Discovery and Analysis of Synthetic TLR Agonists and Antagonists

描述（由申请人提供）：Toll样受体（TLR）很重要，因为它们介导感染的初始意识，还因为它们引起已知最强大的炎症反应。激动剂和拮抗剂均可用于临床目的。然而，人们对TLR如何被触发以产生定性上不同的信号，或者激动剂和拮抗剂结构的“规则”可能是什么，还知之甚少。我们建议合成化学家和TLR生物学家之间的合作，以确定和表征新的和固有的信息TLRs配体。Boger实验室已经使用液相合成来产生肽模拟分子的综合库，专门设计用于探测，模拟和破坏各种类型的蛋白质-蛋白质相互作用。Beutler实验室已经筛选了这个库（95，000种化合物）的TLR激动剂。在信息丰富和全面的α-螺旋模拟物子文库中，几个非常稳健和结构相关的命中揭示了一类新的TLR 4激动剂。生殖系突变小鼠的反应研究表明，与LPS（TLR 4的天然配体），这些分子的信号诱导NF-？B依赖性细胞因子TNF，而不是I型IFN。构效关系（SAR）分析已经开始揭示这些配体的激动剂活性的结构规则，这些配体在构象上类似于具有α-螺旋二级结构的七聚肽。将继续筛选正在进行的新库，以确定是否有半页、半页和半页。使肽模拟配体同时具有激动剂和拮抗剂性质。当它们被发现时，这些分子将进行SAR分析，优化活性，并将使用Beutler实验室中产生和维持的TLR信号突变体对其对TLR信号的影响进行详细研究。这些分子与TLR或相关蛋白质之间的相互作用位点将使用光亲和标记方法进行作图。在适当的情况下，配体将与其同源受体共结晶，用于高分辨率结构分析。还将在体内分析分子的性质（药代动力学、毒性、免疫佐剂作用和保护宿主免受模型感染的能力）。

DESCRIPTION (provided by applicant): The Toll-like receptors (TLRs) are important both because they mediate initial awareness of infection and because they elicit the most powerful inflammatory responses known. Both agonists and antagonists might be used for clinical purposes. Yet there is little understanding of how TLRs are triggered to yield qualitatively distinct signals, or what the "rules" for agonist and antagonist structure might be. We propose a collaboration between synthetic chemists and TLR biologists to identify and characterize new and inherently informative ligands for TLRs. The Boger lab has used solution-phase synthesis to produce a comprehensive library of peptide mimetic molecules, specifically designed to probe, mimic, and disrupt various types of protein-protein interactions. The Beutler lab has screened this library (95,000 compounds) for TLR agonists. Within an information-rich and comprehensive a-helix mimetic sub-library, several remarkably robust and structurally related hits have revealed a new class of TLR4 agonists. Studies of responses in germline mutant mice have shown that unlike LPS (the natural ligand for TLR4), these molecules signal to induce the NF-?B dependent cytokine TNF, but not type I IFNs. Structure-activity relationship (SAR) analyses have begun to reveal structural rules for agonist activity by these ligands, which conformationally resemble heptameric peptides with a-helical secondary structure. Continued screening of new libraries that are in progress will be performed to identify ¿-turn, ¿-sheet, and ?-turn peptidomimetic ligands with both agonist and antagonist properties. As they are discovered, these molecules will be subjected to SAR analysis, optimized for activity, and detailed studies of their effects on TLR signaling will be undertaken using TLR signaling mutants produced and maintained in the Beutler lab. The sites of interaction between these molecules and TLRs or associated proteins will be mapped using photoaffinity labeling methods. Where appropriate, the ligands will be co-crystallized with their cognate receptors for high resolution structural analysis. The properties of the molecules (pharmacokinetics, toxicity, immunoadjuvant effects, and ability to protect the host from model infections) will also be analyzed in vivo.