Kinetic models towards an enhanced understanding of diverse ADC conjugation reactions

The conjugation reaction is the central step in the manufacturing process of
antibody-drug conjugates (ADCs). This reaction generates a heterogeneous and
complex mixture of differently conjugated sub-species depending on the chosen
conjugation chemistry. The parametrization of the conjugation reaction through
mechanistic kinetic models offers a chance to enhance valuable reaction
knowledge and ensure process robustness. This study introduces a versatile
modeling framework for the conjugation reaction of cysteine-conjugated ADC
modalities—site-specific and interchain disulfide conjugation. Various
conjugation kinetics involving different maleimide-functionalized payloads
were performed, while controlled gradual payload feeding was employed to
decelerate the conjugation, facilitating a more detailed investigation of the
reaction mechanism. The kinetic data were analyzed with a reducing reversed
phase (RP) chromatography method, that can readily be implemented for the
accurate characterization of ADCs with diverse drug-to-antibody ratios,
providing the conjugation trajectories of the single chains of the monoclonal
antibody (mAb). ... mehrPossible kinetic models for the conjugation mechanism were
then developed and selected based on multiple criteria. When calibrating the
established model to kinetics involving different payloads, conjugation rates were
determined to be payload-specific. Further conclusions regarding the kinetic
comparability across the two modalities could also be derived. One calibrated
model was used for an exemplary in silico screening of the initial concentrations
offering valuable insights for profound understanding of the conjugation process
in ADC development.