Pyrimidine Triazole Thioether Derivatives as Toll‐Like Receptor 5 (TLR5)/Flagellin Complex Inhibitors

Abstract Protein–protein interactions have been regarded as “undruggable” despite their importance in many biological processes. The complex formed between host toll‐like receptor 5 (TLR5) and flagellin, a globular protein that is the main component of a bacterial flagellum, plays a vital role in a number of pathogen defenses, immunological diseases and cancers. Through high‐throughput screening, we identified two hits with a common pharmacophore, which were used to successfully develop a series of small‐molecule probes as novel inhibitors of flagellin binding to TLR5. In a multitude of assays, 4‐((4‐benzyl‐5‐(pyridin4yl)‐4 H ‐1,2,4‐triazol‐3‐yl)thio)pyrido[3′,2′:4,5]thieno[3,2‐ d ]pyrimidine (TH1020) was identified as a potent antagonist of TLR5 signaling with promising activity (IC 50 =0.85±0.12 μ m ) and specificity. Furthermore, TH1020 was shown to repress the expression of downstream TNF‐α signaling pathways mediated by the TLR5/flagellin complex formation. Based on molecular docking simulation, TH1020 is suggested to compete with flagellin and disrupt its association with TLR5. TH1020 provides a much‐needed molecular probe for studying this important protein–protein interaction and a lead compound for identifying novel therapeutics targeting TLR5.