A whole-animal small molecule screen to identify and characterize modifiers of Apolipoprotein B

用于识别和表征载脂蛋白 B 修饰物的全动物小分子筛选

• 批准号: 10460567
• 负责人: Daniel Kelpsch
• 金额: $ 6.98万
• 依托单位: CARNEGIE INSTITUTION OF WASHINGTON, D.C.
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-03 至 2023-09-02
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10460567
• 关键词: Affect; Affinity; Animal Model; Animals; Apolipoproteins B; Binding; Bioinformatics; Biological; Biological Assay; Biological Markers; Biology; Blood Circulation; Cardiovascular Diseases; Cells; Cholesterol; Chronic; Collaborations; Complex; Data; Dietary Fats; Disease Progression; Dose; Drug Screening; Environment; Etiology; FRAP1 gene; Female; Food; Future; Gene Expression; Homeostasis; Human; Imaging Techniques; Inflammation; Innovative Therapy; Institutes; Insulin Resistance; Intention; Intestines; Larva; Libraries; Lipids; Lipoproteins; Liver; Maleates; Measures; Mediating; Metabolic Diseases; Metabolic dysfunction; Metabolic syndrome; Metabolism; Modeling; Non-Insulin-Dependent Diabetes Mellitus; Optics; Other Genetics; Pathway interactions; Peripheral; Persons; Pharmacologic Substance; Phenotype; Physiology; Plasma; Production; Regulation; Reporter; Research; Risk Factors; Series; Serotonin; Serotonin Antagonists; Source; Syndrome; System; Testing; Therapeutic; Tissues; Transgenic Organisms; Triglycerides; Zebrafish; antagonist; base; career; design; drug repurposing; endoplasmic reticulum stress; experimental study; high throughput screening; high-throughput drug screening; hormonal signals; improved; improved outcome; in vivo; inhibitor; insight; interest; lipid metabolism; lipid transport; microsomal triglyceride transfer protein; nanoluciferase; non-alcoholic fatty liver disease; novel; novel therapeutics; receptor; screening; serotonin receptor; side effect; small molecule; success; transcriptome sequencing

PROJECT SUMMARY Metabolic syndrome, encompassing type 2 diabetes, non-alcoholic fatty-liver disease, and cardiovascular disease, affects more than one billion people worldwide. While its etiology is complex, the best biological marker of metabolic syndrome is increased levels of apolipoprotein B (ApoB)-containing lipoproteins (B-lps). B-lps transport triglycerides and cholesterol through the plasma to peripheral tissues, and excess plasma B-lps are causative to metabolic syndrome. A single ApoB molecule decorates each B-lp and is essential for its function. However, the cellular mechanisms that ultimately regulate ApoB and B-lp production, secretion, transport, and degradation remains to be fully defined. The proposed studies aim to identify new molecules that alter B-lp physiology with the hope of not only generating new therapeutics but to elucidate new cell biological mechanisms of ApoB regulation. Human B-lp biology is remarkably conserved in the zebrafish. Further, zebrafish produce large numbers of progeny, larvae are optically transparent, and larvae do not require an exogenous food source. Thus, the zebrafish is the ideal model to identify novel mechanisms of ApoB modulation. Therefore, the Farber lab generated an in vivo chemiluminescent reporter of ApoB that does not disrupt ApoB function. Thus, I hypothesize that I can identify novel drugs that modulate ApoB regulation, turnover, and function to rectify metabolic dysfunction using this whole-animal reporter of ApoB. I have developed a high-throughput assay to screen chemiluminescence from whole zebrafish. Each compound that reduces ApoB from a drug repurposing library will be further validated by several assays measuring ApoB and B-lps production, size, and turnover. I will also evaluate the effects of each compound on whole-animal physiology. My screening efforts have identified 25 ApoB-lowering compounds. One compound, pimethixene maleate, specifically reduces ApoB levels in a dose- dependent manner. Prior research suggests pimethixene is a serotonin receptor antagonist. Studies suggest that serotonin influence B-lps levels through regulation of the mammalian Target of Rapamycin (mTOR). Thus, I hypothesize that pimethixene-dependent ApoB reduction is mediated by 5-HT2 receptor antagonism. I will determine whether pimethixene directly alters this pathway. Further, I will examine whether this compound improves several risk factors associated with metabolic disease using a series of established transgenic reporter lines and bioinformatic approaches. Ultimately, this research aims not only to identify novel ApoB-modulating therapeutics that would improve outcomes of metabolic disease but would also provide fundamental insights into the regulation and function of ApoB. Together, the research environment of the Farber lab and the Carnegie Institute are ideal for the success of this project and my success in the future as I grow towards an independent career in metabolism research.

项目总结