variants that enhance FcRn binding while maintaining robust biophysical properties and wild type-like binding to activating receptors. An antibody harboring representative Fc designs demonstrates a half-life improvement of > 9 fold in transgenic mice and > 3.5 fold in cynomolgus monkeys, and maintains robust effector functions such as antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity.

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Section: Resultsmentioning

confidence: 99%

Extending human IgG half-life using structure-guided design

Booth

Ramakrishnan

Narayan

et al. 2018

mAbs

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“…Third, the degradation or modification of antibodies during storage can lead to loss of binding affinity for FcRn. For example, methionine oxidation of two residues, Met252 and Met428, in proximity to the FcRn-(human) IgG1 interaction site can lead to decreased in vivo half-life (Bertolotti-Ciarlet et al, 2009; Gao et al, 2014; Wang et al, 2011b). Fourth, when targeting membrane-bound receptors, or multivalent soluble ligands, receptor-antibody internalization and degradation or IC-mediated internalization by FcγRs, respectively, may contribute to clearance (McKeage and Perry, 2002; Rowinsky et al, 2004; Suzuki et al, 2010; Tabrizi et al, 2006).…”

Section: Engineering Fcrn-igg Interactions To Modulate In Vivo Phamentioning

confidence: 99%

Targeting FcRn for the modulation of antibody dynamics

Ward

Devanaboyina

Ober

2015

Molecular Immunology

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The MHC Class I-related receptor, FcRn, is a multitasking protein that transports its IgG ligand within and across cells of diverse origins. The role of this receptor as a global regulator of IgG homeostasis and transport, combined with knowledge of the molecular details of FcRn-IgG interactions, has led to opportunities to modulate the in vivo dynamics of antibodies and their antigens through protein engineering. Consequently, the generation of half-life extended antibodies has shown a rapid expansion over the past decade. Further, FcRn itself can be targeted by inhibitors to induce decreased levels of circulating IgGs, which could have applications in multiple clinical settings. The engineering of antibody-antigen interactions to reduce antibody-mediated buffering of soluble ligand has also developed into an active area of investigation, leading to novel antibody platforms designed to lead to more effective antigen clearance. Similarly, the target-mediated elimination of antibodies by internalizing, membrane bound antigens (receptors) can be decreased using novel engineering approaches. These strategies, combined with subcellular trafficking analyses of antibody/antigen/FcRn behavior in cells to predict in vivo behavior, have considerable promise for the production of next generation therapeutics and diagnostics.

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“…Since both events are critical components of the FcRnIgG interaction, they should be included in a comprehensive analytical strategy as part of the candidate selection, quality control of therapeutic monoclonal antibodies (mAbs) or during quality by design studies [7][8][9]. Binding of IgG antibodies at low pH to FcRn is often characterized by surface plasmon resonance (SPR) binding assays [10][11][12][13]. Nevertheless, changes in FcRn binding measured by SPR did not correlate well with changes in serum half-life [14].…”

mentioning

confidence: 99%

Evaluation of an Fcrn Affinity Chromatographic Method for Igg1-Type Antibodies and Evaluation of Igg Variants

Cymer

Schlothauer²,

Knaupp³

et al. 2017

Bioanalysis

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Aim:The neonatal Fc-receptor (FcRn) mediates long serum half-life of therapeutic IgG-type antibodies. This interaction represents a critical quality attribute in terms of pharmacokinetics and should be covered by respective quality control strategies. Antibodies are taken up by cells unspecifically and can bind to FcRn in early endosomes preventing lysosomal degradation and allowing release back into circulation. Reflecting this complex cycle in an in vitro assay strategy represents a challenging task. Methodology: We report the qualification of an FcRn affinity chromatographic method and, for the first time, establish a noncriticality window. We analyzed different IgG-type antibodies, subtypes, glycoforms as well as mutants. Conclusion: The FcRn affinity chromatographic method allows the assessment of mAb samples with respect to their pH-dependent FcRn interaction. Furthermore, the method's capabilities and current limitations are discussed.

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Effect of Individual Fc Methionine Oxidation on FcRn Binding: Met252 Oxidation Impairs FcRn Binding More Profoundly than Met428 Oxidation

Cited by 91 publications

References 30 publications

Extending human IgG half-life using structure-guided design

Extending human IgG half-life using structure-guided design

Targeting FcRn for the modulation of antibody dynamics

Evaluation of an Fcrn Affinity Chromatographic Method for Igg1-Type Antibodies and Evaluation of Igg Variants

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