Synthesis of Fused Benzoxazole-Imidazo[1,2-c][1,2,3]triazole Hybrids in PEG-400 and Evaluation of Their Anticancer Activity In Vitro and In Silico

Objective: This work aimed to design, synthesize, and biologically evaluate a novel series of fused benzoxazole-imidazo[1,2-c][1,2,3]triazole derivatives. Methods: The target compounds Va–Vo were synthesized via a one-pot, copper-catalyzed cyclization in polyethylene glycol-400 (PEG-400). Their in vitro cytotoxicity was evaluated against the human breast adenocarcinoma cell lines MCF-7 and MDA-MB-231 using the MTT assay. Erlotinib was used as the reference inhibitor. The most active compounds were subjected to molecular docking studies against the epidermal growth factor receptor (EGFR) kinase domain (PDB ID: 4HJO) to elucidate their potential binding mode. Results and Discussion: All synthesized derivatives were characterized by ESI-HRMS, 1H, and 13C NMR spectroscopy. Compounds Vf, Vh, Vk, and Vl displayed significant cytotoxic activity with IC50 values ranging from 4.28 ± 0.53 to 7.44 ± 0.72 µM against both cancer cell lines, showing potency superior to that of erlotinib (IC50 = 4.68– 7.29 µM). Furthermore, these compounds exhibited markedly reduced cytotoxicity towards the non-tumorigenic mammary epithelial cell line MCF-10A, indicating a selective anticancer profile. Docking simulations revealed that the active compounds favorably bind to the EGFR ATP-binding site, with calculated free energies of binding ranging from –9.13 to –8.36 kcal/mol, which correlate with their in vitro potency and exceed the docking score of erlotinib (–7.69 kcal/mol). Conclusions: A new class of benzoxazole-imidazotriazole hybrids was successfully developed using an eco-friendly synthetic protocol. Several derivatives demonstrated potent and selective anticancer activity against breast cancer cell lines, likely mediated through EGFR inhibition, as supported by computational studies. Derivative Vk, with the highest activity and selectivity, is identified as a promising lead for further development.