Pulse Proteolysis: An Orthogonal Tool for Protein Formulation Screening
| Autor: | Wenkui Lan, Rahul Phanse, Mark S. Bolgar, Mary Elizabeth Krause, Scott A. Hart, Meng Xu, Lavanya K. Iyer |
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| Rok vydání: | 2019 |
| Předmět: |
Circular dichroism
Protein Conformation Drug Compounding Recombinant Fusion Proteins Size-exclusion chromatography Pharmaceutical Science Excipient 02 engineering and technology 030226 pharmacology & pharmacy Fluorescence spectroscopy Excipients Protein Aggregates 03 medical and health sciences 0302 clinical medicine Protein structure Differential scanning calorimetry Dynamic light scattering medicine Thermal stability Protein Unfolding Chromatography Calorimetry Differential Scanning Protein Stability Chemistry Antibodies Monoclonal 021001 nanoscience & nanotechnology Proteolysis 0210 nano-technology medicine.drug |
| Zdroj: | Journal of Pharmaceutical Sciences. 108:842-850 |
| ISSN: | 0022-3549 |
| Popis: | Protein formulation stability is difficult to predict a priori and generally involves long-term stability studies. It is of interest to develop an analytical method that can predict stability trends reliably. Here, pulse proteolysis was evaluated as an analytical tool to predict solution-state stability in different formulations. Four proteins formulated in different buffer and excipient compositions were subjected to urea-induced unfolding and brief enzymatic digestion ("pulse" proteolysis), and relative resistance to proteolysis was measured by microfluidics-based capillary electrophoresis-sodium dodecyl sulfate. Biophysical properties of each formulation were measured using orthogonal biophysical techniques such as differential scanning fluorimetry, differential scanning calorimetry, dynamic light scattering, circular dichroism, and fluorescence spectroscopy. Protein stability in all formulations was monitored by size exclusion chromatography on storage at 5°C and 40°C. For all 4 proteins, formulations with greater proteolytic resistance also showed higher monomer content on thermal stability. In contrast, standard biophysical techniques showed reasonable-to-no correlation with size exclusion chromatography data. The data support the use of pulse proteolysis as an orthogonal, quantitative, and predictive tool to measure protein conformational stability and rank-order formulations. |
| Databáze: | OpenAIRE |
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