Towards a simple on-line coupling of ion exchange chromatography and native mass spectrometry for the detailed characterization of monoclonal antibodies

Murisier, Amarande; Duivelshof, Bastiaan L.; Fekete, Szabolcs; Bourquin, Julien; Schmudlach, Andrew; Lauber, Matthew; Nguyen, Jennifer M.; Beck, Alain; Guillarme, Davy; D’Atri, Valentina

doi:10.1016/j.chroma.2021.462499

Cited by 34 publications

(46 citation statements)

References 33 publications

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“…Within the 1D-LC analysis format, there has been an increase in the development of MS-compatible CEX methods and direct online coupling of CEX to MS using volatile salt-based pH gradients, giving rise to Native CEX-MS. This has been demonstrated by multiple groups with variations in the stationary phase (weak and strong ion exchangers) and MS platforms (ESI-MS, IM-MS, and orbitrap) for trastuzumab, adalimumab, infliximab, bevacizumab, and cetuximab (Bailey et al, 2018;Füssl et al, 2018;Sankaran et al, 2018;Jaag et al, 2021;Murisier et al, 2021) (Supplementary Table S7). Multidimensional platforms such as 2D-LC with CEX in first dimension followed by an MS-compatible second dimension, i.e., RP (desalting improves peak capacity and MS compatibility) have also been incorporated in biosimilarity (Alvarez et al, 2011;Stoll et al, 2015) (Supplementary Table S7).…”

Section: Charge Variantsmentioning

confidence: 91%

Section: Global Landscape On Biosimilar Approvalsmentioning

confidence: 92%

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Analytical Similarity Assessment of Biosimilars: Global Regulatory Landscape, Recent Studies and Major Advancements in Orthogonal Platforms

Nupur

Joshi

Gulliarme

et al. 2022

Front. Bioeng. Biotechnol.

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Biopharmaceuticals are one of the fastest-growing sectors in the biotechnology industry. Within the umbrella of biopharmaceuticals, the biosimilar segment is expanding with currently over 200 approved biosimilars, globally. The key step towards achieving a successful biosimilar approval is to establish analytical and clinical biosimilarity with the innovator. The objective of an analytical biosimilarity study is to demonstrate a highly similar profile with respect to variations in critical quality attributes (CQAs) of the biosimilar product, and these variations must lie within the range set by the innovator. This comprises a detailed comparative structural and functional characterization using appropriate, validated analytical methods to fingerprint the molecule and helps reduce the economic burden towards regulatory requirement of extensive preclinical/clinical similarity data, thus making biotechnological drugs more affordable. In the last decade, biosimilar manufacturing and associated regulations have become more established, leading to numerous approvals. Biosimilarity assessment exercises conducted towards approval are also published more frequently in the public domain. Consequently, some technical advancements in analytical sciences have also percolated to applications in analytical biosimilarity assessment. Keeping this in mind, this review aims at providing a holistic view of progresses in biosimilar analysis and approval. In this review, we have summarized the major developments in the global regulatory landscape with respect to biosimilar approvals and also catalogued biosimilarity assessment studies for recombinant DNA products available in the public domain. We have also covered recent advancements in analytical methods, orthogonal techniques, and platforms for biosimilar characterization, since 2015. The review specifically aims to serve as a comprehensive catalog for published biosimilarity assessment studies with details on analytical platform used and critical quality attributes (CQAs) covered for multiple biotherapeutic products. Through this compilation, the emergent evolution of techniques with respect to each CQA has also been charted and discussed. Lastly, the information resource of published biosimilarity assessment studies, created during literature search is anticipated to serve as a helpful reference for biopharmaceutical scientists and biosimilar developers.

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Section: Charge Variantsmentioning

confidence: 91%

Section: Global Landscape On Biosimilar Approvalsmentioning

confidence: 92%

Analytical Similarity Assessment of Biosimilars: Global Regulatory Landscape, Recent Studies and Major Advancements in Orthogonal Platforms

Nupur

Joshi

Gulliarme

et al. 2022

Front. Bioeng. Biotechnol.

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“…nMS and nIM-MS have been promoted for the characterization of biopharmaceuticals thanks to their quite straightforward and rapid workflows, especially through online nondenaturing liquid chromatography couplings. 15 − 17 IM separates ions based on their size, shape, and charge. Arrival time distributions (ATD) of ions offer structural information and may be converted into rotationally averaged collision cross sections (CCS, also labeled Ω), which reflect gas-phase conformations.…”

Section: Introductionmentioning

confidence: 99%

Combination of IM-Based Approaches to Unravel the Coexistence of Two Conformers on a Therapeutic Multispecific mAb

et al. 2022

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Multispecific antibodies, which target multiple antigens at once, are emerging as promising therapeutic entities to offer more effective treatment than conventional monoclonal antibodies (mAbs). However, these highly complex mAb formats pose significant analytical challenges. We report here on the characterization of a trispecific antibody (tsAb), which presents two isomeric forms clearly separated and identified with size exclusion chromatography coupled to native mass spectrometry (SEC-nMS). Previous studies showed that these isomers might originate from a proline cis / trans isomerization in one Fab subunit of the tsAb. We combined several innovative ion mobility (IM)-based approaches to confirm the isomeric nature of the two species and to gain new insights into the conformational landscape of both isomers. Preliminary SEC-nIM-MS measurements performed on a low IM resolution instrument provided the first hints of the coexistence of different conformers, while complementary collision-induced unfolding (CIU) experiments evidenced distinct gas-phase unfolding behaviors upon activation for the two isomers. As subtle conformational differences remained poorly resolved on our early generation IM platform, we performed high-resolution cyclic IM (cIM-MS) to unambiguously conclude on the coexistence of two conformers. The cis / trans equilibrium was further tackled by exploiting the IM n slicing capabilities of the cIM-MS instrument. Altogether, our results clearly illustrate the benefits of combining state-of-the-art nMS and IM-MS approaches to address challenging issues encountered in biopharma. As engineered antibody constructs become increasingly sophisticated, CIU and cIM-MS methodologies undoubtedly have the potential to integrate the drug development analytical toolbox to achieve in-depth conformational characterization of these products.

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“…Reversed-phase liquid chromatography (RPLC) is most commonly employed for separation of intact proteins prior to ESI-MS/MS; this mode of chromatography analyzes proteins in their denatured forms with proteins ionized in high charge states. , Several other separation modalities have been explored for analysis of intact proteins by mass spectrometry, including size-exclusion chromatography (SEC), − ion-exchange chromatography (IEX), − and hydrophobic interaction chromatography (HIC). − Although well-suited for analysis of native proteins using nondenaturing mobile phases, SEC suffers from low-resolution separation and significant sample dilution . IEX typically uses a salt or pH gradient, a factor that may alter MS sensitivity throughout the run, but has proven successful for biopharmaceutical applications .…”

Section: Introductionmentioning

confidence: 99%

Nanohydrophobic Interaction Chromatography Coupled to Ultraviolet Photodissociation Mass Spectrometry for the Analysis of Intact Proteins in Low Charge States

et al. 2022

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The direct correlation between proteoforms and biological phenotype necessitates the exploration of mass spectrometry (MS)-based methods more suitable for proteoform detection and characterization. Here, we couple nano-hydrophobic interaction chromatography (nano-HIC) to ultraviolet photodissociation MS (UVPD-MS) for separation and characterization of intact proteins and proteoforms. High linearity, sensitivity, and sequence coverage are obtained with this method for a variety of proteins. Investigation of collisional cross sections of intact proteins during nano-HIC indicates semifolded conformations in low charge states, enabling a different dimension of separation in comparison to traditional, fully denaturing reversed-phase separations. This method is demonstrated for a mixture of intact proteins from Escherichia coli ribosomes; high sequence coverage is obtained for a variety of modified and unmodified proteoforms.

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Towards a simple on-line coupling of ion exchange chromatography and native mass spectrometry for the detailed characterization of monoclonal antibodies

Cited by 34 publications

References 33 publications

Analytical Similarity Assessment of Biosimilars: Global Regulatory Landscape, Recent Studies and Major Advancements in Orthogonal Platforms

Analytical Similarity Assessment of Biosimilars: Global Regulatory Landscape, Recent Studies and Major Advancements in Orthogonal Platforms

Combination of IM-Based Approaches to Unravel the Coexistence of Two Conformers on a Therapeutic Multispecific mAb

Nanohydrophobic Interaction Chromatography Coupled to Ultraviolet Photodissociation Mass Spectrometry for the Analysis of Intact Proteins in Low Charge States

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