Exploring tetrahydro-β-carboline derivatives as inhibitors of protein disulfide isomerases A1 and A3

Protein disulfide isomerases (PDIs) play a key role for reduction, oxidation, and isomerization of disulfide bonds within the endoplasmic reticulum. Among them, PDIA1 and PDIA3 have been implicated in diverse pathologies, emerging as promising therapeutic targets. Starting from the scaffold of the known PDI inhibitor 16F16, new molecules were designed, synthesized, and tested against PDIA1 and PDIA3. Based on insulin turbidity and protein thermal shift assays, compounds 2a and 2b showed preferential PDIA1 inhibition, while 2c and 2e inhibited both PDIA1 and PDIA3. Comparative proteomic experiments revealed that both 16F16 and 2e primarily target the first highly conserved Cys-Gly-His-Cys motif in PDIA1, while mainly engaging non-catalytic cysteines in PDIA3. Molecular modeling confirmed thiol reactivity in PDIA1 and PDIA3 and identified binding modes to the most involved protein cysteines. On this basis, here we have reported novel potent PDI inhibitors derived from 16F16 and demonstrated a distinct cysteine modification profile in PDIA1 and PDIA3.