Modern chromatographic and mass spectrometric techniques for protein biopharmaceutical characterization

Sandra, Koen; Vandenheede, Isabel; Sandra, Pat

doi:10.1016/j.chroma.2013.11.057

Cited by 136 publications

(115 citation statements)

References 221 publications

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“…The need for high resolution separation prior to MS is essential where multiple proteoforms of biopharmaceuticals, some of which are isobaric or differ by only 1 or a few Daltons and/or are in low abundance. Efforts are being made to develop LC columns suitable for high resolution intact protein separation [228]. At the same time, capillary zone electrophoresis (CZE) represents an alternative and complementary separation approach based on charge and hydrodynamic volume, with the potential for ultra-high efficiency [229][230][231].…”

Section: Introductionmentioning

confidence: 99%

Top-down proteomic analysis of protein pharmaceuticals, mixtures of protein complexes, and ribosomal protein extracts by capillary zone electrophoresis-mass spectrometry

Belov¹

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Section: Introductionmentioning

confidence: 99%

Top-down proteomic analysis of protein pharmaceuticals, mixtures of protein complexes, and ribosomal protein extracts by capillary zone electrophoresis-mass spectrometry

Belov¹

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“…iologics or biopharmaceutical products have seen an exponential growth, in terms of both scientific interest and market value, which is essentially driven by developments in the field of biosimilars and novel biopharmaceuticals [1][2][3][4][5][6]. Owing to the inherent complexity of biomacromolecules, the physicochemical characterization of biologics poses an analytical challenge.…”

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confidence: 99%

“…Nonetheless, comprehensive structural and functional characterization is a prerequisite to determine the quality, as it directly affects the product efficacy, safety, and immunogenicity. Amongst the physicochemical attributes, aggregation, charge heterogeneity, sequence variants, posttranslational modifications, and process-induced chemical modifications are critical to monitoring, and are done utilizing state-of-art analytical techniques [3,[7][8][9][10].…”

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confidence: 99%

Mass Spectrometry Based Mechanistic Insights into Formation of Tris Conjugates: Implications on Protein Biopharmaceutics

Kabadi

Sankaran

Palanivelu

et al. 2016

J. Am. Soc. Mass Spectrom.

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Abstract. We present here extensive mass spectrometric studies on the formation of a Tris conjugate with a therapeutic monoclonal antibody. The results not only demonstrate the reactive nature of the Tris molecule but also the sequence and reaction conditions that trigger this reactivity. The results corroborate the fact that proteins are, in general, prone to conjugation and/or adduct formation reactions and any modification due to this essentially leads to formation of impurities in a protein sample.Further, the results demonstrate that the conjugation reaction happens via a succinimide intermediate and has sequence specificity. Additionally, the data presented in this study also shows that the Tris formation is produced in-solution and is not an in-source phenomenon. We believe that the facts given here will open further avenues on exploration of Tris as a conjugating agent as well as ensure that the use of Tris or any ionic buffer in the process of producing a biopharmaceutical drug is monitored closely for the presence of such conjugate formation.

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“…The complexity of biologics attributes and the implementation of QbD strategies demand a multi-attribute method (MAM) that can monitor multiple biologics PQAs at the molecular level in a single assay. Coupling liquid chromatography (LC) to high resolution and high accuracy mass spectrometry (MS) techniques, LC-MS based peptide mapping has become a MAM approach that can identify and quantify multiple attributes of biologics simultaneously at the molecular level [6,7,11,12], as well as elucidate the mechanisms associated with degradation [7,13,14]. In a typical MAM workflow, therapeutic protein samples are denatured and reduced to break disulfide bonds between the cysteine residues, which are then alkylated by iodoacetamide.…”

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confidence: 99%

“…Analytical methods to identify and quantify these PQAs, especially CQAs, are essential for the development of QTPP and implementation of control strategies. Conventionally, a panel of analytical techniques such as size-exclusion chromatography (SEC), ion-exchange chromatography (IEX), hydrophobic-interaction chromatography (HIC), or capillary electrophoresis (CE) is typically used to monitor product quality consistency as well as product variants and impurities at the intact protein level [7][8][9]. Although these chromatographic and electrophoretic methods widely are used as release assays for biologics [10], they cannot directly monitor biologically relevant PQAs at the molecular level, which does not align with QbD principles.…”

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confidence: 99%

LC-MS multi-attribute method for characterization of biologics

Xu¹,

Qiu²,

Li³

2017

J Appl Bioanal

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Biologics, such as therapeutic monoclonal antibodies (mAbs), are complex protein molecules produced from mammalian tissue culture cells through recombinant DNA technology. As a result of naturally-occurring molecular heterogeneity as well as chemical and enzymatic modifications during manufacture, process, and storage, there are many product quality attributes (PQAs) presenting in therapeutic proteins. These PQAs can potentially include: product-related structural heterogeneity related to glycosylation profile, disulfide bond pattern, and higher order structure; product-related degradants and impurities, such as deamidation, oxidation, sequence variants; and process-related impurities and residuals, such as host cell protein (HCP), host cell DNA, and residual protein A [1]. Regulatory agencies have recently recommended a Quality by Design (QbD) approach for the manufacturing of therapeutic molecules [2][3][4][5], which requires in-depth understanding of these PQAs at the molecular level to ensure that the drug products meet the desired safety and efficacy profiles [6]. The QbD guidelines require development of a quality target product profile (QTPP) that identifies critical quality attributes (CQAs) and implementation of control strategies to ensure that the QTPP is achieved. QTPP is a prospective summary of the quality characteristics of a drug product to be achieved to ensure the desired quality, safety and efficacy [2]. QTPP describes the design criteria for the product and forms the basis for determination of the CQAs, critical process parameters (CPPs), and control strategy. A CQA is a physical, chemical, biological, or microbiological property or characteristic that should be within an appropriate limit, range, or distribution to ensure the desired product quality [2]. A CQA is identified based on the severity of harm to a patient resulting from failure to meet that quality attribute. Analytical methods to identify and quantify these PQAs, especially CQAs, are essential for the development of QTPP and implementation of control strategies. Conventionally, a panel of analytical techniques such as size-exclusion chromatography (SEC), ion-exchange chromatography (IEX), hydrophobic-interaction chromatography (HIC), or capillary electrophoresis (CE) is typically used to monitor product quality consistency as well as product variants and impurities at the intact protein level [7][8][9]. Although these chromatographic and electrophoretic methods widely are used as release assays for biologics [10], they cannot directly monitor biologically relevant PQAs at the molecular level, which does not align with QbD principles. The complexity of biologics attributes and the implementation of QbD strategies demand a multi-attribute method (MAM) that can monitor multiple biologics PQAs at the molecular level in a single assay. Coupling liquid chromatography (LC) to high resolution and high accuracy mass spectrometry (MS) techniques, LC-MS based peptide mapping has become a MAM approach that can identify and quantify multiple ...

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Modern chromatographic and mass spectrometric techniques for protein biopharmaceutical characterization

Cited by 136 publications

References 221 publications

Top-down proteomic analysis of protein pharmaceuticals, mixtures of protein complexes, and ribosomal protein extracts by capillary zone electrophoresis-mass spectrometry

Top-down proteomic analysis of protein pharmaceuticals, mixtures of protein complexes, and ribosomal protein extracts by capillary zone electrophoresis-mass spectrometry

Mass Spectrometry Based Mechanistic Insights into Formation of Tris Conjugates: Implications on Protein Biopharmaceutics

LC-MS multi-attribute method for characterization of biologics

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