In vivo HTS assay for novel modulators of Apolipoprotein B

载脂蛋白 B 新型调节剂的体内 HTS 测定

• 批准号: 10502731
• 负责人: STEVEN A FARBER
• 金额: --
• 依托单位: CARNEGIE INSTITUTION OF WASHINGTON, D.C.
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10502731
• 关键词: Address; Animals; Apolipoproteins B; Area; Biological Assay; Biological Markers; Biological Models; Biology; Cardiovascular Diseases; Cells; Chemicals; Chronic; Clinical; Collaborations; Collection; Communities; Complement; Coupled; Cultured Cells; Development; Diabetes Mellitus; Disease; Disease Progression; Disease model; Drug Modelings; Eligibility Determination; Energy Metabolism; FDA approved; Fatty acid glycerol esters; Fishes; Funding; Genome engineering; Health; Hepatic; Human; Image; Individual; Inflammation; Insulin Resistance; Investigation; Laboratories; Larva; Lead; Libraries; Link; Luciferases; Mediator of activation protein; Metabolic; Metabolic Diseases; Metabolic syndrome; Metabolism; Modeling; Monitor; Mus; Organism; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Phenotype; Physiology; Probability; Process; Production; Reporter; Research; Risk Factors; Robotics; Scientist; Series; Structure; Study models; System; Techniques; Testing; Therapeutic; Toxic effect; Transgenic Organisms; Translations; Triglycerides; United States National Institutes of Health; Universities; Validation; Whole Organism; Zebrafish; assay development; atherogenesis; base; combat; dietary; disease phenotype; drug discovery; efficacy evaluation; endoplasmic reticulum stress; experimental study; fatty liver disease; feeding; high throughput screening; imaging platform; in vivo; internal control; lead candidate; lipid metabolism; metabolic phenotype; novel; novel therapeutics; screening; statistics; success; therapeutic development; therapeutic evaluation; tool

Apolipoprotein-B (ApoB) is both a biomarker and a causal mediator of many central hallmarks of metabolic disease, including insulin resistance, fatty liver disease, atherogenesis, endoplasmic reticulum stress, and chronic inflammation. ApoB therefore serves as a useful phenotypic readout for the identification of compounds that engender diverse metabolic benefits. The present proposal will perform a high-throughput screen (HTS) to identify novel ApoB-lowering compounds using an automated robotics platform that enables screening to take place in live larval zebrafish using a genetically encoded chemiluminescent reporter to sensitively detect ApoB levels in individual fish. To accomplish this effort, we have brought together a team of scientists all located at Johns Hopkins University and leaders in their respective fields. Farber has established the zebrafish as a model for studies of vertebrate lipid metabolism, Mumm has created a powerful HTS zebrafish screening platform, Ahima is a world leader in mammalian energy metabolism and Lectka is an established chemist bringing significant expertise in screen hit prioritization to the effort. The HTS will take place in two iterations, with the first iteration screening a ~3,000 compound library of clinically approved compounds so that hits can be rapidly repurposed to treat a host of disease associated with Apob perturbations. The second iteration screening is much larger effort to maximize compound diversity (30,000 compounds) and discover potentially entirely new avenues for treatment. Hits from the both screens will be subjected to a high-content secondary screen that uses an automated imaging platform to monitor effects on disease progression using a panel of transgenic zebrafish carrying fluorescent reporters of several important metabolic disease risk factors. This secondary screen will efficiently classify and prioritize hits from the primary screen and identify the subset of compounds with validated metabolic benefits in live vertebrate organisms that justify further investigation and therapeutic development. Promising compounds from primary and secondary screening will also be validated for activity in mammalian models, including mouse and human cultured cells. The results of these efforts will be the first ever whole animal HTS for ApoB modifiers coupled with a high-content secondary screen that together will enable the rapid identification of compounds to ameliorate many metabolic disease phenotypes, as well as a collection of hits for the development of novel therapies to combat the growing global burden of metabolic disease.

载脂蛋白-B(ApoB)既是一种生物标志物，也是许多代谢中心特征的因果介质 疾病，包括胰岛素抵抗、脂肪肝、动脉粥样硬化、内质网应激，以及 慢性炎症。因此，载脂蛋白B可作为一种有用的表型读数来鉴定 产生多种新陈代谢益处的化合物。本方案将执行高吞吐量 使用自动化机器人平台筛选(HTS)以识别新的降低载脂蛋白B的化合物 使用基因编码的化学发光报告对活体斑马鱼幼体进行筛选 灵敏地检测单条鱼中的载脂蛋白B水平。为了完成这项工作，我们组建了一个团队 都是约翰·霍普金斯大学的科学家和各自领域的领军人物。法伯有 建立了斑马鱼作为研究脊椎动物脂肪代谢的模型，妈妈创造了一个强大的 HTS斑马鱼筛选平台，Ahima是哺乳动物能量代谢的世界领先者，Lectka是 一位老牌化学家为这项工作带来了银幕热门电影优先排序方面的重要专业知识。HTS将采取 分两次迭代，第一次迭代筛选临床批准的~3,000个化合物文库 化合物，从而可以迅速改变HITS的用途，以治疗与载脂蛋白B相关的一系列疾病 微扰。第二次迭代筛选是更大的努力，以最大化复合多样性(30,000 化合物)，并发现潜在的全新治疗途径。来自两个屏幕的点击量将是 采用高内容二次屏幕，使用自动成像平台监控效果 利用携带几个重要荧光报告基因的转基因斑马鱼的疾病进展 代谢性疾病危险因素。此辅助屏幕将有效地对来自 初步筛选和鉴定在活体脊椎动物中具有有效代谢益处的化合物子集 证明进一步的研究和治疗开发是合理的。来自伯父的有希望的化合物 二次筛查也将在哺乳动物模型中验证活性，包括小鼠和人类 培养的细胞。这些努力的结果将是有史以来第一个用于偶联ApoB修饰剂的完整动物HTS 具有高含量的二次筛选，这将使化合物能够快速识别以 改善了许多代谢性疾病的表型，以及为小说的发展收集了热门作品 应对日益加重的全球代谢性疾病负担的治疗方法。