iNOS/PGE₂ inhibitors as a novel template for analgesic/anti‐inflammatory activity: Design, synthesis, in vitro biological activity and docking studies

Due to the serious gastrointestinal side effects associated with prolonged use of current anti-inflammatory therapies, various strategies such as the regulation of nitric oxide (NO) and prostaglandin E₂ (PGE₂) production have been explored in the field of anti-inflammatory drug development. In this study, a series of disubstituted 1,3,4-oxadiazoles (3a-f and 4a-f) and their cyclized 1,2,4-triazole derivatives (5a-e and 6a-e) were synthesized and tested for their NO, PGE₂, and interleukin-6 (IL-6) releasing inhibition ability. All of the compounds were observed to reduce lipopolysaccharide (LPS)-induced nitrite production in a concentration-dependent manner. Moreover, compounds 3b (50 μM) and 6d (1 μM) exhibited 63% and 49% inhibition, respectively, while indomethacin showed 52% at 100 μM. Based on a preliminary NO inhibition assay, 10 of the compounds (3a, 3b, 3e, 4b, 4d, 6a-e) were selected to be evaluated for in vitro PGE₂, IL-6, and inducible nitric oxide synthase (iNOS) inhibition. Notably, compound 6d proved to be the most active of the series with the lowest dose (1 µM), in comparison to the other further tested compounds (5-100 µM) and the reference drug indomethacin (100 µM). The inhibitory activity of the compounds was supported by docking simulations into the binding site of the iNOS protein receptor (Protein Data Bank [PDB]ID: 3E7G). The data showing that 4d reduced iNOS levels the most can be explained by the H-bond with Tyr347 through oxadiazole and π-halogen interactions through the p-bromo, in addition to aromatic interactions with protoporphyrin IX.