Aggregation-Prone Motifs in Human Immunoglobulin G

Chennamsetty, Naresh; Helk, Bernhard; Voynov, Vladimir; Kayser, Veysel; Trout, Bernhardt L.

doi:10.1016/j.jmb.2009.06.028

Cited by 142 publications

(133 citation statements)

References 41 publications

(45 reference statements)

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“…[25][26][27][28] One such study that is presently ongoing suggests that deglycosylation permits the CH 2 -CH 2 domains of the Fc region to assume conformations in which these two domains are farther apart than in corresponding glycosylated structures (personal communication, TW Patapoff and JP Brandt). This phenomenon may account for the slightly larger hydrodynamic volume of deglycosylated antibodies as detected by SEC.…”

Section: Discussionmentioning

confidence: 99%

The impact of glycosylation on monoclonal antibody conformation and stability

Zheng

Bantog

Bayer

2011

mAbs

268

210

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Section: Discussionmentioning

confidence: 99%

The impact of glycosylation on monoclonal antibody conformation and stability

Zheng

Bantog

Bayer

2011

mAbs

268

210

View full text Add to dashboard Cite

“…[5][6][7]9,18,19,21,23,27 While the former can be limited due to the degree of knowledge of sites of interactions, the latter can be time and resource intensive and may not always be fully successful to a desired degree for all antibodies. Historically, a number of computational and analytical screening methods have been used to predict aggregation [28][29][30] and high concentration formulation risks. 17,[31][32][33][34][35][36] While these approaches can provide valuable information and help screening of potential lead candidates during discovery and early stage development, they do not necessarily provide insight into the underlying molecular mechanisms involved.…”

Section: Introductionmentioning

confidence: 99%

Mitigation of reversible self-association and viscosity in a human IgG1 monoclonal antibody by rational, structure-guided Fv engineering

Geoghegan¹,

Fleming²,

Damschroder³

et al. 2016

mAbs

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Undesired solution behaviors such as reversible self-association (RSA), high viscosity, and liquid-liquid phase separation can introduce substantial challenges during development of monoclonal antibody formulations. Although a global mechanistic understanding of RSA (i.e., native and reversible proteinprotein interactions) is sufficient to develop robust formulation controls, its mitigation via protein engineering requires knowledge of the sites of protein-protein interactions. In the study reported here, we coupled our previous hydrogen-deuterium exchange mass spectrometry findings with structural modeling and in vitro screening to identify the residues responsible for RSA of a model IgG1 monoclonal antibody (mAb-C), and rationally engineered variants with improved solution properties (i.e., reduced RSA and viscosity). Our data show that mutation of either solvent-exposed aromatic residues within the heavy and light chain variable regions or buried residues within the heavy chain/light chain interface can significantly mitigate RSA and viscosity by reducing the IgG's surface hydrophobicity. The engineering strategy described here highlights the utility of integrating complementary experimental and in silico methods to identify mutations that can improve developability, in particular, high concentration solution properties, of candidate therapeutic antibodies.

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“…SAP identifies the location and size of these aggregation-prone regions. Chennamsetty et al [33,34,47] demonstrated that SAP calculations can be used to determine critical regions of aggregation in therapeutic antibodies. The aggregation-prone motifs for the Fc region of antibodies have been identified with use of SAP and other methods [47].…”

Section: Correlation Between Hdx-ms and Sap Resultsmentioning

confidence: 99%

“…Chennamsetty et al [33,34,47] demonstrated that SAP calculations can be used to determine critical regions of aggregation in therapeutic antibodies. The aggregation-prone motifs for the Fc region of antibodies have been identified with use of SAP and other methods [47]. In the FcP1 aggregation study, although no structure is currently available for the full-length protein, the crystal structures of ABP and the human IgG1 Fc domain are available.…”

Section: Correlation Between Hdx-ms and Sap Resultsmentioning

confidence: 99%

Characterization of Aggregation Propensity of a Human Fc-Fusion Protein Therapeutic by Hydrogen/Deuterium Exchange Mass Spectrometry

Huang

Iacob

Krystek

et al. 2016

J. Am. Soc. Mass Spectrom.

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Abstract. Aggregation of protein therapeutics has long been a concern across different stages of manufacturing processes in the biopharmaceutical industry. It is often indicative of aberrant protein therapeutic higher-order structure. In this study, the aggregation propensity of a human Fc-fusion protein therapeutic was characterized. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) was applied to examine the conformational dynamics of dimers collected from a bioreactor. HDX-MS data combined with spatial aggregation propensity calculations revealed a potential aggregation interface in the Fc domain. This study provides a general strategy for the characterization of the aggregation propensity of Fc-fusion proteins at the molecular level.

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Aggregation-Prone Motifs in Human Immunoglobulin G

Cited by 142 publications

References 41 publications

The impact of glycosylation on monoclonal antibody conformation and stability

The impact of glycosylation on monoclonal antibody conformation and stability

Mitigation of reversible self-association and viscosity in a human IgG1 monoclonal antibody by rational, structure-guided Fv engineering

Characterization of Aggregation Propensity of a Human Fc-Fusion Protein Therapeutic by Hydrogen/Deuterium Exchange Mass Spectrometry

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