Abstract
ObjectivesThe effect of different formulations variables on protein integrity were investigated using lysozyme as a model protein for the development of biotherapeutic protein formulations for use in the clinic.ResultsBuffer composition/concentration was the key variable of formulation reagents investigated in determining lysozyme stability and authenticity independent of protein concentration whilst the storage temperature and time, not surprisingly, were also key variables. Tryptic peptide mapping of the protein showed that the modifications occurred when formulated under specific conditions but not others. A model peptide system was developed that reflected the same behavior under formulation conditions as intact lysozyme.ConclusionsPeptide models may mirror the stability of proteins, or regions of proteins, in the same formulations and be used to help develop a rapid screen of formulations for stabilisation of biotherapeutic proteins.Electronic supplementary materialThe online version of this article (doi:10.1007/s10529-015-2014-y) contains supplementary material, which is available to authorized users.
Highlights
The number of biopharmaceutical protein-based drugs on the market and in development continues to increase with biopharmaceuticals making up a significant portion of new drugs in the development pipeline
Quantitative determination of lysozyme solubility and aggregation in different formulations All the formulation variables, concentrations and levels used in this study were based upon those reported in previous studies (Trikha et al 2002; Walsh 2006; Wang et al 2007)
The effect of formulation variables on lysozyme enzymatic activity Lysozyme initial rate activity was determined by measuring the OD500 of a suspension of the substrate Micrococcus lysodeikticus in the presence of lysozyme in each formulation (Fig. 1)
Summary
The number of biopharmaceutical protein-based drugs on the market and in development continues to increase with biopharmaceuticals making up a significant portion of new drugs in the development pipeline. In order for biopharmaceutical drugs to be successful in clinical applications, appropriate formulation(s) for preservation, stability and delivery need to be determined. This is not an easy task as each protein biopharmaceutical is unique and small differences in the amino acid residues result in the need of a specific formulation to deliver maximal stability and activity for each protein. The effect of different formulations variables on protein integrity were investigated using lysozyme as a model protein for the development of biotherapeutic protein formulations for use in the clinic
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