Mapping the Aggregation Behaviour of Biopharmaceuticals: A New Approach

Mapping the Aggregation Behaviour of Biopharmaceuticals: A New Approach

S. Hedberg, J. Heng, D. Williams                       

                                               

ABSTRACT

Protein aggregation is a critical problem for the safety of biopharmaceuticals as they are linked to adverse immunologically related responses in patients. Much effort has been made to gain a better understanding of aggregation, however, the mechanisms leading to protein aggregation are still not fully understood. Protein-protein molecular interactions in solution are known to be involved in protein solution aggregation behaviour and are a common issue for the manufacturing of biopharmaceuticals such as monoclonal antibodies, mAbs.  Therefore, a major industrial and academic challenge is the development of fast and reliable methods, either theoretically or experimentally driven, which allow formulation scientists to identify optimal biopharmaceutical species and solution conditions which deliver stable solution products, thus avoiding aggregation.

In this work we will describe a SIC (self-interaction chromatography) method for determining the osmotic second virial coefficient (B₂₂), where B₂₂ describes the thermodynamics of protein-protein interactions in solution. This experimental biophysical approach allows solution stability maps including solution pH and salt concentration effects to be obtained using mg quantities of species such as mAbs. The approach has been validated against traditional size-exclusion chromatographic methods on samples that were aged over many weeks at elevated temperature. Both methods gave similar stability condition maps, with the SIC method delivering results within days compared with traditional stability trials which would typically take many months.  Over a wide range of test conditions good correlations were found between experimental B₂₂ values and experimentally measured aggregation rates.

Similarly, the conformational and structural changes were investigated for a number of selected conditions over a longer time at continuously increasing temperature. The results proved that SIC easily could predict the conformational changes of mAbs too. From this study it was concluded that improvement in the SIC methods reported here allows accurate and robust mapping of solution stability conditions as well as a good prediction of conformational stability.