title: Characterization of intact antibody-drug conjugates from plasma/serum in vivo by affinity capture capillary liquid chromatography-mass spectrometry.
abstract: Antibody-drug conjugates (ADCs) are designed to facilitate the targeted delivery of cytotoxic drugs to improve their tumor fighting effects and minimize systemic toxicity. However, efficacy and safety can potentially be compromised due to the release of conjugated drugs from the ADC with time while in circulation, resulting in changes in the drug-to-antibody ratio (DAR). Current understanding of this process is limited because existing methods such as immunoassays fail to distinguish ADCs with different DARs. Here we demonstrate a novel method with bead-based affinity capture and capillary liquid chromatography-mass spectrometry to allow direct measurement of drug release by quantifying DAR distributions of the ADC in plasma/serum. This method successfully identified individual intact conjugated antibody species produced due to drug loss from ADCs (e.g., an engineered site-specific anti-MUC16 THIOMAB-drug conjugate) and measured the corresponding DAR distributions in vitro and in vivo. Information obtained can provide insights into the mechanisms involved in drug loss and help to optimize ADC therapeutics. Other potential applications of the method may include characterization of posttranslational modifications, protein adducts, and immunogenicity.
authors.affiliation: Genentech, South San Francisco, CA 94080, USA. xu.keyang@gene.com
authors.forename: Keyang
authors.initials: K
authors.lastname: Xu
authors.forename: Luna
authors.initials: L
authors.lastname: Liu
authors.forename: Ola M
authors.initials: OM
authors.lastname: Saad
authors.forename: Jakub
authors.initials: J
authors.lastname: Baudys
authors.forename: Lara
authors.initials: L
authors.lastname: Williams
authors.forename: Douglas
authors.initials: D
authors.lastname: Leipold
authors.forename: Ben
authors.initials: B
authors.lastname: Shen
authors.forename: Helga
authors.initials: H
authors.lastname: Raab
authors.forename: Jagath R
authors.initials: JR
authors.lastname: Junutula
authors.forename: Amy
authors.initials: A
authors.lastname: Kim
authors.forename: Surinder
authors.initials: S
authors.lastname: Kaur
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000097:blood* / Q000737:chemistry; D019921:Biotinylation; D018394:CA-125 Antigen / Q000276:immunology; D002846:Chromatography, Affinity / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods*; D005260:Female; D000882:Haplorhini; D006801:Humans; D018796:Immunoconjugates / Q000097:blood* / Q000737:chemistry / Q000302:isolation & purification; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008565:Membrane Proteins / Q000276:immunology; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry; D004364:Pharmaceutical Preparations / Q000097:blood* / Q000737:chemistry; D051381:Rats
chemical_list: D000911:Antibodies, Monoclonal; D018394:CA-125 Antigen; D018796:Immunoconjugates; C451759:MUC16 protein, human; D008565:Membrane Proteins; D009842:Oligopeptides; D004364:Pharmaceutical Preparations; C495575:monomethyl auristatin E