Chemical proteomic strategies for deciphering neocarzilin´s mode of action in cancer cells

破译新卡齐林在癌细胞中作用模式的化学蛋白质组学策略

• 批准号: 426512676
• 负责人: Dr. Sabine Schneider
• 金额: --
• 依托单位: Department Chemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2023-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/426512676?language=en
• 关键词: Chemical; proteomic; strategies; deciphering; neocarzilin

Natural products address a plethora of different proteins with essential functions. The deconvolution of their targets represents a unique opportunity for unravelling new anti-cancer strategies. We here apply chemical proteomics to identify the targets of the natural product neocarzilin within living cancer cells. Neocarzilin is a potent anti-cancer compound which was identified decades ago but still lacks functional characterization. In preliminary studies we synthetically equipped the natural product with an alkyne tag and treated proteomes to identify its cellular target(s). Interestingly, only one protein, the vesicle amine transport protein (VAT-1) with an essential role in cell migration, has been identified as prominent hit. As neocarzilin also exhibits strong anti-proliferative effects which are independent of VAT-1, other yet unidentified targets must exist. To decipher these targets we aim to improve our initial probe by incorporation of a signature stereocenter present in the natural product but absent in the first generation probe design. Target identification will follow our established chemical proteomic procedures and selected protein hits will be validated for compound binding by overexpression and crystallography. Another strong focus of this work is the already identified hit VAT-1. Our initial validation revealed a putative role in cancer cell migration, an important feature of malignant tumors. Further chemical proteomic studies are required to understand the cellular function of this poorly characterized protein and to exploit its potential for future cancer treatment. The studies proposed here will include co-immunoprecipitation in presence and absence of the natural product to decipher interaction partners, co-crystallization and structure elucidation with the compound to unravel the binding mode and rationalize improved binders. The overall aims of this proposal are to decipher the full spectrum of neocarzilin targets, understand its mode of action, investigate the role of VAT-1 and find improved derivatives with therapeutic potential.

天然产品涉及大量具有基本功能的不同蛋白质。他们的目标的反卷积为揭示新的抗癌策略提供了独特的机会。我们在这里应用化学蛋白质组学来识别活癌细胞内天然产物新卡齐林的靶标。新卡齐林是一种有效的抗癌化合物，几十年前就被发现，但仍缺乏功能表征。在初步研究中，我们合成地为天然产物配备了炔烃标签并处理蛋白质组以鉴定其细胞靶点。有趣的是，只有一种蛋白质，即在细胞迁移中发挥重要作用的囊泡胺转运蛋白（VAT-1），被认为是重要的目标。由于新卡齐林还表现出独立于 VAT-1 的强大抗增殖作用，因此必定存在其他尚未确定的靶点。为了破译这些目标，我们的目标是通过纳入天然产物中存在但第一代探针设计中不存在的特征立体中心来改进我们的初始探针。目标鉴定将遵循我们既定的化学蛋白质组学程序，并且将通过过表达和晶体学来验证选定的蛋白质命中的化合物结合。这项工作的另一个重点是已经确定的热门 VAT-1。我们的初步验证揭示了其在癌细胞迁移中的假定作用，这是恶性肿瘤的一个重要特征。需要进一步的化学蛋白质组学研究来了解这种特征不明的蛋白质的细胞功能并开发其用于未来癌症治疗的潜力。这里提出的研究将包括在存在和不存在天然产物的情况下进行免疫共沉淀，以破译相互作用伙伴、与化合物的共结晶和结构阐明，以揭示结合模式并合理化改进的结合剂。该提案的总体目标是破译新卡齐林靶点的全谱，了解其作用模式，研究 VAT-1 的作用并找到具有治疗潜力的改进衍生物。