Improved detection of host cell proteins (HCPs) in a mammalian cell‐derived antibody drug using liquid chromatography/mass spectrometry in conjunction with an HCP‐enrichment strategy

Thompson, Jenny Heidbrink; Chung, Wai Keen; Zhu, Min; Liu, Tie; Lu, Yali; Aboulaich, Nabila; Strouse, Robert J.; Mo, Wenjun

doi:10.1002/rcm.6854

Cited by 39 publications

(25 citation statements)

References 25 publications

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“…Recently, others have investigated using HCP enrichment strategies combined with LC‐MS/MS to improve the detection of HCPs from antibodies generated from mammalian cell expression systems (Thompson et al, ). Using such an approach, 19 HCPs were detected in a therapeutic antibody preparation compared to a single HCP identified without the enrichment strategy.…”

Section: Analytical Technologies For Hcp Analysismentioning

confidence: 99%

The future of host cell protein (HCP) identification during process development and manufacturing linked to a risk‐based management for their control

Bracewell

Francis

Smales

2015

Biotech & Bioengineering

151

128

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The use of biological systems to synthesize complex therapeutic products has been a remarkable success. However, during product development, great attention must be devoted to defining acceptable levels of impurities that derive from that biological system, heading this list are host cell proteins (HCPs). Recent advances in proteomic analytics have shown how diverse this class of impurities is; as such knowledge and capability grows inevitable questions have arisen about how thorough current approaches to measuring HCPs are. The fundamental issue is how to adequately measure (and in turn monitor and control) such a large number of protein species (potentially thousands of components) to ensure safe and efficacious products. A rather elegant solution is to use an immunoassay (enzyme‐linked immunosorbent assay [ELISA]) based on polyclonal antibodies raised to the host cell (biological system) used to synthesize a particular therapeutic product. However, the measurement is entirely dependent on the antibody serum used, which dictates the sensitivity of the assay and the degree of coverage of the HCP spectrum. It provides one summed analog value for HCP amount; a positive if all HCP components can be considered equal, a negative in the more likely event one associates greater risk with certain components of the HCP proteome. In a thorough risk‐based approach, one would wish to be able to account for this. These issues have led to the investigation of orthogonal analytical methods; most prominently mass spectrometry. These techniques can potentially both identify and quantify HCPs. The ability to measure and monitor thousands of proteins proportionally increases the amount of data acquired. Significant benefits exist if the information can be used to determine critical HCPs and thereby create an improved basis for risk management. We describe a nascent approach to risk assessment of HCPs based upon such data, drawing attention to timeliness in relation to biosimilar initiatives. The development of such an approach requires databases based on cumulative knowledge of multiple risk factors that would require national and international regulators, standards authorities (e.g., NIST and NIBSC), industry and academia to all be involved in shaping what is the best approach to the adoption of the latest bioanalytical technology to this area, which is vital to delivering safe efficacious biological medicines of all types. Biotechnol. Bioeng. 2015;112: 1727–1737. © 2015 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.

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Section: Analytical Technologies For Hcp Analysismentioning

confidence: 99%

The future of host cell protein (HCP) identification during process development and manufacturing linked to a risk‐based management for their control

Bracewell

Francis

Smales

2015

Biotech & Bioengineering

151

128

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“…[4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] In this methodology, proteins are first digested enzymatically. The resulting peptides are then separated (usually by liquid chromatography), and subsequently detected by mass/ charge via a mass spectrometer.…”

Section: Introductionmentioning

confidence: 99%

“…[4][5][6]11,14 Since HCPs are several orders of magnitude lower in concentration compared to the protein therapeutic, 2-dimensional LC separation helps increase the sensitivity of HCP detection. HCP quantitation is then achieved typically by spiking in one or more protein standards at known concentrations.…”

Section: Introductionmentioning

confidence: 99%

“…Other LC-MS/MS-based techniques have sought to first enrich the HCP population in an attempt to increase the number of host cell protein identifications. Several techniques such as immunoprecipitation, 10 Protein A enrichment, 14 and combinatorial ligand libraries 21 have shown great promise for enriching HCPs; however, the numbers of protein impurities detected were still very low, unless a substantial amount of protein impurities were present in the samples being analyzed (i.e., levels typically observed in the beginning of purification processes). While substantial advances have been made in improving HCP analysis, due to the shortcomings in current methodologies as just described, there is still a great need for alternative techniques to improve HCP characterization.…”

Section: Introductionmentioning

confidence: 99%

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Toward the complete characterization of host cell proteins in biotherapeutics via affinity depletions, LC-MS/MS, and multivariate analysis

Farutin²,

Carbeau³

et al. 2015

mAbs

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“…To address these challenges, methods using advanced liquid chromatography (LC) and mass spectrometry (MS) techniques have been developed, most of which focus on detecting low level HCPs in the final monoclonal antibody (mAb) products. [6][7][8][9][10][11] However, there is currently an unmet need to provide high throughput and robust LC-MS/MS analysis of HCPs in various in-process pools to help design and optimize purification processes toward optimal clearance.…”

Section: Introductionmentioning

confidence: 99%

A modular and adaptive mass spectrometry-based platform for support of bioprocess development toward optimal host cell protein clearance

Walker

Yang

Carver

et al. 2017

mAbs

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A modular and adaptive mass spectrometry (MS)-based platform was developed to provide fast, robust and sensitive host cell protein (HCP) analytics to support process development. This platform relies on one-dimensional ultra-high performance liquid chromatography (1D UHPLC) combined with several different MS data acquisition strategies to meet the needs of purification process development. The workflow was designed to allow HCP composition and quantitation for up to 20 samples per day, a throughput considered essential for real time bioprocess development support. With data-dependent acquisition (DDA), the 1D UHPLC-MS/MS method had excellent speed and demonstrated robustness in detecting unknown HCPs at 50 ng/mg (ppm) level. Combining 1D UHPLC with sequential window acquisition of all theoretical spectra (SWATH) MS enabled simultaneous detection and quantitation of all HCPs in single-digit ng/mg range within 1 hour, demonstrating for the first time the benefit of SWATH MS as a technique for HCP analysis. As another alternative, a targeted MS approach can be used to track the clearance of specific known HCP under various process conditions. This study highlights the importance of designing a robust LC-MS/MS workflow that not only allows HCP discovery, but also affords greatly improved process knowledge and capability in HCP removal. As an orthogonal and complementary detection approach to traditional HCP analysis by enzyme-linked immunosorbent assay, the reported LC-MS/MS workflow supports the development of bioprocesses with optimal HCP clearance and the production of safe and high quality therapeutic biopharmaceuticals.

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Improved detection of host cell proteins (HCPs) in a mammalian cell‐derived antibody drug using liquid chromatography/mass spectrometry in conjunction with an HCP‐enrichment strategy

Abstract: Compared with other published HCP studies with LC/MS, the HCP-enrichment step in our method enables a more practical and relevant application to approved protein therapeutics, which are mostly mammalian cell-derived products with HCPs present at very low levels.

Cited by 39 publications

References 25 publications

The future of host cell protein (HCP) identification during process development and manufacturing linked to a risk‐based management for their control

The future of host cell protein (HCP) identification during process development and manufacturing linked to a risk‐based management for their control

Toward the complete characterization of host cell proteins in biotherapeutics via affinity depletions, LC-MS/MS, and multivariate analysis

A modular and adaptive mass spectrometry-based platform for support of bioprocess development toward optimal host cell protein clearance

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