Synthesis, multi-target enzyme inhibition, and in silico evaluation of phthalimide derivatives with relevance to tuberculosis-associated pathophysiology.

Abstract

Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), remains a significant global health concern requiring innovative therapeutic approaches. Urease is a bacterial enzyme involved in nitrogen acquisition and phagosomal alkalinization in Mtb, whereas thymidine phosphorylase (TP), xanthine oxidase (XO), and butyrylcholinesterase (BChE) are host enzymes associated with nucleotide metabolism, oxidative stress, and cholinergic/immune regulation, respectively, and were included to explore enzyme-inhibitory profiles with potential relevance to tuberculosis-associated pathophysiology. In this study, a new series of phthalimide derivatives was synthesized and structurally characterized through 1H NMR and 13C NMR, with further optimization performed at the DFT level (B3LYP/6-31G). Enzyme inhibition assays demonstrated inhibitory activity across the four targets. Compound C15 emerged as the most effective multitarget enzyme inhibitor, showing strong activity against urease (IC₅₀ = 2.90 ± 0.11 μM), XO (IC₅₀ = 5.61 ± 0.14 μM), and BChE (IC₅₀ = 9.49 ± 0.18 μM), whereas compound C17 displayed the greatest potency towards TP (IC₅₀ = 3.24 ± 0.28 μM). UV-Visible spectrophotometric analysis was conducted to examine enzyme-ligand interactions, revealing concentration-dependent changes consistent with complex formation. Among the series, C17 exhibited the strongest binding affinity towards TP and urease, whereas C15 showed the strongest affinity towards XO. Structure-activity relationship analysis identified benzothiazole, halogen, nitro, sulphonamide, and carboxylic acid functionalities as important contributors to enzyme-inhibitory activity. Collectively, these findings identify C15 and C17 as promising lead compounds for further optimization as multitarget enzyme inhibitors.