Small Molecule Regulators of Beta-Cell Differentiation

β 细胞分化的小分子调节剂

• 批准号: 7114248
• 负责人: MARK MERCOLA
• 金额: $ 13.44万
• 依托单位: SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-05 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7114248
• 关键词: biomedical automation; cell differentiation; cell line; chemical registry /resource; clinical research; combinatorial chemistry; drug discovery /isolation; embryonic stem cell; genetic transduction; green fluorescent proteins; high throughput technology; human tissue; immunocytochemistry; laboratory mouse; pancreatic islets; polymerase chain reaction; small molecule; technology /technique development; transcription factor

DESCRIPTION (provided by applicant): Preliminary data in this Collaborative R01 Project demonstrate the feasibility of screening libraries of small molecules to discover compounds that promote the differentiation of embryonic stem (ES) cells and human pancreatic duct cells. A substantial Institutional investment has been made in chemical libraries, robotics and automated microscopy to perform cell-based screens. The proposed research will build on these successes and this infrastructure by developing high throughput screening technology to discover molecules that promote beta-cell differentiation. A strong synergy is created by the complementary expertise of each laboratory. The Mercola laboratory has led development of cell-based screening efforts at the Institute and provides expertise in robotic screening, ES cell culture and HIV lentiviral technology while the Levine laboratory provides knowledge of diabetes, technology for culture and manipulation of primary human pancreatic duct tissue. Aims 1-3 will develop and perform a pilot screen for molecules that promote ES cells to differentiate into pancreatic endodermal precursors. To do this, we will produce a panel of ES cells with GFP under control of differentiation specific promoters to evaluate differentiation-promoting capability of 10,000 natural product and established drugs. This compound library was chosen for chemical diversity and prior success at yielding modulators of intracellular signaling pathways in several cell types. Aims 4-5 will repeat the screens with adult pancreatic duct cells and an immortalized cell line, both transduced with HIV-lentiviral vector carrying GFP under control of the insulin promoter. An unbiased, chemical approach to beta-cell differentiation is made possible by advances in high throughput technologies, availability of diverse chemical libraries and the ability to easily place reporter proteins such as GFP in relevant cells under the control of genes that characterize distinct stages in pancreatic differentiation. In addition to providing tools to regulate differentiation, knowledge of the "hits" might reveal insight into cellular targets that are involved in pancreatic development, or be used as affinity probes to identify such proteins, in a process that is becoming known as chemical genetics.

描述（由申请人提供）： 该协作R01项目中的初步数据证明了筛选小分子库以发现促进胚胎干细胞（ES）细胞和人类胰管细胞的分化的化合物的可行性。在化学库，机器人技术和自动显微镜上进行了大量的机构投资，以执行基于细胞的屏幕。拟议的研究将通过开发高吞吐量筛选技术来发现促进β细胞分化的分子来建立这些成功和基础设施。 每个实验室的补充专业知识创造了强大的协同作用。 Mercola实验室领导了该研究所的基于细胞的筛查工作的发展，并提供了机器人筛查，ES培养和HIV慢病毒技术方面的专业知识，而Levine Laboratory提供了糖尿病的知识，用于培养和操纵原发性人类胰管组织的技术。 AIMS 1-3将开发并执行一个促进ES细胞以区分胰腺内胚层前体的分子的试验屏幕。为此，我们将在控制特定启动子的控制下生产具有GFP的ES细胞，以评估10,000种天然产物和已建立药物的促进能力。选择该化合物文库以进行化学多样性和先前的成功，以产生几种细胞类型的细胞内信号通路的调节剂。 AIMS 4-5将用成年胰管细胞和永生的细胞系重复筛选，均用携带GFP的HIV静脉病毒载体转导的胰岛素启动子。 通过高通量技术的进步，各种化学文库的可用性以及在相关细胞中易于将报道蛋白（例如GFP）放置在相关细胞中，这些基因的控制表征了胰腺分化的不同阶段。除了提供调节差异化的工具外，对“命中”的知识还可能揭示对胰腺发育中涉及的细胞靶标的见解，或者在被称为化学遗传学的过程中用作识别此类蛋白质的亲和力探针。