Exploring the thymidine phosphorylase inhibitory potential of bis-Schiff derivatives bearing thiobarbituric acid: insights into in vitro and in silico approaches.

Thymidine phosphorylase (TP) is overexpressed in many cancers, promoting metastasis, angiogenesis, and tumor growth, making it a promising therapeutic target for anticancer drug development. In this study, bis-Schiff bases of thiobarbituric acid (3a-3g) were screened for in vitro TP inhibitory activity. In the series, four compounds 3c, 3d, 3b, and 3e exhibited potent inhibition with IC50 values ranging from (IC50 = 2.16 ± 1.94 µM) to (IC50 = 12.27 ± 1.78 µM). Density functional theory (DFT) calculations revealed that nitro-substituted derivatives possess the smallest HOMO and LUMO gaps (2.53-2.59 eV) and highest softness values (0.103-0.113 eV-1), indicating enhanced electronic reactivity. Molecular docking against TP enzyme showed strong binding affinities, with compound 3f achieving favorable docking score (-8.96 kcal/mol). Normal Mode Analysis (NMA) provided dynamic insights into the binding mechanisms, revealing that 3f-4EAD harnesses the intrinsic breathing motion of TP, while 3d-4LHM engages a distal allosteric network. A quantitative structure-activity relationship (QSAR) model (R2 = 0.91) was developed correlating quantum chemical descriptors with biological activity. The ADME/T analysis predicted acceptable pharmacokinetic profiles for the most active compounds. These preliminary findings identify 3c, 3d, and 3f as lead compounds warranting further pharmacological evaluation as TP-targeted anticancer agents.