A Nonisotopic HTS Assay to Elucidate Choline Transporter (CHT) Modulators

用于阐明胆碱转运蛋白 (CHT) 调节剂的非同位素 HTS 测定

• 批准号: 7761581
• 负责人: Alicia M Ruggiero
• 金额: $ 2.5万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7761581
• 关键词: Acetylcholine; Acetylcholinesterase; Ache; Adverse effects; Alternative Therapies; Alzheimer' s Disease; Arousal; Attention; Binding; Biological Assay; Brain; Carrier Proteins; Cell Line; Cell membrane; Cell surface; Central Nervous System Diseases; Chemicals; Choline; Cholinesterase Inhibitors; Cholinesterases; Cognitive; Cognitive deficits; Collaborations; Data; Databases; Dementia; Development; Disease; Dose-Limiting; Drug abuse; Enhancers; Equipment; Exhibits; Future; Hemicholinium 3; Human; Lead; Learning; Measurement; Mediating; Membrane Potentials; Memory; Methodology; Mining; Molecular Conformation; Motor; Mutation; Nature; Neurons; Neurotransmitters; Pharmaceutical Preparations; Play; Process; Proteins; Psychotic Disorders; Radiolabeled; Reagent; Recovery; Research; Role; Screening procedure; Signal Transduction; Surface; System; Work; aging population; base; chemical synthesis; choline transporter; cholinergic; depression; design; high throughput screening; improved; motor control; mutant; neurotransmission; novel; prevent; public health relevance; radiotracer; response; small molecule libraries; tool; trafficking; uptake; voltage

DESCRIPTION (provided by applicant): In the brain, the chemical acetylcholine (ACh) exerts powerful modulatory control over arousal, motor and cognitive circuits, and has been found to be deficient in Alzheimer's Disease (AD). The current drugs available to positively impact cognitive deficits in Alzheimer's Disease (AD) and other dementias are the cholinesterase inhibitors. These prevent the breakdown of the neurotransmitter acetylcholine (ACh), and thus augment Ach function. Due to the limited utility of the cholinesterase inhibitors, alternative therapies to augment ACh deficits are critical in our aging population. Another vital protein, the hemicholinium-3 sensitive choline transporter (CHT) is believed to be responsible for the efficient uptake of choline by neurons to allow for ACh synthesis. We have developed an assay system for high throughput screening to identify compounds with high selectivity for CHT. It is anticipated that these compounds may lead to future cholinergic therapies in AD, and multiple other CNS diseases regulated by cholinergic signaling. Our research aims are to implement a high throughput screen for discovery of novel enhancers of high-affinity choline transporter uptake as mediated by CHT using voltage-sensitive fluorescent measurements; to perform secondary assays aimed at identifying novel modulators that act selectively at the choline transporter (CHT); and to use database mining and chemical synthesis to optimize HTS hits for use in biological assays. Through the development of high-throughput screening (HTS) methodologies that allow for the screening of large chemical libraries, we seek to identify CHT activators and allosteric modulators. These compounds may be able to enhance choline uptake and increase the levels of ACh produced in the neuron. Such reagents would allow greater ACh release from viable neurons not lost in the disease process and could offer advantages over cholinesterase therapy in targeting precursors rather than ACh itself. PUBLIC HEALTH RELEVANCE: Cognitive deficits associated with Alzheimer's Disease (AD) and other dementias are primarily caused by loss of the neurotransmitter acetylcholine (ACh) in the brain and treatments are currently limited to cholinesterase inhibitors. Additional and improved therapies to augment ACh deficits are critical in our aging population and may be accessed by targeting another vital protein, the hemicholinium-3 sensitive choline transporter (CHT). CHT is responsible for the efficient uptake of choline by neurons and we have developed an assay for high throughput screening to identify compounds with high selectivity for CHT for future cholinergic therapies in AD, and other diseases regulated by cholinergic signaling.

描述（由申请人提供）：在大脑中，化学乙酰胆碱（ACH）对唤醒，运动和认知回路具有强大的调节性控制，并且已被发现在阿尔茨海默氏病（AD）中缺乏。目前可用于积极影响阿尔茨海默氏病（AD）和其他痴呆症的认知缺陷的药物是胆碱酯酶抑制剂。这些防止神经递质乙酰胆碱（ACH）的分解，从而增强ACH功能。由于胆碱酯酶抑制剂的效用有限，因此增加ACH缺陷的替代疗法对于我们的老龄化人群至关重要。另一种重要的蛋白质是Hemicholinium-3敏感胆碱转运蛋白（CHT），这是造成神经元有效胆碱的造成ACH合成的原因。我们已经开发了一个用于高吞吐量筛选的测定系统，以鉴定CHT选择性高的化合物。预计这些化合物可能会导致AD中未来的胆碱能疗法，以及通过胆碱能信号调节的多种其他中枢神经系统疾病。我们的研究目的是实施高吞吐量屏幕，以发现使用对电压敏感的荧光测量值介导的高亲和力胆碱转运蛋白摄取的新型增强子；执行旨在识别有选择性作用于胆碱转运蛋白（CHT）的新型调节剂的次要测定；并使用数据库挖掘和化学合成来优化HTS命中以用于生物测定。通过开发允许筛选大型化学文库的高通量筛选（HTS）方法，我们试图鉴定CHT激活剂和变构调节剂。这些化合物可能能够增强胆碱摄取并增加神经元中的ACH水平。这样的试剂将使在疾病过程中不会失去的可行神经元的ACH释放更大，并且可以在靶向前体而不是ACH时提供优于胆碱酯酶治疗的优势。 公共卫生相关性：与阿尔茨海默氏病（AD）和其他痴呆症相关的认知缺陷主要是由于大脑中神经递质乙酰胆碱（ACH）的丧失引起的，目前限于胆碱酯酶抑制剂。增加ACH缺陷的额外和改进的疗法在我们的老龄化人群中至关重要，可以通过靶向另一种重要的蛋白质，即Hemicholin-3敏感胆碱转运蛋白（CHT）来获得。 CHT负责神经元对胆碱的有效摄取，我们为高吞吐量筛选开发了一种测定法，以鉴定CHT高选择性的AD中CHT高选择性的化合物，以及其他受胆碱能信号调节的疾病。