Abstract:
Reversible self-association (RSA) of therapeutic proteins presents major challenges in the development of high-concentration formulations, especially those intended for subcutaneous administration. Understanding self-association mechanisms is therefore critical to the design and selection of candidates with acceptable developability to advance to clinical trials. The combination of experiments and in silico modeling presents a powerful tool to elucidate the interface of self-association. RSA of monoclonal antibodies has been studied extensively under different solution conditions and have been shown to involve interactions for both the antigen-binding fragment and the crystallizable fragment. Novel modalities such as bispecific antibodies, antigen-binding fragments, single-chain-variable fragments, and diabodies constitute a fast-growing class of antibody-based therapeutics that have unique physiochemical properties compared to monoclonal antibodies. In this study, the RSA interface of a diabody-interleukin 22 fusion protein (FP-1) was studied using hydrogen-deuterium exchange coupled with mass spectrometry (HDX-MS) in combination with in silico modeling. Taken together, the results show that a complex solution behavior underlies the self-association of FP-1 and that the interface thereof can be attributed to a specific segment in the variable light chain of the diabody. These findings also demonstrate that the combination of HDX-MS with in silico modeling is a powerful tool to guide the design and candidate selection of novel biotherapeutic modalities.
摘要:
治疗性蛋白质的可逆自缔合(RSA)在高浓度制剂的开发中提出了主要挑战。尤其是那些打算皮下给药的。因此,了解自缔合机制对于设计和选择具有可接受的可开发性的候选人以进行临床试验至关重要。实验和计算机建模的结合提供了阐明自关联界面的强大工具。已经在不同的溶液条件下广泛研究了单克隆抗体的RSA,并且已经显示涉及抗原结合片段和可结晶片段两者的相互作用。新型模态,如双特异性抗体,抗原结合片段,单链可变片段,和双抗体构成了一类快速增长的基于抗体的治疗剂,与单克隆抗体相比具有独特的生理化学性质。在这项研究中,双抗体-白细胞介素22融合蛋白(FP-1)的RSA界面使用氢-氘交换耦合质谱(HDX-MS)结合计算机模拟研究.一起来看,结果表明,复杂的溶液行为是FP-1自缔合的基础,其界面可归因于双抗体可变轻链中的特定片段。这些发现还表明,HDX-MS与计算机建模的组合是指导新型生物治疗方式的设计和候选选择的强大工具。