Design,Synthesis,and Anti-inflammatory Evaluation Of Inhibitors Targeting Toll-like Receptor 4 Signaling

Acute lung injury(ALI)is often caused by systemic inflammatory responses,resulting in extremely high mortality rate.Current pharmacologic treatments of ALI are disappointing.Toll-like receptor(TLR)superfamily play a fundamental role in host defense and inflammation.Among all these TLRs,TLR4 together with its accessory protein myeloid differentiation protein 2(MD2)are the most studied receptor so far.MD2-TLR4 is an essential complex which recognizes lipopolysaccharide(LPS),leading to initiation of inflammation through the activation of TLR4 signaling.Upon activation TLR4 recuited myeloid differentiation primary response protein 88(My D88),eventually leading to activation of nuclear factor-kappa B(NF-?B),activator protein-1(AP-1)and proinflammatory cytokine production.Numerus evidence indicated that targeting the MD2-TLR4-My D88 complex signaling may be a promising way to treat acute inflammatory disorders.Caffeic acid phenethyl ester(CAPE)from propolis of honeybee hives could interfere interactions between LPS and the TLR4-MD2 complex,and thereby has promising anti-inflammatory properties.In chapter two,we designed and synthesized48 CAPE derivatives and evaluated their anti-inflammatory activities in mouse primary peritoneal macrophages(MPMs)activated by LPS.The most active compound,5s,was found to bind with MD2 with high affinity,thereby prevented formation of the LPS-MD2-TLR4 complex.The binding mode of 5s revealed that the major interactions with MD2 were established via a key hydrogen bonds and hydrophobic interactions.Furthermore,5s showed remarkable protective effects against LPS-caused ALI in vivo.Indole and indazole have wide biological applications and they are potentially valuable as building-blocks in medicinal chemistry.In chapter three,we report the design and synthesis of a new series of 3-(indol-5-yl)-indazoles,which were evaluated for their anti-inflammatory activities in MPMs.Among the analogues generated,the promising 3-(indol-5-yl)-indazole analogue 13m inhibited lipopolysaccharide(LPS)-induced expression of tumor necrosis factor alpha(TNF-?)and interleukin-6(IL-6)in MPMs with IC₅₀ values of 0.89 and 0.53?M,respectively.Subsequently,compound13m was then identified as an MD2-TLR4 antagonist in suppressing LPS-induced inflammatory responses.In vivo administration of 13m significantly inhibited macrophage infiltration and ameliorated histopathological changes in lung tissues of LPS-challenged mice.My D88 is an essential adapter protein used by TLRs,is a promising target molecule for the treatment of respiratory inflammatory diseases.Previous studies reported a My D88 activities of novel 2-amino-4-phenylthiazole analogue(TJ-M2010-5)in inflammation-induced cancer,and identified the analogue as an inhibitor of My D88 toll/interleukin-1 receptor(TIR)homology domain homo-oligomerization.In chapter four,we described the synthesis of 47 new analogues by modifying different sites on this lead compound and assessed their anti-inflammatory activities in LPS-induced MPMs.The most promising compound,24d,was found to effectively interact with My D88 protein and prevented formation of the Myddosome complex.Furthermore,24d showed in vivo anti-inflammatory activity in LPS-caused model of acute lung injury.Taken together,our studies have identified new inhibitors targeting MD2-TLR4-My D88 signaling and lay the groundwork for future drug development of anti-inflammatory agents for the treatment of ALI.