Natural Product HCV Drugs from Rare Plant-Microbe Interactions

该项目的主要目标是探索美国稀有和濒危植物及其独特的宿主微生物群落。这些社区将研究其生物合成和生产的新类别的天然产物（NP）的HCV药物导致独特的MOA。该项目涉及与密西西比大学Mark Hamann教授的实验室以及Emory大学/Atlanta VA的RFS Pharma和Raymond Schinazi的Tony Whitaker建立的良好合作关系。 集体的专业知识提供了一个独特的机会，以探索新的来源，治疗丙型肝炎病毒是耐药的目前形式的治疗。 该方案的具体目标包括： 1.每年收集100株濒危植物伴生细菌和真菌菌株并进行系统发育分析。 系统发育多样性和独特的美国濒危植物物种（20）将被评估的细菌和真菌抑制丙型肝炎病毒。 这将在五年的项目期间提供500种独特的细菌和真菌。 2.创新的表观遗传学工具将用于刺激沉默基因产生次级代谢产物。 这将包括商业上可用的监管工具以及来自宿主濒危植物本身的监管工具。 每一个新的菌株将受到10个或更多的表观遗传工具。 将使用具有UV-TOF-ELS检测的高灵敏度HPLC和小体积NMR来评价在标准培养条件下沉默基因的NP生产。3.在本授权申请中，一种独特的HCV真实的实时PCR检测方法作为初步筛选的特征，以鉴定从目标2中获得的提取物，其具有HCV选择性，可用于目标4中的分级分离。4.将使用HPLC结合MS、UV和ELSD检测对目标3的活性先导化合物进行纯化。 将使用真实的时间PCR完成重复生物测定，直至生成HCV选择性代谢产物。 天然产物的结构将使用综合NMR、HPLC-MS、UV、IR和X射线晶体学分析进行归属。

The key objective of this project involves the exploration of US rare and endangered plants and their unique host-microbiome community. These communities will be studied for their biosynthesis and production of new classes of natural products (NPs) HCV drug leads with unique MOAs. The project involves a well established collaborative relationship with the laboratories of Prof Mark Hamann and at the University of Mississippi and Tony Whitaker with RFS Pharma and Raymond Schinazi at Emory University/Atlanta VA. The collective expertise provides a unique opportunity to explore new sources for treatments for HCV which are resistant to current forms of treatment. The specific aims of this program include: 1. A collection and phylogenetic analysis of 100 endangered plant associated bacteria and fungi strains each year. Phylogenetically diverse and unique US endangered plant species (20) will be evaluated for bacteria and fungi which inhibit HCV. This will provide 500 unique bacteria and fungi over the course of the five year project period. 2. Innovative epigenetic tools will be applied to stimulate production of secondary metabolites from silent genes. These will include regulatory tools available commercially as well as those derived from the host endangered plant itself. Each new strain will be subjected to 10 or more epigenetic tools. Highly sensitive HPLC with UV-TOF-ELS detection and small volume NMR will be utilized to evaluate NP production from genes silent under standard culture conditions. 3. A unique HCV real time PCR assay is featured in this grant application as a primary screen to identify extracts obtained from Aim 2 with HCV selectivity for fractionation in aim 4. 4. Active leads from Aim 3 will be purified using HPLC with MS, UV and ELSD detection. Repeated bioassays using real time PCR will be completed until HCV selective metabolites are generated. Natural product structures will be assigned using integrated NMR, HPLC-MS, UV, IR and X-ray crystallographic analysis.