Fragment-Based Ligand Discovery in Proteomes

描述（由申请人提供）：人类蛋白质组的很大一部分（超过50%）仍未表征，这些蛋白质中甚至更大一部分缺乏用于功能分析的共价选择性探针。高通量筛选（HTS）和基于片段的配体发现（FBLD）在开发新的探针和抑制剂方面都取得了成功，但受到有关靶标的生物学洞察和结构信息的先决条件的挑战。为了解决全球识别蛋白质调节新位点的需求，我们已经开始开发基于片段的蛋白质组配体发现（FBLDiP）的化学蛋白质组学平台。我们建议使用这个平台来发现新的配体结合位点和片段，这些位点以大规模并行的方式调节蛋白质活性，从而获得传统上“不可药用”的靶标。FBLDiP同时报告了蛋白质组中数千个半胱氨酸的标记位点和片段结合的构效关系（SAR）。我们已经开始实施FBLDiP平台，并在许多癌症相关蛋白中发现了新的配体结合位点，包括表观遗传酶PRMT 1，转录因子NF-？B、致癌衔接蛋白CRKL、癌症代谢酶IDH 1和表征不佳的激酶ZAK。半胱氨酸反应性片段文库的合成正在进行中，我们预计将产生100多个化合物文库。生物学评价的配体结合对选定的确定的蛋白质的影响已经开始和初步的数据支持，FBLDiP方法是有效的发现新的配体结合位点和探针，这些网站。在对接和结构生物学的指导下，我们将在癌症相关靶点的新配体结合位点产生有效的选择性抑制剂。抑制剂合成将由来自FBLDiP筛选和二级基于凝胶的蛋白质谱分析的目标靶标的SAR谱引导。我们预计，FBLDiP方法将发现广泛的蛋白质调节和铅抑制剂靶向这些网站的新网站。总的来说，这项研究提供了一种令人兴奋的新方法来识别配体结合位点，并指导重要和非靶向的生物相关蛋白质的抑制剂的合成。

DESCRIPTION (provided by applicant): A significant portion of the human proteome (greater than 50%) remains uncharacterized and an even greater fraction of these proteins lack covalent, selective probes for functional analysis. High-Throughput Screening (HTS) and Fragment-Based Ligand Discovery (FBLD) have both had success in the development of new probes and inhibitors but are challenged by prerequisites for biological insight and structural information about a target. To address the need for global identification of new sites for protein modulation, we have begun to develop a chemoproteomic platform for Fragment-Based Ligand Discovery in Proteomes (FBLDiP). We propose to use this platform to discover novel ligand-binding sites and fragments that modulate protein activity at these sites in a massively parallel fashion that will access traditionally 'undruggable' targets. FBLDiP simultaneously reports the site of labeling and the structure activity relationship (SAR) of fragment binding at thousands of cysteines within the proteome. We have already begun to implement the FBLDiP platform and have uncovered novel ligand-binding sites in many cancer relevant proteins, including the epigenetic enzyme PRMT1, the transcription factor NF-?B, the oncogenic adaptor protein CRKL, cancer metabolism enzyme IDH1 and the poorly characterized kinase ZAK. The synthesis of a cysteine-reactive fragment library is ongoing and we anticipate the production of a 100+ compound library. Biological evaluation of the effects of ligand-binding on selected identified proteins has begun and preliminary data supports that the FBLDiP methodology is effective for discovery of novel ligand-binding sites and probes for these sites. Guided by docking and structural biology, we will generate potent, selective inhibitors at novel ligand-binding sites in cancer relevant targets. Inhibitor synthesis will be guided by SAR profiles for targets of interest from FBLDiP screening and secondary gel-based protein profiling. We anticipate that the FBLDiP methodology will uncover a wide range of new sites for protein modulation and lead inhibitors to target these sites. As a whole, this research offers an exciting new means to identify ligand-binding sites and to guide the synthesis of inhibitors for important and untargeted, biologically-relevant proteins.