Integrative computational, synthetic, experimental evaluation of targeted inhibitors against matrix metalloproteinase-9: Toward precision modulation of proteolytic activity

Matrix metalloproteinase-9 (MMP-9) is a zinc-dependent enzyme that degrades the extracellular matrix and is involved in various diseases, including rheumatoid arthritis, atherosclerosis, tumor invasion, and metastasis. Despite the development of inhibitors, none have succeeded in trials. Our goal was to find potential inhibitors to regulate its proteolytic activity. Ligand- and structure-based drug design approaches were explored to identify inhibitors against wild-type (1GKC) and mutant (2OW1) MMP-9. A pharmacophore model was created, and drug-like molecules were prioritized to guide the development of benzamide and 1H-indole-2-carboxamide derivatives. These compounds were synthesized and characterized using ¹H NMR, 13C NMR, and HRMS (ESI). An experimental evaluation assessed their inhibitory potential and IC50 values against MMP-9. Most tested inhibitors fit the pharmacophore model, which consists of three aromatic/hydrophobic spheres and two hydrogen-bond donors/acceptors. Compounds 1, 2, 8, 10, 20, 21, 27, and 29 exhibited significant inhibition (P < 0.0001) of over 60%. Compounds 2 and 20 inhibited growth by over 70%, with IC50 values of 28.59 μM and 30.82 μM, respectively. The IF docking showed strong binding for these, with scores of -9.179 and -10.739 kcal/mol. The alignment between the computational approach and experimental validation reinforces the inhibitor's specificity and potency, confirms the docking model, and suggests that the predicted binding pose represents key biological interactions.