Understanding drug metabolism is a critical component for successful drug development . Complex macromolecules such as ADCs present unique challenges to gain such understanding. Therefore, we developed and employed several analytical approaches to profile ADC biotransformation in circulation comprehensively. The results help to better understand factors affecting the underlying pharmacokinetic profiles of the various molecular species which are formed when AZD8205 is administered in vivo, and thereby aid in developing better understanding of SAR for thio-succinimidelinked ADCs. In the future, these findings could better inform translation of PK/PD from animal models to the clinical setting. Biotransformation profiling of protein conjugates can be further studied in patient populations, or in specific organ/tissue with this or similar method.