A Genomic Approach to Discovering Novel Cathepsin Inhibitors from Cyanobacteria

从蓝藻中发现新型组织蛋白酶抑制剂的基因组方法

基本信息

批准号：
10531556
负责人：
Nicole Elizabeth Avalon
金额：
$ 7.16万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-12-01 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10531556
关键词：
Active Sites Anabaena Anabolism Aneurysmal Subarachnoid Hemorrhages Animal Model Antibiotic Resistance Architecture Atlases Behavioral Bioinformatics Biological Assay Biology Bypass Caspase Cathepsin L Cathepsins Cathepsins B Cells Cerebral Ischemia Cerebral hemisphere hemorrhage Cessation of life Chemicals Collection Complex Cyanobacterium Data Docking Drug Targeting Environmental Impact Enzymes FDA approved Fellowship Gene Cluster Gene Expression Genome Genomic Library Genomic approach Genomics Goals In Vitro Injury Institution Ion Channel Ischemia Knock-out Lead Libraries Literature Methods Mining Modeling Molecular Morbidity - disease rate Mus National Research Service Awards Natural Products Neurocognitive Neurocognitive Deficit Neurologic Neurological outcome Neuronal Injury Neurons Oceanography Organism Outcome Pathway interactions Pattern Peptide Hydrolases Peptides Permeability Pharmaceutical Preparations Pharmacologic Substance Pharmacology Phylogeny Plasmids Postdoctoral Fellow Protease Inhibitor Proteins Reporter Ribosomes Scheme Secondary to Source Specificity Structure Techniques Technology Testing Traumatic Brain Injury Tubulin United States United States National Institutes of Health analog behavioral outcome clinical development design drug candidate drug discovery experimental study improved in silico inhibitor innovation interdisciplinary approach interest markov model mortality nanomolar natural product inspired neuroinflammation neuropathology neuroprotection novel novel therapeutics overexpression peptide natural products peptide synthase pharmacophore polyketide synthase polyketides post-doctoral training pre-clinical prevent professor prognostic resistance gene small molecule synergism tandem mass spectrometry tool virtual yeast two hybrid system

项目摘要

Project Abstract Traumatic brain injury (TBI) is a leading cause of morbidity and mortality world-wide and in the United States;1,2 however, there are no FDA-approved medications to ameliorate the devastating consequences of secondary injury caused by complex neuropathological cascades following the initial neuronal insult. Cathepsin B and L represent promising drug targets, as knock-out and inhibition studies of these cysteine proteases in animal models reveal improvement in behavioral and neurocognitive sequelae.3–7 There is a clear need for new therapeutic options to treat secondary injury following TBI, and cathepsin B and L inhibitors have pharmaceutical promise. Over 60% of FDA-approved drugs are derived from or inspired by natural products (NPs).8 Cyanobacteria are known to produce NPs with a range of bioactivity, including activity as protease inhibitors.9,10 NPs are biosynthesized by megaenzymes that are encoded as discrete genomic packages called biosynthetic gene clusters (BGCs).11 We hypothesize that a genomic approach can be used to enhance drug discovery efforts from cyanobacteria for novel cathepsin B and L inhibitors through the three aims outlined below. First, an innovative pharmacophore-based genome mining pipeline will be developed (Aim 1). In this aim, pharmacophores deduced from virtual docking experiments and known cathepsin inhibitors will be used to predict enzymatic domains responsible for the creation of the desired pharmacophore. This retrobiosynthetic prediction will be used to make bioinformatic models to interrogate sequenced cyanobacterial genomes to find candidate BGCs. The BGCs of highest interest will be identified, delineated, and the compounds produced through either cultivation or heterologous expression (Aim 2). Using a full retrobiosynthetic prediction for gallinamide A, a compound that demonstrates nanomolar cathepsin L inhibition, we will search for the BGC within the genomic library at Scripps or in new cyanobacterial collections. Additional BGCs from Aim 1 will be developed in this aim as well. If the BGC is not constitutively expressed or if the BGC is not associated with a cultivatable organism, heterologous expression will be pursued (Aim 3). Compound isolation and molecular networking to analyze and annotate the chemical space of the natural products produced will follow. Compounds will be tested in bioassays to evaluate cathepsin B and L activity on purified enzymes as well as in neuronal and glial cellular studies. The gap between the identified pharmaceutical need and the discovery of novel bioactive molecules can be bridged by the innovative multidisciplinary approach presented in this application. The proposed project will be carried out as part of an NIH F32 NRSA Fellowship at Scripps Institution of Oceanography, UC San Diego, under the co-sponsorship of Professors William Gerwick and Vivian Hook and a team of collaborators that will train the postdoctoral fellow in virtual docking experiments, bioinformatics, heterologous gene expression, and cathepsin-related bioassays.

项目摘要创伤性脑损伤 (TBI) 是全世界和美国发病率和死亡率的主要原因；1,2 然而，目前还没有 FDA 批准的药物可以减轻继发性炎症的破坏性后果最初的神经元损伤后由复杂的神经病理级联反应引起的损伤。有前景的药物靶点，如动物体内这些半胱氨酸蛋白酶的敲除和抑制研究模型揭示了行为和神经认知后遗症。3-7 显然需要新的治疗 TBI 后继发性损伤的治疗选择，组织蛋白酶 B 和 L 抑制剂已超过 60% 的 FDA 批准药物源自或受天然产品启发。 (NP).8 已知蓝藻可产生具有一系列生物活性的 NP，包括蛋白酶活性抑制剂。9,10 NP 由大酶生物合成，这些大酶被编码为离散基因组包，称为生物合成基因簇 (BGC)。11 我们发现基因组方法可用于增强药物通过概述的三个目标，蓝藻对新型组织蛋白酶 B 和 L 抑制剂的发现工作首先，将开发一种基于药效基团的创新基因组挖掘管道（目标 1）。目的，从虚拟对接实验推导出来的药效团和已知的组织蛋白酶抑制剂将用于预测负责产生所需药效基团的酶结构域。预测将用于建立生物信息学模型来询问测序的蓝藻基因组以发现将识别、描述最感兴趣的候选 BGC，并生成化合物。通过培养或异源表达（目标 2）使用完整的逆转录生物合成预测。没食子酰胺 A 是一种具有纳摩尔组织蛋白酶 L 抑制作用的化合物，我们将寻找 BGC 斯克里普斯的基因组文库或来自 Aim 1 的新蓝藻保藏中的其他 BGC 将是。如果 BGC 没有被组成性地表达或者 BGC 不与某个目标相关联。可培养的生物体，将追求异源表达（目标 3）。随后将建立网络来分析和注释所生产的天然产物的化学空间。化合物将在生物测定中进行测试，以评估组织蛋白酶 B 和 L 对纯化酶的活性以及神经元和神经胶质细胞研究之间的差距已确定的药物需求和发现之间的差距。新颖的生物活性分子可以通过本文中提出的创新的多学科方法来桥接拟议的项目将作为斯克里普斯 NIH F32 NRSA 奖学金的一部分进行。加州大学圣地亚哥分校海洋学研究所，由 William Gerwick 教授和 Vivian Hook 和一个合作者团队将培训博士后研究员进行虚拟对接实验，生物信息学、异源基因表达和组织蛋白酶相关生物测定。