PROCESS DEVELOPMENT FOR ROBUST REMOVAL OF AGGREGATES USING CATION EXCHANGE CHROMATOGRAPHY IN MONOCLONAL ANTIBODY PURIFICATION WITH IMPLEMENTATION OF QUALITY BY DESIGN

Aggregate removal is one of the most important aspects in monoclonal antibody (mAb) purification. Cation-exchange chromatography (CEX), a widely used polishing step in mAb purification, is able to clear both process-related impurities and product-related impurities. In this study, with the implementation of quality by design (QbD), a process development approach for robust removal of aggregates using CEX is described. First, resin screening studies were performed and a suitable CEX resin was chosen because of its relatively better selectivity and higher dynamic binding capacity. Second, a pH-conductivity hybrid gradient elution method for the CEX was established, and the risk assessment for the process was carried out. Third, a process characterization study was used to evaluate the impact of the potentially important process parameters on the process performance with respect to aggregate removal. Accordingly, a process design space was established. Aggregate level in load is the critical parameter. Its operating range is set at 0-3% and the acceptable range is set at 0-5%. Equilibration buffer is the key parameter. Its operating range is set at 40 +/- 5mM acetate, pH 5.0 +/- 0.1, and acceptable range is set at 40 +/- 10 mM acetate, pH 5.0 +/- 0.2. Elution buffer, load mass, and gradient elution volume are non-key parameters; their operating ranges and acceptable ranges are equally set at 250 +/- 10 mM acetate, pH 6.0 +/- 0.2, 45 +/- 10 g/L resin, and 10 +/- 20% CV respectively. Finally, the process was scaled up 80 times and the impurities removal profiles were revealed. Three scaled-up runs showed that the size-exclusion chromatography (SEC) purity of the CEX pool was 99.8% or above and the step yield was above 92%, thereby proving that the process is both consistent and robust.