Kiese Sylvia, Papppenberger Astrid, Friess Wolfgang, Mahler Hanns-Christian
Formulation R&D Biologics, Pharmaceutical and Analytical R&D, F. Hoffmann-La Roche Ltd, Basel, Switzerland.
J Pharm Sci. 2008 Oct;97(10):4347-66. doi: 10.1002/jps.21328.
Protein aggregation is known to occur under different stress conditions and displays a wide variety of morphologies. In this work, the aggregation behavior of a monoclonal antibody (IgG1) was investigated using two different mechanical stress methods namely stirring and shaking at two temperatures, various fill volumes and headspaces and different amounts of polysorbate present in the formulation. The detection of aggregates in terms of size and number was carried out using various analytical techniques including visible particle inspection, turbidity, sub-visible particle analysis, size exclusion chromatography and dynamic light scattering. The data showed that shaking and stirring resulted in different species of aggregates both qualitatively and quantitatively, where stirring was found more stressful than shaking on the IgG1 formulation. Mechanical stress testing performed at 5 and 25 degrees C only showed a difference on samples stressed by shaking and not by stirring. The headspace in the vials had great influence on the stability of the protein formulation when stressed by shaking. The presence of polysorbate had a protective effect on the antibody, however certain polysorbate concentrations even resulted in increased protein aggregation. An array of analytical methods was essential in order to cover the vast aggregate morphologies, which occurred during agitation.
已知蛋白质聚集会在不同的应激条件下发生，并呈现出各种各样的形态。在这项工作中，使用两种不同的机械应激方法，即在两个温度、不同填充体积和顶空条件以及制剂中存在不同量聚山梨酯的情况下进行搅拌和振荡，研究了一种单克隆抗体（IgG1）的聚集行为。使用包括可见颗粒检查、浊度、亚可见颗粒分析、尺寸排阻色谱和动态光散射在内的各种分析技术，对聚集体的大小和数量进行检测。数据表明，振荡和搅拌在定性和定量方面都导致了不同种类的聚集体，其中发现搅拌对IgG1制剂的压力比振荡更大。在5摄氏度和25摄氏度下进行的机械应激测试仅显示出受振荡而非搅拌应激的样品存在差异。小瓶中的顶空在振荡应激时对蛋白质制剂的稳定性有很大影响。聚山梨酯的存在对抗体有保护作用，然而某些聚山梨酯浓度甚至会导致蛋白质聚集增加。为了涵盖搅拌过程中出现的大量聚集体形态学特征，一系列分析方法至关重要。
