Comparative higher-order structure analysis of antibody biosimilars using combined bottom-up and top-down hydrogen-deuterium exchange mass spectrometry

Pan, Jingxi; Zhang, Suping; Borchers, Christoph H.

doi:10.1016/j.bbapap.2016.08.013

Cited by 30 publications

(23 citation statements)

References 53 publications

(91 reference statements)

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“…Arguably less well‐recognized is the rapidly developing capability of MS instruments to analyze large, intact biomolecules under native‐like conditions . This, combined with coupled techniques like ion mobility spectrometry (IMS) and hydrogen/deuterium exchange (HDX) allows MS to support higher order structure analysis, in some cases at single residue resolution …”

Section: Introductionmentioning

confidence: 99%

“…The argument can always be made, however, that the gas‐phase conformational properties monitored by IMS do not adequately reflect conformational properties in solution. To allay these concerns, IMS analyses can be complemented by global HDX, which provides similar information (essentially a measure of relative global conformational stability and heterogeneity) based on isotope labeling in solution . In this work, we used a variation of HDX that involves short (ms timescale) isotope labeling times, called “time‐resolved” HDX.…”

Section: Introductionmentioning

confidence: 99%

“…[20][21][22][23][24][25] This, combined with coupled techniques like ion mobility spectrometry (IMS) 26,27 and hydrogen/deuterium exchange (HDX) [28][29][30] allows MS to support higher order structure analysis, in some cases at single residue resolution. 31,32 IMS is a relatively recent addition to the MS-based analysis toolbox, with a number of commercial instruments now including IMS technologies. IMS is essentially a gas-phase analog of electrophoresis, allowing for (usually pre-mass spectrometric) separation of analytes on the basis of size, shape, and charge.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Rapid characterization of structural and functional similarity for a candidate bevacizumab (Avastin) biosimilar using a multipronged mass‐spectrometry‐based approach

Brown

Rajendran

Dowd

et al. 2019

Drug Testing and Analysis

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The ongoing shift from small molecule drugs to protein therapeutics in the pharmaceuticals industry presents a considerable challenge to generic drug developers who are increasingly required to demonstrate biosimilarity for biological macromolecules, a task that is decidedly more complex than doing the same for small molecule drugs. In this work, we demonstrate a multipronged mass‐spectrometry‐based workflow that allows rapid and facile molecular characterization of antibody‐based protein therapeutics, applied to biosimilars development. Specifically, we use a combination of native mass spectrometry (MS), ion mobility spectrometry (IMS), and global time‐resolved hydrogen deuterium exchange (HDX) to provide an unambiguous assessment of the structural, dynamic, and chemical similarity between Avastin (bevacizumab) and a biosimilar in the late stages of pre‐clinical development. Minor structural and dynamic differences between the biosimilar and Avastin, and between lots of the biosimilar, were tested for functional relevance using Surface Plasmon Resonance‐derived kinetic and equilibrium binding parameters.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Rapid characterization of structural and functional similarity for a candidate bevacizumab (Avastin) biosimilar using a multipronged mass‐spectrometry‐based approach

Brown

Rajendran

Dowd

et al. 2019

Drug Testing and Analysis

View full text Add to dashboard Cite

show abstract

“…In the recent past, separations at < 0 °C, hereafter referred to as subzero, have been explored in order to limit the reversion of deuterium to hydrogen [9–13]. The intrinsic rate of exchange, k ex , is slowed by reducing the temperature [5].…”

Section: Introductionmentioning

confidence: 99%

“…Recently, Venable et al [9] showed this theory in practice when they demonstrated that reversion of deuterium to hydrogen in the analysis step can be greatly reduced by lowering the LC analysis temperature to −30 °C but preventing freezing with solvent modifiers (ethylene glycol and methanol) in the mobile phase. The increase in deuterium recovery afforded at subzero temperatures has also been employed to extend the ECD/ETD data accumulation window for top-down analyses of labeled proteins [10, 12, 13]. …”

Section: Introductionmentioning

confidence: 99%

Subzero Celsius separations in three-zone temperature controlled hydrogen deuterium exchange mass spectrometry

Wales

Fadgen

Eggertson

et al. 2017

Journal of Chromatography A

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Hydrogen deuterium exchange mass spectrometry (HDX MS) reports on the conformational landscape of proteins by monitoring the exchange between backbone amide hydrogen atoms and deuterium in the solvent. To maintain the label for analysis, quench conditions of low temperature and pH are required during the chromatography step performed after protease digestion but before mass spectrometry. Separation at 0 °C is often chosen as this is the temperature where the most deuterium can be recovered without freezing of the typical water and acetonitrile mobile phases. Several recent reports of separations at subzero Celsius emphasize the promise for retaining more deuterium and using a much longer chromatographic gradient or direct infusion time. Here we present the construction and validation of a modified Waters nanoACQUITY HDX manager with a third temperature-controlled zone for peptide separations at subzero temperatures. A new Peltier-cooled door replaces the door of a traditional main cooling chamber and the separations and trapping column are routed through the door housing. To prevent freezing, 35% methanol is introduced post online digestion. No new pumps are required and online digestion is performed as in the past. Subzero separations, using conventional HPLC column geometry of 3 μm particles in a 1x50mm column, did not result in major changes to chromatographic efficiency when lowering the temperature from 0 to −20 °C. There were significant increases in deuterium recovery for both model peptides and biologically relevant protein systems. Given the higher levels of deuterium recovery, expanded gradient programs can be used to allow for higher chromatographic peak capacity and therefore the analysis of larger and more complex proteins and systems.

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Considerations in the Analysis of Hydrogen Exchange Mass Spectrometry Data

Eggertson¹,

Fadgen²,

Engen³

et al. 2019

Mass Spectrometry Data Analysis in Proteomics

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Comparative higher-order structure analysis of antibody biosimilars using combined bottom-up and top-down hydrogen-deuterium exchange mass spectrometry

Cited by 30 publications

References 53 publications

Rapid characterization of structural and functional similarity for a candidate bevacizumab (Avastin) biosimilar using a multipronged mass‐spectrometry‐based approach

Rapid characterization of structural and functional similarity for a candidate bevacizumab (Avastin) biosimilar using a multipronged mass‐spectrometry‐based approach

Subzero Celsius separations in three-zone temperature controlled hydrogen deuterium exchange mass spectrometry

Considerations in the Analysis of Hydrogen Exchange Mass Spectrometry Data

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