Discovery of novel seven-membered ring derivatives of ONC212 as caseinolytic protease P protein activators using the ring expansion strategy: Rational design, synthesis, and antibacterial evaluation.

Caseinolytic protease P (ClpP) protein is essential for prokaryotic and eukaryotic protein homeostasis, and has emerged as a promising bactericidal target for controlling bacterial infection and evading the emergent of drug-resistance risk. As the unique bactericidal mechanism, ClpP loses its substrate specificity and undergoes uncontrolled protein hydrolysis in the presence of an activator, leading to causing bacterial death. To further expand the chemotype of ClpP activator, a series of ONC212 derivatives were synthesized using a ring expansion strategy. Surprisingly, compound A14 showed optimal antibacterial activity against Xanthomonas oryzae pv. oryzae (Xoo) in vitro (EC50 value = 3.86 μg/mL), which was >6 × more potent than the EC50 value of ONC212 (26.42 μg/mL). Molecular docking, fluorescence titration and degradation experiments of the XooFtsZ protein indicated that compound A14 was a strong degrader of XooClpP towards XooFtsZ. In vivo assays showed that the control activity of compound A14 (200 μg/mL) reached 47.47 %, compared to 41.57 %, 36.72 %, and 30.43 % for ONC212, thiodiazole copper, and bismerthiazol, respectively. Overall, this study led to the identification compound A14, which not only showed improved antibacterial potency, and maintained the binding XooClpP, but also highlighted the ring expansion strategy as a promising approach for bactericide discovery.