A toolbox of promiscuous immobilisation chemistries to identify the targets of natural compounds by chemoproteomics (PromisChemProt)

混杂固定化化学工具箱，用于通过化学蛋白质组学识别天然化合物的靶标 (PromisChemProt)

• 批准号: 452256511
• 负责人: Professor Dr. Bernhard Küster, since 10/2022
• 金额: --
• 依托单位: Lehrstuhl für Proteomik und Bioanalytik / Experimentelle Bioinformatik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/452256511?language=en
• 关键词: toolbox; promiscuous; immobilisation; chemistries; identify

Natural products have evolved to benefit their producing organisms: contrary to synthetic man-made compounds, they have been selected and optimized by evolution to interact with biological targets for example as attack or defense mechanisms. Natural products thus constitute a promising pool of innovative chemical biology tools and potential drugs that would benefit from systematic exploration. Target proteins can be involved in essential cellular pathways of other organisms, and are many times not yet validated drug targets for human medicine. Phenotypic screenings have uncovered many natural compounds with anti-tumor activity but it is often unclear which cellular targets are engaged and lead to the observed phenotypic outcome. Yet, acquiring this information, i.e. target deconvolution, is of utmost importance to convert observable phenotypes into actionable novel chemotypes and/or novel biological targets. Target deconvolution of those screening hits is often deterred by the lack of generic experimental approaches in the context of complex cellular systems. In particular, chemical synthesis of an ad-hoc linkable analogue is often used to interrogate the target space of a molecule. However natural products are often difficult to synthesize which complicates e.g. activity-based or affinity-based target deconvolution approaches. With this research proposal, we ambition to develop a robust chemoproteomics pipeline that allows to perform target deconvolution of any chosen unmodified natural product: More precisely, we will build a toolbox of chemistries able to immobilize small amounts of natural compounds on beads. We will investigate diazirine for carbene insertion, Mitsunobu reaction, gold catalyzed cycloaddition and alkoxylation as well as enyne metathesis as means to promiscuously react natural products with reagent-loaded beads. The obtained affinity matrices will enable the affinity purification of targets of natural compounds from cellular lysates and their characterization by quantitative mass spectrometry, building on our chemical proteomics expertise. Because of the wealth of alkenes and alcohol functionalities present in natural products, we anticipate that at least one of the utilized chemistries will produce a linkage compatible with the molecule:target binding event. Since such propitious linkage site cannot be predicted for unknown targets, our concept relies on the parallel agnostic use of a set of chemistries, which will be applied to forty natural compounds confirmed to act on cancer cell lines. Any confirmed causality link between the engagement of the newly identified target by the natural product and he phenotype will constitute an actionable starting point for pharmaceutical research, where the lack of validated targets is pressing for alternative drug discovery paradigms.

天然产物的进化使其生产生物体受益：与合成的人造化合物相反，它们通过进化被选择和优化，以与生物靶标相互作用，例如作为攻击或防御机制。因此，天然产品是一个有希望的创新化学生物学工具和潜在药物库，将受益于系统的探索。靶蛋白可以参与其他生物体的基本细胞途径，并且很多时候尚未被验证为人类医学的药物靶标。表型筛选已经发现了许多具有抗肿瘤活性的天然化合物，但通常不清楚哪些细胞靶点参与并导致观察到的表型结果。然而，获取该信息，即靶标去卷积，对于将可观察的表型转化为可操作的新型化学型和/或新型生物靶标是至关重要的。这些筛选命中的目标去卷积通常由于在复杂细胞系统的背景下缺乏通用实验方法而受到阻碍。特别是，化学合成的ad-hoc的类似物通常用于询问分子的目标空间。然而，天然产物通常难以合成，这使得例如基于活性或基于亲和力的目标去卷积方法复杂化。通过这项研究计划，我们的目标是开发一个强大的化学蛋白质组学管道，允许对任何选定的未修饰的天然产物进行目标解卷积：更确切地说，我们将建立一个能够在珠子上添加少量天然化合物的化学工具箱。我们将研究用于卡宾插入、Mitsunobu反应、金催化的环加成和烷氧基化以及烯炔复分解的二氮杂环丙烷，作为使天然产物与试剂负载珠混杂反应的手段。所获得的亲和基质将能够从细胞裂解物中亲和纯化天然化合物的靶标，并通过定量质谱法对其进行表征，建立在我们的化学蛋白质组学专业知识的基础上。由于天然产物中存在丰富的烯烃和醇官能团，我们预期至少一种所利用的化学物质将产生与分子相容的连接：靶结合事件。由于这种有利的连接位点无法预测未知的目标，我们的概念依赖于一组化学的平行不可知论的使用，这将被应用于40个天然化合物证实对癌细胞系的作用。任何确认的因果关系之间的联系，新确定的目标的天然产品和他的表型的参与将构成一个可行的起点，药物研究，缺乏验证的目标是紧迫的替代药物发现范式。