(320b) Quantifying Host Cell Protein Interactions with Antibodies Using Interaction Chromatography

Protein aggregation and impurities such as host cell proteins, HCPs, are critical problems 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. Similarly, the removal of host cell proteins and aggregates from the product is associated with very high costs leading to lengthy and costly manufacturing processes for biopharmaceuticals.

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). The osmotic second virial coefficient (B₂₂) is a fundamental physicochemical property that describes the protein-protein interactions in solution that has been linked to predict aggregation, protein phase behavior and solubility in previous research. Self-Interaction Chromatography (SIC) and Cross-Interaction Chromatography (CIC) have recently become popular tools to determine, B₂₂, and protein-protein interactions. These tools are based on quantifying the interactions between one protein that is immobilized on resins and packed into a column and another which is injected through the column (same protein for SIC and different protein for CIC).

SIC has shown successes in previous research predicting protein aggregation in downstream processing and bioprocess formulation. However, limited research using these tools have been done to study the interactions that exist between host cell proteins and purified proteins for us to better understand the interface between upstream and downstream operations and get a better prediction of aggregation early on in the process.

In this study we utilized CIC and characterized the interactions using cell culture samples containing different amounts of host cell proteins and purified mAbs. These interactions were then compared with specific interactions between two purified proteins. Early results displayed that samples with HCPs and other impurities displayed much stronger interactions and more negative virial coefficients than samples with less HCPs and impurities. Next phase will be to evaluate specific HCPs and HCP loads and the interactions associated with these. This will then be able to provide direct guidelines on difficult-to-remove HCPs and added mechanical insights that could aid future downstream process development.