Multi-attribute method based characterization of antibody drug conjugates (ADC) at the intact and subunit levels

Martelet, Armelle; Garrigue, Valérie; Zhang, Zoe; Genet, Bruno; Guttman, András

doi:10.1016/j.jpba.2021.114094

Cited by 16 publications

(9 citation statements)

References 36 publications

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“…Complementary MS in QC methodology may be considered for more complex molecule designs, such as antibody-drug conjugates, vaccine products, combination products, and multi-specific platforms. 32 , 38 , 39 This may also offer flexibility to continuously improve a CQA focused analytical control strategy that is customized to the specific structural properties and manufacturing process of the commercial mAb.…”

Section: Discussionmentioning

confidence: 99%

Targeted CQA analytical control strategy for commercial antibody products: Replacing ion-exchange chromatography methods for charge heterogeneity with multi-attribute monitoring

Evans,

Mulholland,

Lewis

et al. 2024

mAbs

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Peptide mapping with mass spectrometry (MS) is an important tool for protein characterization in the biopharmaceutical industry. Historically, peptide mapping monitors post-translational modifications (PTMs) of protein products and process intermediates during development. Multi-attribute monitoring (MAM) methods have been used previously in commercial release and stability testing panels to ensure control of selected critical quality attributes (CQAs). Our goal is to use MAM methods as part of an overall analytical testing strategy specifically focused on CQAs, while removing or replacing historical separation methods that do not effectively distinguish CQAs from non-CQAs due to co-elution. For example, in this study, we developed a strategy to replace a profile-based ion-exchange chromatography (IEC) method using a MAM method in combination with traditional purity methods to ensure control of charge variant CQAs for a commercial antibody (mAb) drug product (DP). To support this change in commercial testing strategy, the charge variant CQAs were identified and characterized during development by high-resolution LC-MS and LC-MS/MS. The charge variant CQAs included PTMs, high molecular weight species, and low molecular weight species. Thus, removal of the IEC method from the DP specification was achieved using a validated LC-MS MAM method on a QDa system to directly measure the charge variant PTM CQAs in combination with size exclusion chromatography (SE-HPLC) and capillary electrophoresis (CE-SDS) to measure the non-PTM charge variant CQAs. Bridging data between the MAM, IEC, and SE-HPLC methods were included in the commercial marketing application to justify removing IEC from the DP specification. We have also used this MAM method as a test for identity to reduce the number of QC assays. This strategy has received approvals from several health authorities.

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Section: Discussionmentioning

confidence: 99%

Targeted CQA analytical control strategy for commercial antibody products: Replacing ion-exchange chromatography methods for charge heterogeneity with multi-attribute monitoring

Evans,

Mulholland,

Lewis

et al. 2024

mAbs

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“…However, the large molecular weight of intact mAbbased drugs still represents a major challenge for MS/MS sequencing [33][34][35][36][37]. This can be substantially improved when fragmenting subunits [32,[38][39][40][41][42][43]. For instance, the complete sequencing of complementarity determining regions (CDRs) has been described on abovehinge subunits with multiple activation techniques [32,[44][45][46][47].…”

Section: Significance Statementmentioning

confidence: 99%

Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top‐down and middle‐down mass spectrometry

et al. 2023

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Monoclonal antibodies (mAbs) have established themselves as the leading biopharmaceutical therapeutic modality. Once the developability of a mAb drug candidate has been assessed, an important step is to check its in vivo stability through pharmacokinetics (PK) studies. The gold standard is ligand‐binding assay (LBA) and liquid chromatography‐mass spectrometry (LC‐MS) performed at the peptide level (bottom‐up approach). However, these analytical techniques do not allow to address the different mAb proteoforms that can arise from biotransformation. In recent years, top‐down and middle‐down mass spectrometry approaches have gained popularity to characterize proteins at the proteoform level but are not yet widely used for PK studies. We propose here a workflow based on an automated immunocapture followed by top‐down and middle‐down liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) approaches to characterize mAb proteoforms spiked in mouse plasma. We demonstrate the applicability of our workflow on a large concentration range using pembrolizumab as a model. We also compare the performance of two state‐of‐the‐art Orbitrap platforms (Tribrid Eclipse and Exploris 480) for these studies. The added value of our workflow for an accurate and sensitive characterization of mAb proteoforms in mouse plasma is highlighted.

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“…However, the large molecular weight of intact mAb-based drugs still represents a major challenge for MS/MS sequencing [33][34][35][36][37]. This can be substantially improved when fragmenting subunits [32,[38][39][40][41][42][43]. For instance, the complete sequencing of Complementarity Determining Regions (CDRs) has been described on above-hinge subunits with multiple activation techniques [32,[44][45][46][47].…”

Section: Introductionmentioning

confidence: 99%

Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top-down and middle-down mass spectrometry

Dhenin

Lafont

Dupré

et al. 2023

Preprint

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Monoclonal antibodies (mAbs) have established themselves as the leading biopharmaceutical therapeutic modality. Once the developability of a mAb drug candidate has been assessed, an important step is to check its in vivo stability through pharmacokinetics (PK) studies. The gold standard is ligand-binding assay (LBA) and LC–MS (Liquid Chromatography- Mass Spectrometry) performed at the peptide level (bottom-up approach). However, these analytical techniques do not allow to address the different mAb proteoforms that can arise from biotransformation. In recent years, top-down and middle-down mass spectrometry approaches have gained popularity to characterize proteins at the proteoform level but are not yet widely used for PK studies. We propose here a workflow based on an automated immunocapture followed by top-down and middle-down LC-MS/MS approaches to characterize mAb proteoforms spiked in mouse plasma. We demonstrate the applicability of our workflow on a large concentration range using pembrolizumab as a model. We also compare the performance of two state-of-the-art Orbitrap platforms (Tribrid Eclipse and Exploris 480) for these studies. The added value of our workflow for an accurate and sensitive characterization of mAb proteoforms in mouse plasma is highlighted.

show abstract

Multi-attribute method based characterization of antibody drug conjugates (ADC) at the intact and subunit levels

Cited by 16 publications

References 36 publications

Targeted CQA analytical control strategy for commercial antibody products: Replacing ion-exchange chromatography methods for charge heterogeneity with multi-attribute monitoring

Targeted CQA analytical control strategy for commercial antibody products: Replacing ion-exchange chromatography methods for charge heterogeneity with multi-attribute monitoring

Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top‐down and middle‐down mass spectrometry

Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top-down and middle-down mass spectrometry

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