Antibody-Drug Conjugates Methods and Protocols

 This volume looks at key methodologies that are commonly used across antibody drug conjugates (ADCs) programs. The chapters in this book cover topics such as conjugations to endogenous cysteine residues; click chemistry conjugations; antibody conjugations via glycosyl remodeling; analysis of ADCs by native mass spectrometry; characterization of ADCs by capillary electrophoresis; LC/MS methods for studying lysosomal ADC catabolism; and determination of ADC concentration by ligand-binding assays. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

 

Cutting-edge and practical, Antibody-Drug Conjugates: Methods and Protocols is a valuable resource that aims to lower the “activation barrier” when undertaking a new discipline, and provides a “toolbox” for the next generation of ADC scientists.

An Overview of the Current ADC Discovery Landscape.- Pushing the Envelope: Advancement of ADCs Outside of Oncology.- Conjugations to Endogenous Cysteine Residues.- Site-Specific Conjugation to Cys-Engineered THIOMAB™ Antibodies.- Transglutaminase-Mediated Conjugations.- Click Chemistry Conjugations.- Utilizing Solid-Phase to Enable High-Throughput, Site-Specific Conjugation and Dual Labeled Antibody and Fab Conjugates.- Bridged Cysteine Conjugations.- Antibody Conjugations via Glycosyl Remodeling.- ADC Analysis by Hydrophobic Interaction Chromatography.- Two-Dimensional Liquid Chromatography Coupled to High Resolution Mass Spectrometry for the Analysis of ADCs.- Drug Loading and Distribution of ADCs after Reduction or IdeS Digestion and Reduction.- Analysis of ADCs by Native Mass Spectrometry.- High Resolution Characterization of ADCs by Orbitrap LCMS.- Conjugation Site Analysis by MS/MS Protein Sequencing.- Conjugation Site Analysis of Lysine-Conjugated ADCs.- Characterization of ADCs by Capillary Electrophoresis.- Characterization of the Primary Structure of Cysteine-Linked Antibody-Drug Conjugates using Capillary Electrophoresis with Mass Spectrometry.- Purification of ADCs by Hydrophobic Interaction Chromatography.- Detection and Removal of Small-Molecule and Endotoxin Contaminants in ADC Preparations.- Physical Stability Studies of Antibody Drug Conjugates (ADCs) Under Stressed Conditions.- Biophysical Methods for Characterization of Antibody Drug Conjugates.- Determination of ADC Cytotoxicity in Immortalized Human Cell Lines.- LC/MS Methods for Studying Lysosomal ADC Catabolism.- Assessing ADC Plasma Stability by LC/MS Methods.- Determination of ADC Concentration by Ligand-Binding Assays.