What Makes Polysorbate Functional? Impact of Polysorbate 80 Grade and Quality on IgG Stability During Mechanical Stress

Abstract

Polysorbate 80 (PS80) is a commonly used surfactant in therapeutic protein formulations to mitigate adsorption and interface-induced protein aggregation. Several PS80 grades and qualities are available on the market for parenteral application. The role of PS80 grade on protein stability remains debatable, and the impact of (partially) degraded PS on protein aggregation is not yet well understood. In our study, a monoclonal antibody (IgG) was subjected to 3 different mechanical stress conditions in the presence of multicompendial (MC) and Chinese pharmacopeia (ChP) grade PS80. Furthermore, IgG formulations were spiked with (partly) hydrolyzed PS80 to investigate the effect of PS80 degradants on protein stability. PS80 functionality was assessed by measuring the extent of protein aggregation and particle formation induced during mechanical stress by using size-exclusion chromatography, dynamic light scattering, backgrounded membrane imaging, and flow imaging microscopy. No distinguishable differences in PS80 functionality between MC and ChP grade were observed in the 3 stress tests. However, with increasing degree of PS80 hydrolysis, higher counts of subvisible particles were measured after stress. Furthermore, higher levels of PS80 degradants at a constant PS80 concentration may destabilize the IgG. In conclusion, MC and ChP grade PS80 are equally protective, but PS80 degradants compromise IgG stability.

The uniqueness of flow in probing the aggregation behavior of clinically relevant antibodies

“…Using an array of proteins, including three IgG1 mAbs, we have also showed that extensional (and subsequent shear) flow induced by the EFD can trigger protein aggregation and that these effects were extremely protein‐ and flow‐field specific . More recently, EFDs of similar design to that employed in our study have been constructed and mAb aggregation observed under extensional and shear flows . These and other studies have shown that the presence and chemical composition of interfaces and presence of localized high protein concentrations within flow devices also contribute to the aggregation cascade.…”

“… 59 , 60 More recently, EFDs of similar design to that employed in our study have been constructed and mAb aggregation observed under extensional and shear flows. 87 , 88 These and other studies 57 , 79 , 85 , 89 , 90 , 91 , 92 , 93 have shown that the presence and chemical composition of interfaces and presence of localized high protein concentrations within flow devices also contribute to the aggregation cascade. Others have suggested that the friction between the solid components of flow devices, for example, Couette cylinders 89 or plungers and syringe barrels, can also induce aggregation.…”

Effect of Fatty Acid Composition in Polysorbate 80 on the Stability of Therapeutic Protein Formulations

“…Commercial Polysorbate 80 NF excipients can exhibit significant variation in fatty acid composition, but the impact of fatty acid as a variable in terms of polysorbate's role in causing particle formation is not completely understood. Recently, several groups have evaluated the functionality of common and novel polysorbate 80 excipients when subjected to varying mechanical and oxidative stresses (27)(28)(29). Kranz et al evaluated enzyme-mediated hydrolysis of different grades of polysorbate 80 using an esterase from Saccharomyces cerevisiae, and that both the commercial excipient grade of Polysorbate 80, commonly used in US, Europe and Japan, and a novel polysorbate 80 grade synthesized from pure oleic acid were similarly susceptible to enzymatic hydrolysis.…”

“…It is possible that in formulations that had undergone some fatty acid hydrolysis sufficient PS-80 was still present to effectively protect the therapeutic protein from any interfacial stress (6). Indeed, previous studies have demonstrated that a 10% loss of polysorbate did not reduce functionality of either compendial or higher purity grades of polysorbate (28). It has also been shown that other physical stressors, such as dropping or even shipping, can lead to particle formation and changes in the therapeutic protein (27,28,34,35).…”

Characterization of Polysorbate 80 by Liquid Chromatography-Mass Spectrometry to Understand Its Susceptibility to Degradation and Its Oxidative Degradation Pathway

“…13 It was reported that MC and ChP grade PS80 are equally protective, but a higher level of PS80 degradants may destabilize proteins. 30 The degradants generated from each degradation pathway, as discussed above, may adversely affect product quality, stability, and efficacy. 15 Therefore, it is crucial to develop analytical methods to monitor polysorbate content and its degradants as part of the polysorbate analytical control strategy.…”