序列消化可以识别和量化单克隆抗体轻链CDR中的快速酸异构化
Jérôme Jonveaux1, Marc Faudon1, Pauline Heymes1
1Lonza AG, Drug Product Services, Hochbergerstrasse 60G, Basel CH-4057, Switzerland.
Journal of pharmaceutical and biomedical analysis
|February 4, 2026
概括
单克隆抗体 (mAb) 从酸 (Asp) 变为异酸 (isoAsp) 的异构化,通过序列酶化消化精确地局部化. 这种方法在DS图案中发现了异构,随着时间的推移,液体配方的异构增加.
科学领域:
- 生物化学 生物化学
- 分析化学 分析化学
- 蛋白质化学 蛋白质化学
背景情况:
- 在单克隆抗体 (mAbs) 中,酸 (Asp) 到异酸 (isoAsp) 的异构化可以影响抗原结合亲和力并改变物理化学性质.
- 在配方开发过程中,准确地定位Asp异构化对于理解mAb稳定性至关重要.
研究的目的:
- 调查分析策略,以确定染色学和电泳学概况变化的根本原因在mAb.
- 开发一种方法来精确地定位和量化mAb互补性确定区域 (CDRs) 内的Asp异构位点.
主要方法:
- 高分辨率液态色谱-质谱学 (LC-MS) 分析减少的mAb.
- 使用序列酶性消化 (trypsin 跟着Asp-N) 的片映射.
- 减少的mAb的LC-UV量化.
主要成果:
- 顺序消化使得在mAb CDR中精确地定位Asp异构,克服了单酶消化的局限性.
- 首要的异构化部位被确定为Asp-Ser (DS) 基因,随着时间的推移,液体配方中的水平显著增加.
- 在冷化配方中没有观察到显著的异构,结果与LC-UV数据相对应良好.
结论:
- 序列酶化消化是一种强大的策略,用于准确地定位和量化mAbs.中的Asp异构化.
- 在DS基因的asp异构化是液体mAb配方中的一个时间依赖的降解途径.
- 解热化有效地减轻了Asp的异体化,保持了mAb的完整性.
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