Abstract
For sugars to act as successful stabilizers of proteins during lyophilization and subsequent storage, they need to have several characteristics. One of them is that they need to be able to form interactions with the protein and for that miscibility is essential. To evaluate the influence of protein-sugar miscibility on protein storage stability, model protein IgG was lyophilized in the presence of various sugars of different molecular weight. By comparing solid-state nuclear magnetic resonance spectroscopy relaxation times of both protein and sugar on two different timescales, i.e., (1)H T1 and (1)H T1ρ, miscibility of the two components was established on a 2-5- and a 20-50-nm length scale, respectively, and related to protein storage stability. Smaller sugars showed better miscibility with IgG, and the tendency of IgG to aggregate during storage was lower for smaller sugars. The largest sugar performed worst and was phase separated on both length scales. Additionally, shorter protein (1)H T1 relaxation times correlated with higher aggregation rates during storage. The enzyme-linked immunosorbent assay (ELISA) assay showed overlapping effects of aggregation and chemical degradation and did not correspond as well with the miscibility. Because of the small scale at which miscibility was determined (2-5nm) and the size of the protein domains (∼2.5 × 2.5 × 5nm), the miscibility data give an indirect measure of interaction between protein and sugar. This reduced interaction could be the result of steric hindrance, providing a possible explanation as to why smaller sugars show better miscibility and storage stability with the protein.
Highlights
Protein pharmaceuticals have become increasingly important in the pharmaceutical industry over the past decade, with 10 out of the 41 drugs approved by the American Food and Drug Administration (FDA) in 2014 being proteins [1]
The samples which were reconstituted with 1 mL ultrapure water were diluted further to a concentration of 1 mg/mL immunoglobulin G (IgG) with 20 mM NaH2PO4 with 150 mM NaCl
Baseline corrections based on the signals between 6 and 7 and 26–27 min were carried out for the obtained chromatograms
Summary
Protein pharmaceuticals have become increasingly important in the pharmaceutical industry over the past decade, with 10 out of the 41 drugs approved by the American Food and Drug Administration (FDA) in 2014 being proteins [1].Electronic supplementary material The online version of this article (doi:10.1208/s12248-016-9937-7) contains supplementary material, which is available to authorized users.Storrs, Connecticut, USA. 6 To whom correspondence should be addressed. (e-mail: Most protein formulations are produced as solutions requiring refrigerated storage and handling, the so-called cold chain. The cold chain can be circumvented by drying the protein formulation in the presence of a stabilizer, using methods such as lyophilization and spray drying [2, 3]. For these situations, sugars are often used as stabilizers
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