High-Content Screening for Peroxisome Biogenesis for Type-II Diabetes.

II 型糖尿病过氧化物酶体生物发生的高内涵筛选。

• 批准号: 8210816
• 负责人: JAY BRENMAN
• 金额: $ 31.15万
• 依托单位: NORTH CAROLINA CENTRAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-01-01 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8210816
• 关键词: Adipose tissue; Adverse effects; Agonist; Animal Model; Animals; Biogenesis; Biological Assay; Biological Factors; Boxing; Cardiovascular system; Cell Culture Techniques; Cells; Chemicals; Complex; Development; Diet; Disease; Diversity Library; Dose; Drug Industry; Fatty Acids; Fatty Liver; Fatty acid glycerol esters; Fibrates; Future; Gene Expression; Gene Expression Profiling; Genes; Gluconeogenesis; Hepatocyte; Human; Hyperglycemia; Image; Individual; Informatics; Institutes; Insulin Resistance; Label; Lead; Libraries; Link; Lipids; Liver; Measurement; Messenger RNA; Metabolic Diseases; Metabolic syndrome; Metabolism; Metformin; Modeling; Molecular; Molecular Bank; Molecular Chaperones; Molecular Profiling; Morphology; Mus; Names; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Pathway interactions; Peroxisome Proliferator-Activated Receptors; Pharmaceutical Preparations; Phenotype; Phenylbutyrates; Physiology; Plasma; Production; Proteins; Reporter; Risk Factors; Rodent; Rodent Model; Role; Screening procedure; Site; Symptoms; Therapeutic; Thiazolidinediones; Tissues; Translating; Very Long Chain Fatty Acid; Western Blotting; assay development; base; bioimaging; catalase; cell type; diabetic; fatty acid oxidation; feeding; high throughput screening; improved; in vivo; interest; mRNA Expression; novel; novel therapeutics; oxidation; peroxisome; prevent; public health relevance; response; rosiglitazone; stable cell line; therapeutic evaluation; therapeutic target; uptake

DESCRIPTION (provided by applicant): We will optimize a high-content assay to identify compounds that promote peroxisome biogenesis in human cells. Compounds we identify can be valuable chemical probes for modeling peroxisome biogenesis disorders and evaluating the contribution of peroxisomes to normal metabolism and physiology in animal models. In addition, compounds may have therapeutic potential for the treatment of metabolic syndrome (MetS) and type 2 diabetes (T2D). The initial assay development will be performed in a high-content/image based screening format, with the intent of translating the assay to the Molecular Libraries Production Centers Network (MLPCN). Peroxisomes represent a major site of fatty acid -oxidation and the only site of very long-chain fatty acid (VLCFA) -oxidation in the cell. Compounds that increase peroxisome biogenesis in human cells might provide a novel therapeutic avenue for type-II diabetes and metabolic syndrome. Increased plasma fatty acids contribute to insulin resistance and hyperglycemia, which are risk factors for both metabolic syndrome and T2D. The lipotoxicity hypothesis suggests that elevated plasma fatty acids may lead to ectopic lipid accumulation in non- adipose tissue, particularly in the liver ("fatty liver"), which would impair its function. Therefore, preventing lipid accumulation in non-adipose tissue and decreasing fatty acids in plasma by increasing fatty acid oxidation could provide treatments for metabolic disorders. Structurally unrelated compounds that increase peroxisome biogenesis do improve metabolic syndrome and diabetic symptoms in rodents, validating this phenotypic target. We will use a high- content screening assay to identify compounds that promote peroxisome biogenesis in human cells using mechanisms independent of direct PPAR activation (non-classical peroxisomal biogenesis). Compounds we identify will be counter-screened with multiple orthogonal secondary assays to confirm increased peroxisomal functionality in cell culture. Gene expression profiling will be performed with compounds stimulating peroxisome biogenesis to identify distinct mechanisms. Chemical probes identified in this study can be used for modeling peroxisome biogenesis disorders and evaluating the contribution of peroxisomes to normal metabolism and physiology in animal models. The assay development will be performed in a high-content/image based screening format, with the intent of transferring the assay to the Molecular Libraries Production Centers Network (MLPCN) high-content specialized screening center (Burnham Institute). PUBLIC HEALTH RELEVANCE: This proposal utilizes a novel high throughput screening (HTS) assay to identify novel regulators of peroxisome biogenesis and function. Due to peroxisomes function in fatty acid oxidation and their known roles in alleviating symptoms of metabolic syndrome/Type 2 diabetes in mice, compounds that could increase peroxisome biogenesis/functionality in human cells could provide a first step towards developing novel therapeutics for metabolic syndrome and Type 2 diabetes. We have validated a known therapeutic compound in mice as a potent activator in our assay to use for compound screening and secondary/orthogonal assay development.

描述（由申请人提供）：我们将优化高内涵测定，以鉴定促进人类细胞中过氧化物酶体生物发生的化合物。我们鉴定的化合物可以作为有价值的化学探针，用于模拟过氧化物酶体生物发生障碍并评估过氧化物酶体对动物模型中正常代谢和生理学的贡献。此外，化合物可能具有治疗代谢综合征(MetS)和2型糖尿病(T2D)的潜力。最初的检测开发将以基于高内容/图像的筛选格式进行，目的是将检测转化为分子图书馆生产中心网络（MLPCN）。过氧化物酶体代表了细胞中脂肪酸氧化的主要位点和极长链脂肪酸（VLCFA）氧化的唯一位点。增加人类细胞中过氧化物酶体生物发生的化合物可能为 II 型糖尿病和代谢综合征提供新的治疗途径。血浆脂肪酸增加会导致胰岛素抵抗和高血糖，这是代谢综合征和 T2D 的危险因素。脂毒性假说表明血浆脂肪酸升高可能导致非脂肪组织中异位脂质积累，特别是肝脏（“脂肪肝”），这会损害其功能。因此，通过增加脂肪酸氧化来防止非脂肪组织中的脂质积累并减少血浆中的脂肪酸可以为代谢紊乱提供治疗。增加过氧化物酶体生物合成的结构不相关的化合物确实可以改善啮齿动物的代谢综合征和糖尿病症状，从而验证了这一表型目标。我们将使用高内涵筛选测定法，利用独立于直接 PPAR 激活的机制（非经典过氧化物酶体生物发生）来鉴定促进人类细胞中过氧化物酶体生物发生的化合物。我们鉴定的化合物将通过多个正交二次测定进行反筛选，以确认细胞培养物中过氧化物酶体功能的增加。将使用刺激过氧化物酶体生物发生的化合物进行基因表达谱分析，以确定不同的机制。本研究中确定的化学探针可用于模拟过氧化物酶体生物发生障碍并评估过氧化物酶体对动物模型中正常代谢和生理学的贡献。检测开发将以基于高内容/图像的筛选格式进行，目的是将检测转移到分子图书馆生产中心网络 (MLPCN) 高内容专业筛选中心（伯纳姆研究所）。 公共健康相关性：该提案利用新型高通量筛选（HTS）测定法来鉴定过氧化物酶体生物合成和功能的新型调节剂。由于过氧化物酶体在脂肪酸氧化中的功能及其在减轻小鼠代谢综合征/2型糖尿病症状中的已知作用，能够增加人类细胞中过氧化物酶体生物发生/功能的化合物可以为开发代谢综合征和2型糖尿病的新疗法迈出第一步。我们已经在小鼠中验证了一种已知的治疗化合物作为我们的测定中的有效激活剂，用于化合物筛选和二次/正交测定开发。