Design and synthesis of 1-Phenyl-2-thioaryl-4-benzyl-5-methylimidazoles as dual STAT3/NF-κB inhibitors for triple-negative breast Cancer therapy

Triple-negative breast cancer (TNBC) lacks targeted therapies due to the absence of hormone receptors. The oncogenic crosstalk between STAT3 and NF-κB pathways drives tumor proliferation, epithelial-mesenchymal transition (EMT), metastasis, and therapeutic resistance, underscoring a critical need for novel dual inhibitors. Here, we report the rational design and synthesis of new 1-phenyl-2-thioaryl-4-benzyl-5-methylimidazole derivatives targeting this dual pathway axis. Optimized, metal-free S-alkylation reactions of imidazole-2-thione afforded twelve new analogs (5a,b, 6, 7a-g, 8a,b) in excellent yields. Their antiproliferative activity was evaluated in 4 T1 and MDA-MB-231 cells, while the mechanistic leads (5a,b and 8a,b) were assessed for specific STAT3/NF-κB inhibition using luciferase reporter assays together with migration and invasion assays. Structure-activity relationship (SAR) analysis identified the difluorobenzylthio derivative 5b as the optimal lead candidate. At a sub-cytotoxic concentration (10 μM), 5b exhibited potent and specific targeted blockade, suppressing NF-κB and STAT3 transcriptional activities by ∼80% and ∼ 60%, respectively. In contrast, structurally related analogs (5a, 8a, 8b) only exhibited inhibitory effects that were inextricably linked to severe cytotoxicity. Furthermore, 5b demonstrated robust antimetastatic efficacy at non-toxic doses, maintaining >91% wound persistence in 4 T1 cells at 24 h and markedly blocking Matrigel invasion. Molecular docking studies at the STAT3-SH2 domain corroborated the biological data, confirming that the flexibility of the thioether linkage in the 5a and 5b provided optimal binding energies compared to the more rigid or bulkier inactive analogs. This newly accessible imidazole class successfully establishes dual STAT3/NF-κB inhibition with direct antimetastatic efficacy, positioning 5b as a highly promising TNBC lead warranting further in vivo validation and clinical translation.