Synthesis and discovery of triazolo-pyridazine-6-yl-substituted piperazines as effective anti-diabetic drugs; evaluated over dipeptidyl peptidase-4 inhibition mechanism and insulinotropic activities

A family of 12 triazolo-pyridazine-6-yl-substituted piperazines (5a-l) was synthesized and evaluated for their Dipeptidyl peptidase-4 (DPP-4) inhibition potentials in order to develop them as anti-diabetic medications. In the two-step synthesis process, 6-chloro-3-(m-tolyl)-[1,2,4]triazolo[4,3-b]pyridazine was synthesized with one-pot mode using pyridine, 3,6-dichloropyridazine 5-(3-methyl-phenyl)tetrazole in toluene. Conjugating corresponding 2° amines with 6-chloro-3-(m-tolyl)-[1,2,4]triazolo[4,3-b]pyridazine afforded the target triazolo-pyridazine-6-yl-substituted piperazines (5a-l). DPP-4 inhibition potential of these compounds was testified in silico and in nitro along with their insulinotropic activities in 832/13 INS-1 cells. H₂O₂ radical scavenging assay and MTT assay were conducted to assess the antioxidant and cytotoxicity of these compounds respectively. Molecular docking and ELISA based Enzyme inhibition assay results revealed the strong inhibition potential of the target compounds. MTT assay results indicated a maximum dose of 2.5 nM (IC₅₀ 1.25 nM) could be used and above this level vital for the cells. Compounds 5a, 5c, 5g and 5i were found with excellent antioxidant and insulinotropic activity up to 99%.