Sample displacement chromatography of monoclonal antibody charge variants and aggregates.

Abstract

The rise of biosimilar monoclonal antibodies has renewed the interest in monoclonal antibody (mAb) charge variants composition and separation. The sample displacement chromatography (SDC) has the potential to overcome the low separation efficiency and productivity associated with bind-elute separation of mAb charge variants. SDC in combination with weak cation exchanging macroporous monolithic chromatographic column was successfully implemented for a separation of charge variants and aggregates of monoclonal IgG under overloading conditions. The charge variants composition was at-line monitored by a newly developed, simple and fast analytical method, based on weak cation exchange chromatography. It was proven that basic charge variants acted as displacers of IgG molecules with lower pI, when the loading was performed 1 to 1.5pH unit below the pI of acidic charge variants. The efficiency of the SDC process is flow rate independent due to a convection-based mass transfer on the macroporous monolith. The productivity of the process at optimal conditions is 35mg of purified IgG fraction per milliliters of monolithic support with 75-80% recovery. As such, an SDC approach surpasses the standard bind-elute separation in the productivity for a factor of 3, when performed on the same column. The applicability of the SDC approach was confirmed for porous particle-based column as well, but with 1.5 lower productivity compared to the monoliths.