Deconjugation of thio-succinimide-conjugated linker-payloads can result from two possible reactions: 1) retro-Michael elimination at the conjugation site, exposing the free thiol; and 2) step-wise linker cleavage, generating a series of antibody backbone species with partial linker moieties. In evaluating ADCs 1-4, only species consistent with reaction 1 deconjugation were observed. Therefore, the deconjugation and thio-succinimide hydrolysis processes for ADCs 1-4 are two competing reactions. There is a clear structure-stability relationship and in vivo biotransformation reaction preference observed between the ADCs with propionyl-PEG8 vs. caproyl spacer within the linker. This provides mechanistic basis for improved in vivo DAR stability for ADC1 (AZD8205) and ADC4 versus ADC2 and ADC3.