Structure−Guided Optimization of Niclosamide Derivatives as Direct STAT3 Inhibitors Targeting a Novel Binding Site

Signal transducer and activator of transcription 3 (STAT3), a key oncogenic driver activated in approximately 70% of cancers, represents a highly attractive therapeutic target. Although various inhibitors of the STAT3 pathway have been developed, none directly target the STAT3 protein, and no FDA-approved STAT3 inhibitors are currently available. In this study, we identified a novel binding site for niclosamide on STAT3 using X-ray crystallography, revealing unique interactions spanning the coiled-coil, DNA-binding, and linker domains. Niclosamide was shown to disrupt STAT3-DNA binding with an IC₅₀ value of 219 ± 43.4 μM, as determined by fluorescence polarization (FP) assays. We synthesized a library of over 45 niclosamide analogues. Among this expanded set of compounds, MQ021 inhibited STAT3-DNA binding by 60% at a concentration of 100 μM, comparable to the 40% inhibition observed with niclosamide. Microscale thermophoresis (MST) assays confirmed the comparable activity of the compound 21 (Kd = 155 ± 59 μM) relative to niclosamide (Kd = 281 ± 55 μM). These results offer insights into the structure–activity relationships (SAR) of niclosamide derivatives and provide a foundation for the development of STAT3-targeted therapeutics for cancer treatment that exploit this new binding site.