QC Assay Development

Analytical methodology is one of the key components of antibody-drug conjugate (ADC) development for clinical trials and commercialization. This chapter focuses on the release, stability, and characterization methods and product attributes with special considerations and challenges for developing assays designed for ADCs. Characterization of drug loading includes assays to measure average drug- to-antibody ratio (DAR), drug loading distribution, drug loading variants, and positional isomers. DAR is determined by using the following analytical methods: UV spectrophotometry, electrospray ionization mass spectrometry (MS), hydrophobic interaction chromatography (HIC), and reversed-phased high-performance liquid chromatography (RP-HPLC). Drug loading distribution has been commonly used as a product characterization tool. The positional isomer measures the conjugation site on the antibody for cysteine-linked ADCs. Drug-related substances are the unconjugated drug-related impurities in the ADC sample. Drugs used in ADCs are usually too small to elicit an immune response by themselves.