Collision-Induced Unfolding Reveals Stability Differences in Infliximab Therapeutics under Native and Heat Stress Conditions.
Collision-Induced Unfolding Reveals Stability Differences in Infliximab Therapeutics under Native and Heat Stress Conditions.
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碰撞诱导的展开揭示了英夫利昔单抗治疗在天然和热应激条件下的稳定性差异。
DOI：
10.1021/acs.analchem.1c03946
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发表时间：
2021
影响因子：
7.4
通讯作者：
Ruotolo,BrandonT
中科院分区：
文献类型：
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作者：
Vallejo,DanielD;Kang,Jukyung;Coghlan,Jill;Ramírez,CarolinaRojas;Polasky,DanielA;Kurulugama,RuwanT;Fjeldsted,JohnC;Schwendeman,AnnaA;Ruotolo,BrandonT
已证明基于单克隆抗体（mAb）的生物治疗剂的离子迁移率-质谱（IM-MS）和碰撞诱导解折叠（CIU）测定对二硫键结构、糖基化模式和小分子缀合水平敏感。尽管有前景的先前报告详细说明了IM-MS和CIU区分生物仿制药、通用mAb治疗剂的能力，但围绕CIU对应激时发生的mAb结构变化的敏感性、跨IM-MS平台的此类测量的再现性以及CIU和差示扫描量热法（DSC）数据集之间的相关性仍然存在问题。在本报告中，我们描述了三种英夫利昔单抗（类克、Inflectra和Renflexis）采集的综合IM-MS和CIU数据集。我们通过热强制降解对每种英夫利西单抗样品进行强制降解，并观察到这三种生物治疗药物的稳定性模式大致相似但略有不同。我们发现，CIU是能够跟踪的mAb高阶结构（HOS）的差异，在强制热应力降解过程中给予的，DSC是不太敏感的这些变化相比。此外，我们在两个仪器平台（沃茨G2和Agilent 6560）上收集了全面的IM-MS和CIU数据，两者都产生了相似的区分mAb的能力，同时也揭示了两种仪器上获得的结果之间的微小差异。最后，我们证明，基于CIU的热图和分类可以快速评估英夫利西单抗分析中最具差异性的电荷状态，并且使用多重分类，我们保守估计进行mAb稳定性和HOS测量所需的时间缩短了30倍。标准DSC工具。
Ion mobility-mass spectrometry (IM-MS) and collision-induced unfolding (CIU) assays of monoclonal antibody (mAb)-based biotherapeutics have proven sensitive to disulfide bridge structures, glycosylation patterns, and small molecule conjugation levels. Despite promising prior reports detailing the capabilities of IM-MS and CIU to differentiate biosimilars, generic mAb therapeutics, there remain questions surrounding the sensitivity of CIU to mAb structure changes that occur upon stress, the reproducibility of such measurements across IM-MS platforms, and the correlation between CIU and differential scanning calorimetry (DSC) datasets. In this report, we describe a comprehensive IM-MS and CIU dataset acquired for three Infliximabs: Remicade, Inflectra, and Renflexis. We subject each infliximab sample to forced degradation through heat stress and observe broadly similar yet subtly different stability patterns for these three biotherapeutics. We find that CIU is capable of tracking differences in mAb higher-order structure (HOS) imparted during forced heat stress degradation and that DSC is less sensitive to these alterations in comparison. Furthermore, we collected our comprehensive IM-MS and CIU data across two instrument platforms (Waters G2 and Agilent 6560), with both producing similar abilities to differentiate mAbs while also revealing minor differences between the results obtained on the two instruments. Finally, we demonstrate that CIU-based heatmaps and classification allow for rapid assessment of the most differentiating charge states for the analysis of infliximab, and using multiplexed classification, we conservatively estimate a 30-fold improvement in the time required to perform mAb stability and HOS measurements over standard DSC tools.