Fungal Acquisition, Isolation Chemistry, Structure Elucidation, and Bioassay

项目3包括三角研究所（RTI）和Mycosynthetix公司之间的合作努力。 [MSX)都位于北卡罗来纳州的三角研究园地区。后者提供了一个下- 探索源材料（丝状真菌;具体目标1），以补充收集 项目2中描述的蓝细菌的培养和培养。将从MSX收集中选择真菌 以确保最大的化学多样性的方式，这将涉及地理， 生态学和分类学信息，以及菌株产生次生代谢物能力的信息。 代谢物。真菌将使用先前显示的复杂培养基培养，以支持次级 新陈代谢. MSX将扩大有前途的潜在客户的培养规模，提供几乎无限的所需 化合物. RTI将开展天然产物分离和结构解析研究（具体目标2）， 这些将被监控，使用整个计划可用的生物资源的全部广度， 包括RTI的内部测定，用于恢复EZH 2对组蛋白H3的赖氨酸27的三甲基化活性 （H3-K27）和/或相对于非致瘤性乳腺腺癌细胞对致瘤性的选择性活性 （具体目标3）以及项目1、核心A和核心C中描述的那些生物靶标。 EZH 2表达最近以直接和因果的方式与肿瘤的转化和转移有关。 人前列腺癌、乳腺癌和肾细胞癌及其H3-K27三甲基化活性抑制 在体外和人肿瘤异种移植物中最终由细胞存活信号调节。浸提液检测 恢复细胞EZH 2 H3-K27三甲基化活性的化合物应该揭示基于机制的 抗肿瘤药物先导。 项目3将与项目1互动，以协助通过LC-MS进行样品的去复制， 药理学评价他们的植物为基础的铅。它将与项目2相互作用， 样品通过文献数据库（NAPRALERT），结构解析研究将通过 与微线圈核磁共振合作。此外，还将在可供使用的生物测定中对样本进行检测。 整个团队，如引言中的图3所示。这样做的最终目标是获得最 从化学和真菌学的努力，为整个计划的利益药理价值。在这 A1应用程序，我们还重新设计了我们的数据管理计划，整个计划将使用 关系型真实的时间数据库，用于协调、交换、存储数据并帮助确定数据的优先级（参见核心 D）。 总的来说，项目3带来了多样化的源材料、成熟的化学物质以及创新和互补性 生物目标，都是为了整个计划自最初的改革以来， 这些数据大多是对初步数据的补充（C节）。

Project 3 comprises a collaborative effort between Research Triangle Institute (RTI) and Mycosynthetix, Inc. [MSX), both located in the Research Triangle Park area of North Carolina. The latter provides an under- explored source material (filamentous fungi; specific aim 1) that serves to complement the collection of Dlants and culturing of cyanobacteria described in Project 2. Fungi will be selected from the MSX collection n a manner that insures maximum chemical diversity, and this will involve a combination of geographic, ecological, and taxonomic information, as well as information on a strains ability to produce secondary metabolites. Fungi will be cultured using complex media shown previously to support secondary metabolism. MSX will scale-up the cultures of promising leads, providing a nearly limitless supply of desired compounds. RTI will carry out natural product isolation and structure elucidation studies (specific aim 2), and these will be monitored using the full breadth of biological resources available to the entire program, including in-house assays at RTI for restoration of EZH2 trimethylation activity toward lysine 27 of histone H3 (H3-K27) and/or selective activity towards tumorigenic over non-tumorigenic breast adenocarcinoma cells (specific aim 3) as well as those biological targets described in Project 1, Core A, and Core C. EZH2 expression has been linked recently in a direct and causal fashion to transformation and metastasis of human prostate, breast, and renal cell carcinoma, with inhibition of its H3-K27 trimethylation activity regulated ultimately by cell survival signals in vitro and in human tumor xenografts. Testing of extracts for compounds that restore cellular EZH2 H3-K27 trimethylation activity should reveal mechanism-based antitumor drug leads. Project 3 will interact with Project 1 for assistance with dereplication of samples via LC-MS and for pharmacological evaluation of their plant-based leads. It will interact with Project 2 for prioritization of samples via literature databases (NAPRALERT), and structure elucidation studies will be facilitated via collaboration with the micro-coil NMR. Also, the samples will be tested in biological assays available to the entire team, as illustrated in Figure 3 of the Introduction. In doing so, the ultimate goal is to derive the most pharmacological value from the chemistry and mycology efforts for the benefit of the entire Program. In this A1 application, we have also re-designed our data management plans, and the entire Program will use a relational, real time database to coordinate, to exchange, to store, and to help prioritize the data (see Core D). In total, Project 3 brings diverse source materials, proven chemistry, and innovative and complementary biological targets, all for the use of the entire Program. Minor changes have been instituted since the initial application, and most of these constitute additions to the preliminary data (section C).