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Submicron Aggregation of Chemically Denatured Monoclonal Antibody
Molecular Pharmaceutics ( IF 4.5 ) Pub Date : 2018-08-24 00:00:00 , DOI: 10.1021/acs.molpharmaceut.8b00690 Jacob B. Rowe 1 , Rhiannon P. Flynn 1 , Harrison R. Wooten 1 , Hailey A. Noufer 1 , Rachel A. Cancel 1 , Jifeng Zhang 2 , J. Anand Subramony 2 , Sergei Pechenov 2 , Ying Wang 1
Molecular Pharmaceutics ( IF 4.5 ) Pub Date : 2018-08-24 00:00:00 , DOI: 10.1021/acs.molpharmaceut.8b00690 Jacob B. Rowe 1 , Rhiannon P. Flynn 1 , Harrison R. Wooten 1 , Hailey A. Noufer 1 , Rachel A. Cancel 1 , Jifeng Zhang 2 , J. Anand Subramony 2 , Sergei Pechenov 2 , Ying Wang 1
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Isothermal chemical denaturation (ICD) has been widely used to evaluate the conformational stability of therapeutic proteins such as monoclonal antibodies. However, the chemical unfolding pathway and the subsequent aggregation of antibodies are not yet well-understood. In the present work, we conducted a systematic study on an ICD-induced aggregation of a pharmaceutical monoclonal antibody. Using dynamic light scattering, we monitored formation and growth of submicron aggregates in various buffers. Our experiments revealed a nucleation-controlled submicron aggregation of the antibody in the presence of chemical denaturant. After the unfolded protein reached a steady state, we reduced the denaturant concentration by dilution or dialysis to trigger further aggregation after ICD. In this way, we studied the pH effect on aggregation of the stressed protein after removal of denaturant. The ICD-dilution experiment provides a practical means for studying the propensity of unfolded proteins to form aggregates under various formulation conditions. This unique method allows us to control the degree of protein unfolding and the initiation of post-ICD aggregation.
中文翻译：
化学变性单克隆抗体的亚微米聚集
等温化学变性（ICD）已广泛用于评估治疗性蛋白质（如单克隆抗体）的构象稳定性。但是，化学展开途径和随后的抗体聚集尚不为人所理解。在目前的工作中，我们对ICD诱导的药物单克隆抗体的聚集进行了系统的研究。使用动态光散射，我们监测了各种缓冲液中亚微米聚集体的形成和生长。我们的实验揭示了在化学变性剂存在下抗体的成核控制亚微米聚集。展开的蛋白质达到稳态后，我们通过稀释或透析降低变性剂浓度，以引发ICD后的进一步聚集。通过这种方式，我们研究了去除变性剂后pH对应力蛋白聚集的影响。ICD稀释实验为研究未折叠蛋白在各种配制条件下形成聚集体的倾向提供了一种实用的方法。这种独特的方法使我们能够控制蛋白质的解折叠程度和ICD后聚集的启动。
更新日期：2018-08-24
中文翻译：
化学变性单克隆抗体的亚微米聚集
等温化学变性（ICD）已广泛用于评估治疗性蛋白质（如单克隆抗体）的构象稳定性。但是，化学展开途径和随后的抗体聚集尚不为人所理解。在目前的工作中，我们对ICD诱导的药物单克隆抗体的聚集进行了系统的研究。使用动态光散射，我们监测了各种缓冲液中亚微米聚集体的形成和生长。我们的实验揭示了在化学变性剂存在下抗体的成核控制亚微米聚集。展开的蛋白质达到稳态后，我们通过稀释或透析降低变性剂浓度，以引发ICD后的进一步聚集。通过这种方式，我们研究了去除变性剂后pH对应力蛋白聚集的影响。ICD稀释实验为研究未折叠蛋白在各种配制条件下形成聚集体的倾向提供了一种实用的方法。这种独特的方法使我们能够控制蛋白质的解折叠程度和ICD后聚集的启动。
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