Characterization of intact antibody-drug conjugates from plasma/serum in vivo by affinity capture capillary liquid chromatography-mass spectrometry |
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Authors: | Xu Keyang Liu Luna Saad Ola M Baudys Jakub Williams Lara Leipold Douglas Shen Ben Raab Helga Junutula Jagath R Kim Amy Kaur Surinder |
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Institution: | aGenentech, South San Francisco, CA 94080, USA;bEmergency Response and Air Toxicant (ERAT) Branch, Centers for Disease Control and Prevention (CDC), Atlanta, GA 30341, USA;cGrays Harbor College, Aberdeen, WA 98520, USA |
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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. |
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Keywords: | Antibody&ndash drug conjugate THIOMAB&ndash drug conjugate Drug loss Drug-to-antibody ratio Affinity capture Capillary liquid chromatography Mass spectrometry Electrospray ionization |
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