以姜黄素类似物为先导的MD2抑制剂药物设计及其对脓毒血症的药理作用研究

Sepsis, a general acute inflammatory syndrome, is mainly caused by infection and endotoxin. Toll-like receptor 4 / myeloid Differentiation-2 complex is the main receptor of endotoxin LPS and mediates the LPS-induced inflammatory response. Although the evidence showed that MD2 gene knockout attenuated the response to LPS, MD2, as a accessory protein of TLR4 signaling, is always in a secondary research position, and there are a few reports on the drug design and pharmacology of MD2 inhibitors. .Our group has carried out the medicinal chemical and pharmacological research on curcumin analogues as anti-inflammatory agents for a long term. In the previous study, two novel compounds A13 and C12 were found to be able to directly bind to MD2 protein, which contributes to their inhibition against MD2 activity and subsequent LPS-induced inflammation in vitro and in vivo. Thus, we hypothesize that, with the leading compounds A13/C12, we may demonstrate the MD2-inhibitor binding mechanism and sites, and may design and find a series of new candidates for the treatment of sepsis. Also, these studies will enhance our recognition towards MD2 as a new therapeutic target..In the present project, we plan to demonstrate the A13/C12-MD2 binding site and mechanism via a series of enzymological experiments, to design and synthesized about 60 A13/C12 analogues with the computer-assistant molecular docking and virtual screening, and to test the anti-inflammatory activity and pharmacological function of these synthetic analogues. The performation of this project will provide a new target and a kind of new leading structure for the anti-sepsis drug development.

脓毒血症是全身性急性炎症反应综合征，主要有感染和内毒素引起。Toll样受体4/髓样分化蛋白2（TLR4/MD2）是内毒素LPS的受体，介导LPS急性炎症反应。虽然MD2基因敲除可以降低LPS敏感性，但是作为TLR4的辅助蛋白，MD2研究一直处于次要地位，MD2抑制剂抗炎药物和药理研究鲜有报道。我们长期从事以姜黄素为先导的抗炎药物化学和药理研究，前期工作中发现类似物A13和C12可以直接结合MD2蛋白并显著抑制LPS诱导的体内外炎症反应。我们假设：通过以姜黄素、A13和C12为先导，了解其与MD2的结合模式和机制，有望设计和发现新的治疗脓毒血症的候选新药，增强对MD2靶点的认识。本项目拟通过系列酶学研究深入阐明A13/C12与MD2的结合机制；以A13/C12等类似物为先导基于蛋白结构分子对接设计合成一系列类似物并进行抗炎测试和体内外药理研究。本研究将为脓毒症药物设计提供新的靶点和结构先导。

脓毒血症是全身性急性炎症反应综合征，主要有感染和内毒素引起。Toll样受体4/髓样分化蛋白2（TLR4/MD2）是内毒素LPS的受体，介导LPS急性炎症反应。虽然MD2基因敲除可以降低LPS敏感性，但是作为TLR4的辅助蛋白，MD2研究一直处于次要地位，MD2抑制剂抗炎药物和药理研究鲜有报道。我们长期从事以姜黄素为先导的抗炎药物化学和药理研究，前期工作中发现姜黄素类似物A13和C12可以直接结合MD2蛋白并显著抑制LPS诱导的体内外炎症反应。在本课题中我们采用多种分子生物学手段首先阐明了姜黄素与MD2蛋白的结合机制和结合方式；基于姜黄素的化学结构设计合成了68个姜黄素类似物，并且通过波谱学手段表征了它们的化学结构，大部分合成的化合物在细胞层面显著抑制LPS诱导的炎症因子TNF-α和IL-6的释放，随后对活性化合物的抗炎机制和抗炎靶点进行了研究，确定活性化合物是通过靶向MD2蛋白抑制LPS/TLR4诱导的炎症反应缓解脓毒症和急性肺损伤；最后我们对前期发现的抗炎小分子L6H21是MD2蛋白的特异性小分子抑制剂，能够靶向MD2在体内外发挥抗炎活性。通过本课题的研究，我们发现了3个MD2特异性小分子抑制剂，确证了MD2蛋白介导LPS诱导的脓毒症和急性肺损伤，为后续MD2小分子抑制剂的研发提供了先导化合物。