Understanding the mechanisms and prevention of aggregation of biopharmaceutic proteins

Biopharmaceutic proteins, especially monoclonal antibodies, are the fastest growing class of pharmaceutics globally, due to their high affinity, specificity and non-toxicity. Despite their versatility and wide applicability, their production is costly and poses unique challenges due to their high molecular weight and structural complexity as well as from instabilities like aggregation that lead to loss of quality and efficacy, limit their shelf life and can induce life-threatening immunogenic response. Protein aggregation is thus considered one of the major challenges in biopharmaceutic protein formulation with various strategies being employed to characterise protein aggregation and prevent it. Protein aggregation occurs through association of aggregation prone regions (APR) which can be located on protein surface or that get exposed upon partial unfolding of the proteins. These regions are experimentally difficult to detect and isolate due to their transient nature therefore high-resolution structural insight into APR composition remains elusive. Motivated by recent advances showing that these APRs can be detected by peptide array screening and that results of such screening correlate with aggregation propensity of proteins, we propose to develop a peptide library that will be used for a high-throughput nuclear magnetic resonance (NMR) screening that will provide structural insight into aggregation prone regions of native as well as partially unfolded protein species. NMR screening will be complemented with biomolecular simulations and traditional biophysical methods for characterisation of protein aggregation. We will also assess how small molecule excipients that bind to APRs influence the protein-protein interactions and mitigate protein aggregation. This project will contribute to understanding of molecular basis of protein self-association and aggregation which is crucial for rational design of more stable biopharmaceutic proteins in the future, that will reduce waste and significantly lower their production cost making them more accessible to all patients.