Determining the Clearance of Degraded Protein via a Monoclonal Antibody Purification Process in Support of Cleaning Carryover Limits in Multiproduct Facilities

Abstract

Cleaning validation acceptance criteria in multiproduct facilities are established using maximum allowable carryover calculations. Carryover calculations incorporate the shared equipment surface area between two products to ensure that an acceptable limit for residue from the previously manufactured product to the subsequent product is determined. The shared surface area can be limited to areas where carryover presents the highest risk to product quality or patient safety. In these cases, specifically for biologic drug substance manufacturing, the shared surface area is limited to equipment after the purification process based on the assumption that the purification process would remove potential product fragment residues from the previous product. Until now, this assumption has been based on empirical knowledge without experimental data quantifying the clearance or removal of potential residues. We present a three-part study that determined the effects of cleaning conditions on selected monoclonal antibodies (mAbs) and the generation of degraded fragments and evaluated the clearance of both the degraded mAb1 in a laboratory setting and the degraded fragments in the presence of a subsequent product, assessing the risk of co-purification. Several analytical techniques were used, including gel electrophoresis, capillary zone electrophoresis/laser-induced florescence detection, and liquid chromatography-mass spectrometry. Protein fragment generation was demonstrated for five different mAbs from different immunoglobulin G subclasses. The clearance of the degraded fragments in the absence and presence of the subsequent product was demonstrated by calculating fold clearance and log reduction value (LRV) for each chromatography step. The data showed that the fragments generated during cleaning could be removed by the purification process. The fold clearances were determined to be values of 5400 (3.7 LRV) in the absence of subsequent product and 4428 (3.6 LRV) in the presence of subsequent product. The results supported the removal of product residues from shared surface areas by the purification process in multiproduct biologic drug substance manufacturing facilities.

Keywords: Carryover limits; Cleaning; Degraded protein clearance.