Countercurrent preferential precipitation of acidic variants from monoclonal antibody pools

Abstract

The process of countercurrent multistage precipitation (CnMP) was developed for reduction of the acidic variant (av) content in monoclonal antibody (mAb) pools. The process was performed in polyethylene glycol (PEG) solutions of low ionic strength, at which av was the most prone to precipitate. To design the process, a mathematical model was formulated, which consisted of steady-state mass balance equations and underlying thermodynamic dependencies. The model was solved for different combinations of the process variables, including the protein concentration in the feed material and the PEG concentration in subsequent precipitation stages. The model solution was projected onto two-dimensional planes, where the performance indicators, such as the separation yield and the av reduction level, were correlated with the process variables. CnMP overperformed crosscurrent multistage precipitation (CsMP) and ion exchange chromatography (IEX) used in previous studies; in the comparison with CsMP, the 3-stage CnMP process allowed up to 28% increase in yield and up to 25% increase in the av reduction level, and in the comparison with IEX, up to 37% increase in yield and up 12.5% increase in the av reduction level. The yield benefit of CnMP depended on the composition of the feed material and the demand for the av reduction. The concept was experimentally verified; the 2- and 3-stage CnMP processes were performed using the input process variables provided by the model solution. The performance indicators calculated by the model and experimentally measured were in a good agreement, which indicated both feasibility and predictability of CnMP.