Interfacial Stress in the Development of Biologics: Fundamental Understanding, Current Practice, and Future Perspective

Abstract

Biologic products encounter various types of interfacial stress during development, manufacturing, and clinical administration. When proteins come in contact with vapor–liquid, solid–liquid, and liquid–liquid surfaces, these interfaces can significantly impact the protein drug product quality attributes, including formation of visible particles, subvisible particles, or soluble aggregates, or changes in target protein concentration due to adsorption of the molecule to various interfaces. Protein aggregation at interfaces is often accompanied by changes in conformation, as proteins modify their higher order structure in response to interfacial stresses such as hydrophobicity, charge, and mechanical stress. Formation of aggregates may elicit immunogenicity concerns; therefore, it is important to minimize opportunities for aggregation by performing a systematic evaluation of interfacial stress throughout the product development cycle and to develop appropriate mitigation strategies. The purpose of this white paper is to provide an understanding of protein interfacial stability, explore methods to understand interfacial behavior of proteins, then describe current industry approaches to address interfacial stability concerns. Specifically, we will discuss interfacial stresses to which proteins are exposed from drug substance manufacture through clinical administration, as well as the analytical techniques used to evaluate the resulting impact on the stability of the protein. A high-level mechanistic understanding of the relationship between interfacial stress and aggregation will be introduced, as well as some novel techniques for measuring and better understanding the interfacial behavior of proteins. Finally, some best practices in the evaluation and minimization of interfacial stress will be recommended.

Synergistic effects of flow and interfaces on antibody aggregation

“…For instance, in both Hamilton syringes and silica nanoparticles, antibody molecules adsorb to glass via charge‐charge interactions, leading to the formation of protein films. Hydrodynamic flow and mechanical scraping provide removal and renewal of the layer of protein aggregates formed at the interfaces (Li et al, ; Liu et al, ; Rudiuk et al, ). Removal of the absorbed protein layer results in shedding of protein particles into the bulk solution and causes re‐exposure of the surfaces offered by the experimental setup to the protein solution.…”

“…For instance, in both Hamilton syringes and silica nanoparticles, antibody molecules adsorb to glass via charge-charge interactions, leading to the formation of protein films. Hydrodynamic flow and mechanical scraping provide removal and renewal of the layer of protein aggregates formed at the interfaces (Li et al, 2019;Liu et al, 2012;Rudiuk et al, 2012).…”

“…In addition to air–water interfaces, also oil‐water (Bee, Randolph, Carpenter, Bishop, & Dimitrova, ; Jones et al, ; Mehta et al, ; Perevozchikova et al, ; Thirumangalathu et al, ) and solid–liquid interfaces (Bee, Chiu et al, ; Biddlecombe et al, ; Hoehne et al, ; Kalonia et al, ; Tyagi et al, ), including nanometer‐scale surface roughness (Biddlecombe et al, ; Biddlecombe et al, ) and subvisible solid particles shed from surfaces during bioprocessing (Carpenter et al, ; Tyagi et al, ), have been reported to have detrimental effects on the stability of protein formulations. In general, it appears that absorption of protein molecules to interfaces can result in the formation of protein films which can then be dislodged and broken up by mechanical forces including shear, compression‐dilation, and rupture (Li et al, ; Liu, Randolph, & Carpenter, ; Rudiuk, Cohen‐Tannoudji, Huille, & Tribet, ). Also, the aggregates and particles released into the bulk solution can promote further aggregation and additional interactions with interfaces.…”

Stress Factors in mAb Drug Substance Production Processes: Critical Assessment of Impact on Product Quality and Control Strategy

“…There is a fine line between creating a reducing state and having an environment with an excess of ROS which can result in oxidation and chemical modification of proteins. While some of the chemical modifications are benign, others can affect the active site and the bioactivity of the protein, or result in partial protein unfolding creating new (often more hydrophobic) 57 epitopes which could potentially increase the amount of aggregates and perhaps immunogenicity.…”

“…166 For proteins that have a strong tendency to self-associate especially at high concentrations, often needed for low-volume subcutaneous delivery, the UF/DF operation becomes more difficult to manage. 18,57 A recent article thoroughly reviews the impact of interfacial interactions on protein aggregation.…”