title: Identification of an IgG CDR sequence contributing to co-purification of the host cell protease cathepsin D.
abstract: Recombinant therapeutic monoclonal antibodies (mAbs) must be purified from host cell proteins (HCPs), DNA, and other impurities present in Chinese hamster ovary (CHO) cell culture media. HCPs can potentially result in adverse clinical responses in patients and, in specific cases, have caused degradation of the final mAb product. As reported previously, residual traces of cathepsin D caused particle formation in the final product of mAb-1. The current work was focused on identification of a primary sequence in mAb-1 responsible for the binding and consequent co-purification of trace levels of CHO cathepsin D. Surface plasmon resonance (SPR) was used to detect binding between immobilized CHO cathepsin D and a panel of mAbs. Out of 13 mAbs tested, only mAb-1 and mAb-6 bound to cathepsin D. An LYY motif in the HC CDR2 was common, yet unique, to only these two mAbs. Mutation of LYY to AAA eliminated binding of mAb-1 to cathepsin D providing confirmation that this sequence motif was involved in the binding to CHO cathepsin D. Interestingly, the binding between mAb-1 and cathepsin D was weaker than that of mAb-6, which may be related to the fact that two aspartic acid residues near the LYY motif in mAb-1 are replaced with neutral serine residues in mAb-6. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:140-145, 2017.
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: Jared S
authors: 0000-0002-4391-0560
authors: JS
authors: Bee
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: LeeAnn M
authors: LM
authors: Machiesky
authors: Antibody Discovery and Protein Engineering, MedImmune, Gaithersburg, MD, 20878.
authors: Li
authors: L
authors: Peng
authors: Purification Process Sciences, MedImmune, Gaithersburg, MD, 20878.
authors: Kristin C
authors: KC
authors: Jusino
authors: Purification Process Sciences, MedImmune, Gaithersburg, MD, 20878.
authors: Matthew
authors: M
authors: Dickson
authors: Purification Process Sciences, MedImmune, Gaithersburg, MD, 20878.
authors: Jeffrey
authors: J
authors: Gill
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: Douglas
authors: D
authors: Johnson
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: Hung-Yu
authors: HY
authors: Lin
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: Kenneth
authors: K
authors: Miller
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: Jenny
authors: J
authors: Heidbrink Thompson
authors: Analytical Sciences, MedImmune, One MedImmune Way, Gaithersburg, MD, 20878.
authors: Richard L
authors: RL
authors: Remmele
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000302:isolation & purification*; D016466:CHO Cells; D002402:Cathepsin D / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D003412:Cricetulus; D003470:Culture Media / Q000737:chemistry*; D055767:Immobilized Proteins / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D000911:Antibodies, Monoclonal; D003470:Culture Media; D055767:Immobilized Proteins; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D002402:Cathepsin D
keywords: aspartyl protease; cathepsin D; host cell protein; monoclonal antibody; surface plasmon resonance