Development and application of self-interaction chromatography as a rational approach to measuring the osmotic second virial coefficient (B) for protein formulation development

Abstract

Formulation development is an integral step in the successful commercialization of protein-based products, both in the biotechnology and pharmaceutical industries. As the number and complexity of these protein formulations increases, so does the need for innovative approaches to characterize physical product stability. This dissertation demonstrates the development and application of self-interaction chromatography (SIC) for measuring the osmotic second virial coefficient (B) of proteins in the presence of biologically and industrially relevant mixtures of cosolvents including salts, sugars, amino acids, and polyols, and as a function of temperature and pH. Early studies focused on model proteins in very controlled systems. The application of SIC was then advanced to demonstrate rational formulation screening of three industrially relevant enzymes. SIC measured trends in B demonstrated excellent agreement with available B data from static light scattering (SLS) studies. In the absence of B measurements from other characterization approaches, SIC measured trends in B were correlated with trends in solubility and enzymatic activity. The outcome of the work presented in this dissertation suggests that SIC is a useful means to augment current approaches to characterizing the physical stability of protein-based formulations. I believe these efforts to develop a better fundamental understanding and characterization approach of protein self-interaction in complex cosolvent systems will ultimately lead to direct improvements in macromolecular formulation chemistry and structural biology endeavors.