Charge variants characterization of a monoclonal antibody by ion exchange chromatography coupled on-line to native mass spectrometry: Case study after a long-term storage at+5 °C
Charge variants characterization of a monoclonal antibody by ion exchange chromatography coupled on-line to native mass spectrometry: Case study after a long-term storage at+5 °C
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DOI：
10.1016/j.jchromb.2017.02.017
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发表时间：
2017-03-24
影响因子：
3
通讯作者：
Chevreux, G.
中科院分区：
文献类型：
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作者：
Leblanc, Y.;Ramon, C.;Chevreux, G.
许多假定的翻译后修饰可能会引起单克隆抗体电荷分布的变化，从而可能影响其生物活性。这些电荷变体的表征和监控是确保稳定性和工艺一致性的关键质量要求。电荷变体通常通过制备型离子交换色谱、级分收集以及随后的反相液相色谱和质谱分析来表征。虽然这个过程可以通过在线二维色谱实现自动化，但它通常仍然复杂且耗时。因此，非常需要一种简单的在线电荷变体分析方法，分析实验室正在积极努力克服这一挑战。在这项研究中，开发了一种使用挥发性盐并在线耦合到天然质谱的混合模式离子交换色谱方法，并结合中上方法对单克隆抗体电荷变体进行详细表征。通过这种方法成功地研究了一种呈现复杂电荷变体特征的老化单克隆抗体作为案例研究。结果表明，5℃长期储存后，重链的脱酰胺化是主要的降解途径，而氧化程度相当低。该方法对于识别抗体的所有剪切形式也非常有用。 (C) 2017 LFB 生物技术公司。由 Elsevier B.V 出版。这是一篇遵循 CC BY-NC-ND 许可的开放获取文章
Numerous putative post-translational modifications may induce variations of monoclonal antibodies charge distribution that can potentially affect their biological activity. The characterization and the monitoring of these charge variants are critical quality requirements to ensure stability and process consistency. Charge variants are usually characterized by preparative ion exchange chromatography, collection of fractions and subsequent reverse-phase liquid chromatography with mass spectrometry analysis. While this process can be automatized by on-line two-dimensional chromatography, it remains often complex and time consuming. For this reason, a straightforward on-line charge variant analysis method is highly desirable and analytical laboratories are actively pursuing efforts to overcome this challenge.In this study, a mixed mode ion exchange chromatographic method using volatile salts and coupled online to native mass spectrometry was developed in association with a middle-up approach for a detailed characterization of monoclonal antibodies charge variants. An aged monoclonal antibody, presenting a complex charge variant profile was successfully investigated by this methodology as a case study. Results demonstrate that deamidation of the heavy chain was the major degradation pathway after long-term storage at 5 degrees C while oxidation was rather low. The method was also very useful to identify all the clipped forms of the antibody. (C) 2017 LFB Biotechnologies. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license