Griffin Valerie P, Nottar Escobar Estephanie L, Kanthe Ankit, Gokhale Madhushree, Dhar Prajnaparamita
Department of Chemical and Petroleum Engineering, The University of Kansas, 1530 W 15th Street, Lawrence, Kansas 66045, United States.
Drug Product Development, Bristol Myers Squibb, New Brunswick, New Jersey 08903, United States.
ACS Appl Bio Mater. 2025 Aug 18;8(8):6928-6940. doi: 10.1021/acsabm.5c00661. Epub 2025 Jul 14.
Therapeutic proteins, such as monoclonal antibodies (mAbs), can form two-dimensional films at the air-water interface, which, when ruptured or stressed, can lead to protein instability in solution. However, the details of the dynamics of mAb film formation, particularly the transitions in the rheological properties of the film, are not well understood. The emergence of novel protein-based modalities also raises the question of whether our understanding of mAb film formation can be extended to other novel biotherapeutic modalities. This work aims to understand the dynamics of film formation in novel therapeutic proteins by correlating protein adsorption kinetics, measured using surface tensiometry, with the evolution of the interfacial rheological properties, measured using a passive microrheology technique, for two protein modalities: IgG mAb and Fc-Fusion protein. Further, to accurately record how differences in packing density at the interface might lead to differences in the rheological properties in these different protein modalities during film aging, the bulk protein concentration was varied over 3 orders of magnitude. Our results indicate that the multistage protein adsorption process seen in both protein modalities can result in transitions in film rheology from purely viscous to viscoelastic and elastic films with time, depending on the bulk concentration and protein modality. Fc-Fusion proteins demonstrated an earlier onset of viscoelastic film transition compared with IgG mAbs for all bulk concentrations studied. Further, polysorbate 80 (PS80), often added to protein solutions to mitigate protein aggregation, prevents this transition in IgG mAb solutions but not in Fc-Fusion protein solutions. Together, our results provide rheological characterization of the protein films during adsorption and film aging in these novel biotherapeutics and will help inform the development of appropriate mitigation strategies to maintain their interfacial stability.
治疗性蛋白质，如单克隆抗体（mAb），可在气-水界面形成二维膜，当该膜破裂或受力时，会导致溶液中的蛋白质不稳定。然而，mAb膜形成动力学的细节，尤其是膜流变学性质的转变，目前尚不清楚。新型蛋白质类药物的出现也引发了一个问题，即我们对mAb膜形成的理解是否能扩展到其他新型生物治疗药物。这项工作旨在通过将使用表面张力测定法测量的蛋白质吸附动力学与使用被动微流变技术测量的界面流变学性质的演变相关联，来了解新型治疗性蛋白质中膜形成的动力学，涉及两种蛋白质类型：IgG mAb和Fc融合蛋白。此外，为了准确记录在膜老化过程中界面堆积密度的差异如何导致这些不同蛋白质类型的流变学性质差异，将本体蛋白质浓度在3个数量级范围内变化。我们的结果表明，在这两种蛋白质类型中观察到的多阶段蛋白质吸附过程可导致膜流变学随时间从纯粘性膜转变为粘弹性膜和弹性膜，这取决于本体浓度和蛋白质类型。在所研究的所有本体浓度下，与IgG mAb相比，Fc融合蛋白表现出更早的粘弹性膜转变。此外，聚山梨酯80（PS80）通常添加到蛋白质溶液中以减轻蛋白质聚集，它可防止IgG mAb溶液中的这种转变，但不能防止Fc融合蛋白溶液中的转变。总之，我们的结果提供了这些新型生物治疗药物在吸附和膜老化过程中蛋白质膜的流变学特征，并将有助于为制定适当的缓解策略以维持其界面稳定性提供参考。
