title: Identification and characterization of host cell protein product-associated impurities in monoclonal antibody bioprocessing.
abstract: Downstream processing of monoclonal antibodies (mAbs) has evolved to allow the specific process for a new product to be developed largely by empirical specialization of a platform process that enables removal of impurities of different kinds. A more complete characterization of impurities and the product itself would provide insights into the rational design of efficient downstream processes. This work identifies and characterizes host cell protein (HCP) product-associated impurities, that is, HCP species carried through the downstream processes via direct interactions with the mAb. Interactions between HCPs and mAbs are characterized using cross-interaction chromatography under solution conditions typical of those used in downstream processing. The interacting species are then identified by two-dimensional gel electrophoresis and mass spectrometry. This methodology has been applied to identify product-associated impurities in one particular purification step, namely protein A affinity chromatography, for four therapeutic mAbs as well as the Fab and Fc domains of one of these mAbs. The results show both the differences in HCP-mAb interactions among different mAbs, and the relative importance of product association compared to co-elution in protein A affinity chromatography.
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of Delaware, Newark, Delaware, 19716.
authors.forename: Nicholas E
authors.initials: NE
authors.lastname: Levy
authors.forename: Kristin N
authors.initials: KN
authors.lastname: Valente
authors.forename: Leila H
authors.initials: LH
authors.lastname: Choe
authors.forename: Kelvin H
authors.initials: KH
authors.lastname: Lee
authors.forename: Abraham M
authors.initials: AM
authors.lastname: Lenhoff
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D016466:CHO Cells; D002846:Chromatography, Affinity / Q000379:methods*; D006224:Cricetinae; D003412:Cricetulus; D015180:Electrophoresis, Gel, Two-Dimensional; D011506:Proteins / Q000032:analysis* / Q000737:chemistry*; D011994:Recombinant Proteins / Q000302:isolation & purification*
chemical_list: D000911:Antibodies, Monoclonal; D011506:Proteins; D011994:Recombinant Proteins
keywords: host cell protein impurities; mAb process development; protein A affinity chromatography; protein-protein interactions