Discovery of 3-bromoflavone derivative D5 as a novel TLR4-MD2 antagonist for treatment of acute lung injury and sepsis

Abstract

The aberrant activation of immune responses mediated by Toll-like receptor 4 (TLR4) generates severe inflammatory syndromes such as acute lung injury (ALI) and sepsis, thus targeting TLR4 is attractive as a potential pharmacological therapy for these diseases. The aim of this study is to develop a novel structural TLR4 inhibitor for the treatment of ALI and sepsis. Herein, we designed and synthesized a new series of 3-bromoflavone derivatives by screening and incorporating molecular docking. Their anti-inflammatory effects were evaluated by ELISA in macrophages and compound D5 was identified as the most potential anti-inflammatory candidate. Compound D5 inhibited lipopolysaccharide (LPS)-induced expression of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in RAW264.7 cells with IC50 values of 0.215 and 4.211 µM, respectively. Immunoprecipitation, SPR, and immunoblotting demonstrated that D5 targeted TLR4-MD2 complex and competitively inhibited the binding of LPS to TLR4-MD2, resulting in inhibition of NF-κB and MAPK signaling pathways. In vivo administration of D5 significantly ameliorated various pathological indexes in LPS-induced ALI model and promoted the survival rate in LPS-induced septic model. Overall, our studies have identified a 3-bromoflavone derivative, D5, as a potent TLR4-MD2 antagonist and represented a very promising starting point for the development of anti-inflammatory agents for the treatment of ALI and sepsis.