(319d) Characterizing Self-Association and Clustering in Antibody Solutions at High Concentrations Using x-Ray Scattering, Rheology and Coarse Grained Simulation

Monoclonal antibodies (mAbs) are promising candidates for subcutaneous delivery at high concentration (100 to 300 mg/mL). However, short-range attractive protein-protein interactions (PPI) can lead to formation of clusters which can raise viscosity, lower stability, and lead to irreversible aggregate formation. Conventional low concentration characterization techniques are not predictive of undesirable behavior of mAbs at higher concentrations. In this study, we combine small angle x-ray scattering (SAXS) and microfluidic rheometry with coarse grained molecular dynamics (MD) simulations to characterize PPI and cluster formation in concentrated mAb solutions and investigate their impact on viscosity and colloidal stability. The simulations utilize 12-bead models with van der Waals attraction across all beads and additional, short-ranged attractions between specific beads. The resulting cluster size distribution and cluster properties are characterized as a function of strength of short-range attraction. Since our model captures key features of the protein shape, it can describe the experimental SAXS scattering profiles for solutions of 10-200 mg/mL protein with only a small (<1 k_BT) variation in the model’s attraction strength. The models are then compared to the SAXS profiles of mAbs under various solution conditions, including a range of ionic strengths, pH and cosolutes. Key indicators for cluster formation are identified from the scattering curves upon considering a wide range of q values and compared to viscosity measurements. Clustering in these systems is further probed by studying shear thinning behavior at various concentrations.