Electrostatic interactions modulate the differential aggregation propensities of IgG1 and IgG4P antibodies and inform charged residue substitutions for improved developability
Electrostatic interactions modulate the differential aggregation propensities of IgG1 and IgG4P antibodies and inform charged residue substitutions for improved developability
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DOI：
10.1093/protein/gzz046
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发表时间：
2019-06-01
影响因子：
2.4
通讯作者：
Lawson, Alastair D. G.
中科院分区：
文献类型：
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作者：
Heads, James T.;Lamb, Richard;Lawson, Alastair D. G.
天然状态聚集是治疗性抗体开发中的重要关注点。增强mAb天然状态聚集机制的知识将允许基于序列的选择和设计具有改善的可开发性的治疗性mAb。我们研究了静电相互作用如何影响7个人IgG1和IgG4P mAb同种型对的天然状态聚集，每对具有相同的可变结构域，而每组IgG1和IgG4P构建体的可变结构域不同。在pH 7.4（代表生理条件）和pH 5.0（代表常用储存条件）下测定相对聚集倾向。我们的工作表明，相对于恒定结构域的净电荷状态的可变结构域的净电荷状态是主要负责不同的天然状态的聚集行为的IgG1和IgG4P单抗。这一观察结果表明，多结构域蛋白质的全局净电荷不是聚集倾向的可靠预测因子。此外，我们展示了可变域的框架中的设计策略，以减少被确定为聚集倾向的单克隆抗体的原生状态聚集倾向。重要的是，用替代的人种系残基取代特异性鉴定的残基以优化Fv电荷，导致在pH 5.0和7.4下聚集潜力降低，从而增加可开发性。
Native state aggregation is an important concern in the development of therapeutic antibodies. Enhanced knowledge of mAb native state aggregation mechanisms would permit sequence-based selection and design of therapeutic mAbs with improved developability. We investigated how electrostatic interactions affect the native state aggregation of seven human IgG1 and IgG4P mAb isotype pairs, each pair having identical variable domains that are different for each set of IgG1 and IgG4P constructs. Relative aggregation propensities were determined at pH 7.4, representing physiological conditions, and pH 5.0, representing commonly used storage conditions. Our work indicates that the net charge state of variable domains relative to the net charge state of the constant domains is predominantly responsible for the different native state aggregation behavior of IgG1 and IgG4P mAbs. This observation suggests that the global net charge of a multi domain protein is not a reliable predictor of aggregation propensity. Furthermore, we demonstrate a design strategy in the frameworks of variable domains to reduce the native state aggregation propensity of mAbs identified as being aggregation-prone. Importantly, substitution of specifically identified residues with alternative, human germline residues, to optimize Fv charge, resulted in decreased aggregation potential at pH 5.0 and 7.4, thus increasing developability.