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Rapid screening of IgG quality attributes – effects on Fc receptor binding
The interactions of therapeutic antibodies with fragment crystallizable γ (Fcγ) receptors and neonatal Fc receptors (FcRn) are measured in vitro as indicators of antibody functional performance. Antibodies are anchored to immune cells through the Fc tail, and these interactions are important for the...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5623700/ https://www.ncbi.nlm.nih.gov/pubmed/28979843 http://dx.doi.org/10.1002/2211-5463.12283 |
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author | Geuijen, Karin P. M. Oppers‐Tiemissen, Cindy Egging, David F. Simons, Peter J. Boon, Louis Schasfoort, Richard B. M. Eppink, Michel H. M. |
author_facet | Geuijen, Karin P. M. Oppers‐Tiemissen, Cindy Egging, David F. Simons, Peter J. Boon, Louis Schasfoort, Richard B. M. Eppink, Michel H. M. |
author_sort | Geuijen, Karin P. M. |
collection | PubMed |
description | The interactions of therapeutic antibodies with fragment crystallizable γ (Fcγ) receptors and neonatal Fc receptors (FcRn) are measured in vitro as indicators of antibody functional performance. Antibodies are anchored to immune cells through the Fc tail, and these interactions are important for the efficacy and safety of therapeutic antibodies. High‐throughput binding studies on each of the human Fcγ receptor classes (FcγRI, FcγRIIa, FcγRIIb, FcγRIIIa, and FcγRIIIb) as well as FcRn have been developed and performed with human IgG after stress‐induced modifications to identify potential impact in vivo. Interestingly, we found that asparagine deamidation (D‐N) reduced the binding of IgG to the low‐affinity Fcγ receptors (FcγRIIa, FcγRIIb, FcγRIIIa, and FcγRIIIb), while FcγRI and FcRn binding was not impacted. Deglycosylation completely inhibited binding to all Fcγ receptors, but showed no impact on binding to FcRn. On the other hand, afucosylation only impacted binding to FcγRIIIa and FcγRIIIb. Methionine oxidation at levels below 7%, multiple freeze/thaw cycles and short‐term thermal/shake stress did not influence binding to any of the Fc receptors. The presence of high molecular weight species, or aggregates, disturbed measurements in these binding assays; up to 5% of aggregates in IgG samples changed the binding and kinetics to each of the Fc receptors. Overall, the screening assays described in this manuscript prove that rapid and multiplexed binding assays may be a valuable tool for lead optimization, process development, in‐process controls, and biosimilarity assessment of IgGs during development and manufacturing of therapeutic IgGs. |
format | Online Article Text |
id | pubmed-5623700 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-56237002017-10-04 Rapid screening of IgG quality attributes – effects on Fc receptor binding Geuijen, Karin P. M. Oppers‐Tiemissen, Cindy Egging, David F. Simons, Peter J. Boon, Louis Schasfoort, Richard B. M. Eppink, Michel H. M. FEBS Open Bio Research Articles The interactions of therapeutic antibodies with fragment crystallizable γ (Fcγ) receptors and neonatal Fc receptors (FcRn) are measured in vitro as indicators of antibody functional performance. Antibodies are anchored to immune cells through the Fc tail, and these interactions are important for the efficacy and safety of therapeutic antibodies. High‐throughput binding studies on each of the human Fcγ receptor classes (FcγRI, FcγRIIa, FcγRIIb, FcγRIIIa, and FcγRIIIb) as well as FcRn have been developed and performed with human IgG after stress‐induced modifications to identify potential impact in vivo. Interestingly, we found that asparagine deamidation (D‐N) reduced the binding of IgG to the low‐affinity Fcγ receptors (FcγRIIa, FcγRIIb, FcγRIIIa, and FcγRIIIb), while FcγRI and FcRn binding was not impacted. Deglycosylation completely inhibited binding to all Fcγ receptors, but showed no impact on binding to FcRn. On the other hand, afucosylation only impacted binding to FcγRIIIa and FcγRIIIb. Methionine oxidation at levels below 7%, multiple freeze/thaw cycles and short‐term thermal/shake stress did not influence binding to any of the Fc receptors. The presence of high molecular weight species, or aggregates, disturbed measurements in these binding assays; up to 5% of aggregates in IgG samples changed the binding and kinetics to each of the Fc receptors. Overall, the screening assays described in this manuscript prove that rapid and multiplexed binding assays may be a valuable tool for lead optimization, process development, in‐process controls, and biosimilarity assessment of IgGs during development and manufacturing of therapeutic IgGs. John Wiley and Sons Inc. 2017-09-05 /pmc/articles/PMC5623700/ /pubmed/28979843 http://dx.doi.org/10.1002/2211-5463.12283 Text en © 2017 The Authors. Published by FEBS Press and John Wiley & Sons Ltd. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Articles Geuijen, Karin P. M. Oppers‐Tiemissen, Cindy Egging, David F. Simons, Peter J. Boon, Louis Schasfoort, Richard B. M. Eppink, Michel H. M. Rapid screening of IgG quality attributes – effects on Fc receptor binding |
title | Rapid screening of IgG quality attributes – effects on Fc receptor binding |
title_full | Rapid screening of IgG quality attributes – effects on Fc receptor binding |
title_fullStr | Rapid screening of IgG quality attributes – effects on Fc receptor binding |
title_full_unstemmed | Rapid screening of IgG quality attributes – effects on Fc receptor binding |
title_short | Rapid screening of IgG quality attributes – effects on Fc receptor binding |
title_sort | rapid screening of igg quality attributes – effects on fc receptor binding |
topic | Research Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5623700/ https://www.ncbi.nlm.nih.gov/pubmed/28979843 http://dx.doi.org/10.1002/2211-5463.12283 |
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