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Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate

Deamidation, a common post-translational modification, may impact multiple physiochemical properties of a therapeutic protein. MEDI7247, a pyrrolobenzodiazepine (PBD) antibody–drug conjugate (ADC), contains a unique deamidation site, N102, located within the complementarity-determining region (CDR),...

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Autores principales: Huang, Yue, Yuan, Jiaqi, Mu, Ruipeng, Kubiak, Robert J., Ball, Kathryn, Cao, Mingyan, Hussmann, G. Patrick, de Mel, Niluka, Liu, Dengfeng, Roskos, Lorin K., Liang, Meina, Rosenbaum, Anton I.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10594446/
https://www.ncbi.nlm.nih.gov/pubmed/37873863
http://dx.doi.org/10.3390/antib12040066
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author Huang, Yue
Yuan, Jiaqi
Mu, Ruipeng
Kubiak, Robert J.
Ball, Kathryn
Cao, Mingyan
Hussmann, G. Patrick
de Mel, Niluka
Liu, Dengfeng
Roskos, Lorin K.
Liang, Meina
Rosenbaum, Anton I.
author_facet Huang, Yue
Yuan, Jiaqi
Mu, Ruipeng
Kubiak, Robert J.
Ball, Kathryn
Cao, Mingyan
Hussmann, G. Patrick
de Mel, Niluka
Liu, Dengfeng
Roskos, Lorin K.
Liang, Meina
Rosenbaum, Anton I.
author_sort Huang, Yue
collection PubMed
description Deamidation, a common post-translational modification, may impact multiple physiochemical properties of a therapeutic protein. MEDI7247, a pyrrolobenzodiazepine (PBD) antibody–drug conjugate (ADC), contains a unique deamidation site, N102, located within the complementarity-determining region (CDR), impacting the affinity of MEDI7247 to its target. Therefore, it was necessary to monitor MEDI7247 deamidation status in vivo. Due to the low dose, a sensitive absolute quantification method using immunocapture coupled with liquid chromatography–tandem mass spectrometry (LBA-LC-MS/MS) was developed and qualified. We characterized the isomerization via Electron-Activated Dissociation (EAD), revealing that deamidation resulted in iso-aspartic acid. The absolute quantification of deamidation requires careful assay optimization in order not to perturb the balance of the deamidated and nondeamidated forms. Moreover, the selection of capture reagents essential for the correct quantitative assessment of deamidation was evaluated. The final assay was qualified with 50 ng/mL LLOQ for ADC for total and nondeamidated antibody quantification, with qualitative monitoring of the deamidated antibody. The impact of deamidation on the pharmacokinetic characteristics of MEDI7247 from clinical trial NCT03106428 was analyzed, revealing a gradual reduction in the nondeamidated form of MEDI7247 in vivo. Careful quantitative biotransformation analyses of complex biotherapeutic conjugates help us understand changes in product PTMs after administration, thus providing a more complete view of in vivo pharmacology.
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spelling pubmed-105944462023-10-25 Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate Huang, Yue Yuan, Jiaqi Mu, Ruipeng Kubiak, Robert J. Ball, Kathryn Cao, Mingyan Hussmann, G. Patrick de Mel, Niluka Liu, Dengfeng Roskos, Lorin K. Liang, Meina Rosenbaum, Anton I. Antibodies (Basel) Article Deamidation, a common post-translational modification, may impact multiple physiochemical properties of a therapeutic protein. MEDI7247, a pyrrolobenzodiazepine (PBD) antibody–drug conjugate (ADC), contains a unique deamidation site, N102, located within the complementarity-determining region (CDR), impacting the affinity of MEDI7247 to its target. Therefore, it was necessary to monitor MEDI7247 deamidation status in vivo. Due to the low dose, a sensitive absolute quantification method using immunocapture coupled with liquid chromatography–tandem mass spectrometry (LBA-LC-MS/MS) was developed and qualified. We characterized the isomerization via Electron-Activated Dissociation (EAD), revealing that deamidation resulted in iso-aspartic acid. The absolute quantification of deamidation requires careful assay optimization in order not to perturb the balance of the deamidated and nondeamidated forms. Moreover, the selection of capture reagents essential for the correct quantitative assessment of deamidation was evaluated. The final assay was qualified with 50 ng/mL LLOQ for ADC for total and nondeamidated antibody quantification, with qualitative monitoring of the deamidated antibody. The impact of deamidation on the pharmacokinetic characteristics of MEDI7247 from clinical trial NCT03106428 was analyzed, revealing a gradual reduction in the nondeamidated form of MEDI7247 in vivo. Careful quantitative biotransformation analyses of complex biotherapeutic conjugates help us understand changes in product PTMs after administration, thus providing a more complete view of in vivo pharmacology. MDPI 2023-10-17 /pmc/articles/PMC10594446/ /pubmed/37873863 http://dx.doi.org/10.3390/antib12040066 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Huang, Yue
Yuan, Jiaqi
Mu, Ruipeng
Kubiak, Robert J.
Ball, Kathryn
Cao, Mingyan
Hussmann, G. Patrick
de Mel, Niluka
Liu, Dengfeng
Roskos, Lorin K.
Liang, Meina
Rosenbaum, Anton I.
Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate
title Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate
title_full Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate
title_fullStr Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate
title_full_unstemmed Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate
title_short Multiplex Bioanalytical Methods for Comprehensive Characterization and Quantification of the Unique Complementarity-Determining-Region Deamidation of MEDI7247, an Anti-ASCT2 Pyrrolobenzodiazepine Antibody–Drug Conjugate
title_sort multiplex bioanalytical methods for comprehensive characterization and quantification of the unique complementarity-determining-region deamidation of medi7247, an anti-asct2 pyrrolobenzodiazepine antibody–drug conjugate
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10594446/
https://www.ncbi.nlm.nih.gov/pubmed/37873863
http://dx.doi.org/10.3390/antib12040066
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