以核受体FXR为靶标的配体药物筛选及其结构与功能机理研究

The study of structure-activity relationship (SAR), the relationship between the 3D structure of a molecule and its biological activity, is critical for the development of new drugs with improved potency and efficacy but reduced side effects. Our interests are focused on using X-ray crystal structural, molecular and biochemical multidisciplinary approaches and in vivo models to study the protein structure and function especially SAR analysis of nuclear receptors, with emphasis on their implications on drug discovery and therapeutics in the areas of human disease like diabetes and cancer. Nuclear receptors work as ligand inducible transcription factors and are among the most successful drug targets in history. However, clinical use of many ligands is limited by a number of side effects. It is thus critical to develop detailed structure-function relationships to discover potent nuclear receptor ligands that retain the beneficial activities without the undesired side effects..Along with the restriction of the TZD drug Avandia, the global research and development (R&D) of alternative diabetes drugs have reached a new climax. Nuclear receptors are important targets for drug discovery. FXR, a nuclear receptor, plays critical roles in glucose metabolism. FXR ligands, such as GW4064, showed some efficacy on diabetes, but their regulatary mechanism are still not clear. In addition, GW4064 revealed obvious side effects. As such, a new drug design strategy for FXR ligands is needed to yield more efficacious FXR-targeted drugs with less adverse effect. We have obtained a novel FXR ligand (NFL) through advanced high-throughput screening based on Alphascreen techniques. Prelimitary data showed that NFL treatment reduced blood insulin, glucose, triglyceride, free fatty acid, and cholesterol plasma levels. To gain the mechanistic understanding of this promising FXR ligand, we have obtained preliminary crystal structure of FXR in complex with NFL, and also studied the downstream genes related with FXR signaling in metabolism. Our proposed following studies will focus on optimization of the crystal structure of FXR with new ligands and also the anlysis of detailed structure-activity relatinship. Also the FXR dependent NFL function will be further investigated on FXR knockout mice. In summary, we will study the function of novel FXR ligand NFL in metabolism and its correlation with the nuclear receptor FXR signaling, as well as its structural and molecular mechanism in a comprehensive manner, from atomic structures, molecular mechanisms to animal model. This study will provide a potential new drug or a new drug design template for metabolic diseases targeting FXR, which will play great role in guiding drug R&D targeting nuclear receptors.

伴随着文迪雅等有效药物被限用、禁用的危机，全球研发治疗糖尿病药物进入了一个新高潮。核受体是药物研发的重要靶标，FXR是一种核受体转录因子，在代谢方面具有重要调控作用。已有报道FXR的配体GW4064对糖尿病有一定疗效，但其机理尚不是很清楚，且已显示出明显副作用。基于此，申请人已通过高通量Alphascreen筛选方法获得了一种新的FXR配体NFL，初步研究显示NFL对糖尿病小鼠有很好的疗效。申请人已获得FXR与NFL结合的初步晶体结构，对调节代谢的分子机理也进行了初步研究，后期将对其晶体结构进行进一步完善，并利用FXR基因敲除鼠验证NFL功能的受体依赖性，全面地从原子结构、分子机理、模型动物水平阐明NFL通过FXR调节代谢的功能及其结构和分子机理。该项目的研究，将对以FXR为靶标的代谢性疾病提供可能的新药或新药设计结构模板，对以核受体为靶标的药物研发提供极具指导作用的科学依据。

核受体是药物研发的重要靶标。法尼醇受体（FXR）作为核受体家族的一员，在机体内细胞分化、生长发育、新陈代谢等许多生理过程中发挥重要作用，更与系列重大疾病如糖尿病、肿瘤、炎症、心血管疾病等密切相关。但是由于已发现FXR配体的毒副作用，迄今为止，FDA批准的以FXR为靶标的药物屈指可数。鉴于FXR参与调节的重要功能，对以FXR为靶标的新药研究将具有重要的社会意义和重大经济价值。. 通过本项目的研究，我们筛选发现现有药物伊维菌素是FXR的配体。研究了伊维菌素与靶标FXR结合的晶体结构，并利用疾病模型小鼠、靶标FXR基因敲除小鼠研究了伊维菌素在体内通过靶标FXR调节的生理病理功能，发现伊维菌素能通过靶标FXR具有调节糖脂代谢的新功能。通过对伊维菌素系列衍生物进行分析，发现只有能结合FXR的伊维菌素衍生物才能调节糖脂代谢。研究了FXR选择性识别并结合伊维菌素的这些部分衍生物的结构特点，并对其功能进行了详尽的研究。本研究还发现天然产物萜类化合物tschimgine和feroline是FXR的新配体，并且首次发现这类配体与FXR结合的双口袋结合模式。天然产物三萜类化合物常春藤次酸是FXR的新配体，该配体与FXR结合的口袋与经典配体结合口袋明显不同。研究了这些新型萜类化合物与靶标FXR结合的晶体结构，并详细解析了其结构特征。还利用疾病模型小鼠、靶标FXR基因敲除小鼠研究了这些新配体化合物在体内通过靶标FXR调节的生理病理功能，发现它们能通过靶标FXR具有抑制炎症的功能。通过本项目的研究，我们在国际上有影响的刊物包括顶级刊物Nature子刊上发表了研究论文3篇，其中影响因子大于10的论文一篇，大于5的论文两篇；另有2篇论文正在投稿中。还申请了9项国内外专利，其中有两项获得了中国专利授权。本项目的研究成果，在现有药物再定位方面取得了突破性进展，为以FXR为模板的药物设计提供了多种结构模板，为以FXR为靶标的新药研究提供了非常丰富的科学数据和基础。这些研究，将会为我国获得自主知识产权的新药做出应有的贡献。