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。将研究这些生物群落的生物合成和生产具有独特moa的新型天然产物（NPs） HCV药物先导物。该项目涉及与密西西比大学Mark Hamann教授的实验室、RFS Pharma的Tony Whitaker实验室和埃默里大学/亚特兰大弗吉尼亚州的Raymond Schinazi实验室建立良好的合作关系。集体的专业知识为探索对当前治疗形式有耐药性的丙型肝炎病毒治疗的新来源提供了独特的机会。该计划的具体目标包括：1.；每年收集100种濒危植物相关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）将对抑制HCV的细菌和真菌进行评估。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被应用于刺激沉默基因产生次生代谢物。这些工具将包括商业上可用的管制工具以及来自宿主濒危植物本身的管制工具。每一个新菌株都将经受10个或更多的表观遗传工具。高灵敏度HPLC、UV-TOF-ELS检测和小体积NMR将被用于评估标准培养条件下沉默基因的NP生产。3. 一种独特的HCV实时PCR检测在本资助申请中被用作主要筛选，以鉴定从Aim 2中获得的提取物在Aim 4中具有HCV选择性。4. Aim 3的活性导联将采用HPLC、MS、UV和ELSD检测进行纯化。使用实时PCR重复生物测定将完成，直到产生HCV选择性代谢物。天然产物结构将通过核磁共振、高效液相色谱-质谱、紫外、红外和x射线晶体分析来确定。

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.