(200j) Development of a Cation Exchange Chromatography Step for Robust Impurity Clearance and Improved Polysorbate 80 Stability in the Drug Product of a Monoclonal Antibody

It has been reported that polysorbate 80 (PS80) degradation not only occurs due to auto-oxidation, but also due to lipases generated by the host organism. Specific lipases linked to PS80 degradation, such as phospholipase B-like 2 (PLBL2) and lipoprotein lipase (LPL), are difficult to remove in the downstream purification process, causing concerns for PS80 stability in the drug product (DP) as well as potential immunogenicityin humans.

A case study is presented on the development of a cation exchange chromatography (CEX) step for mAb A. After adding CEX as the second polishing step in the purification process, additional lipase clearance was achieved and PS80 stability in the DP improved. To further characterize the CEX step, a design of experiment (DOE) was conducted which elucidated critical step parameters and impurity clearance robustness. The DOE was a split plot central composite design involving 40 column runs operating in bind-and-elute mode. Five CEX parameters were studied: pH and conductivity of the elution buffer and load as well as resin loading. All 40 runs were evaluated for step yield and product quality. Results from HCP ELISA, UP-SEC, and HP-IEX analysis indicated high statistical dependence on elution buffer pH and conductivity (p<0.001). Overall, as elution buffer pH and conductivity increased, impurity clearance decreased and yield increased. LC-MS results indicated non-detectable lipases levels in the CEX pools for all runs in the elution buffer design space studied. These results suggest CEX is a robust process step to remove lipases, and hence, improve PS80 stability in the DP.