Aggregates Removal in Monoclonal Antibody Purification
The presence of aggregates in therapeutic monoclonal antibody (mAb) drugs is a major concern because they can decrease product efficacy and cause an immunogenic response in patients.¹,² Presently, mAb aggregation is an increasing concern due the very high upstream solution titers that can be achieved. Drug manufacturers usually target less than 1% aggregates in the final drug formulation.¹
The aggregates, which are large clusters of irreversibly denatured antibodies, can form during cell culture, purification, or product storage.² The separation of soluble aggregates from the desired monomeric mAbs can be challenging due to their inherent physical and chemical similarities. However, since aggregates typically have higher surface charge or hydrophobicity compared to the monomer, ion-exchange or hydrophobic interaction chromatography (HIC) are effective tools for their separation.¹
When ion-exchange is employed, aggregates will bind to the column more strongly due to their higher charge, and in turn elute at higher conductivity compared to the monomer. Similarly, their increased hydrophobicity results in stronger column binding in HIC, and a lower concentration of salt will be required for elution compared to the monomer.
Multimodal membrane chromatography media, such as Purilogics Purexa™ MQ, can be a powerful tool for aggregates removal, since it can separate species based on both charge and hydrophobic interactions. Purexa™ MQ, differentiated from competing membrane products, has been shown to be effective in removal of aggregates from commercial mAb streams in a two-step chromatography scheme.³
References:
1. Evans, W. Removing Aggregates in Monoclonal Antibody Purification, Pharmaceutical Technology, 39, 3 (2015). http://www.pharmtech.com/removing-aggregates-monoclonal-antibody-purification
2. Joshi, V.; Yadav, N.; Rathore, R. S. Aggregation of Monoclonal Antibody Products: Formation and Removal, BioPharm International, 26, 3 (2013). https://www.biopharminternational.com/aggregation-monoclonal-antibody-products-formation-and-removal?id=&pageID=1&sk=&date=
3. Forsythe, A.; Henn, D.; Temples, G.; Zhou, J.; Husson, S.M. Application of novel multimodal anion-exchange membrane chromatography columns in a two-step mAb purification scheme – Aggregates and HCP removal. 257th ACS National Meeting, Orlando, FL (2019).