LC-MS multi-attribute method for characterization of biologics

Abstract

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 ...

A View on the Importance of “Multi-Attribute Method” for Measuring Purity of Biopharmaceuticals and Improving Overall Control Strategy

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Quantitation and modeling of post-translational modifications in a therapeutic monoclonal antibody from single- and multiple-dose monkey pharmacokinetic studies using mass spectrometry

“…PTMs are a therapeutic mAb product quality attributes (PQAs). Controlling PQAs within predefined acceptance criteria is vital to the biopharmaceutical industry because it ensures consistent product quality and reduces potential impacts on drug safety and efficacy[5]. The modifications that occur during drug production and storage can be reliably monitored and controlled.…”

Isotyping and Semi-Quantitation of Monkey Anti-Drug Antibodies by Immunocapture Liquid Chromatography-Mass Spectrometry

“…With the rapid development of new instrumentation and methods, liquid chromatography-mass spectrometry (LC-MS) has emerged as a critical tool for the characterization of biologicals (13)(14)(15). Many efforts have been made to adopt the LC-MS-based approach as a supplementary tool for ADA analysis (16).…”