CADDMR: a Cell-based Assay for Drug Discovery for Mental Retardation Disorders

CADDMR：一种基于细胞的精神发育迟滞药物发现检测方法

• 批准号: 7130980
• 负责人: LINDA L RESTIFO
• 金额: $ 20.39万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7130980
• 关键词: Drosophilidae; brain; brain disorders; culture; drug discovery /isolation; drug screening /evaluation; gene mutation; genes; genetics; mental retardation; morphology; mutant; neurons; phenotype; tissue /cell culture

DESCRIPTION (provided by applicant): Mental retardation (MR), for which few effective treatments are available, is a common manifestation of developmental brain disorders. This proposal is based on the hypothesis that pharmacological correction of neuronal defects of morphology or plasticity will enhance the acquisition of cognitive skills in MR patients. Mutations of many different genes can cause MR, and many MR genes are members of interconnected genetic pathways that are very highly conserved among animal species. Such conservation provides an opportunity to use genetic model systems to explore novel therapeutic strategies prior to clinical testing. We have developed a primary cell culture assay for identification of abnormal morphology or plasticity of brain neurons with MR-gene mutations, as well as for identification of compounds that normalize the defects. The neurons are from fruit fly (Drosophila melanogaster) mushroom bodies, the best-characterized insect brain region involved in learning and memory, harvested during a stage of peak steroid-hormone-dependent developmental plasticity. When wild-type neurons are cultured in vitro they elaborate an arbor with a highly stereotyped morphology. The assay has revealed normal sexual dimorphism, as well as phenotypes due to single-gene mutations. In some cases, the in vitro culture system is highly sensitive, unmasking defects whose in vivo manifestations are too subtle to be detected by conventional microscopy. To enhance the speed of screening, we are developing NeuronMorphometrics, a set of Java plug-ins for ImageJ that perform semi-automated data analysis. About 75% of human MR genes have a counterpart in Drosophila that is similar enough by sequence to plausibly perform similar functions in the brain. We will screen a subset of Drosophila MR genes which we believe are prime candidates for revealing neuronal morphology and plasticity defects in vitro. In parallel studies, we will focus on the "filagree" phenotype, an abnormal mode of neurite outgrowth in vitro due to genetic loss of a cytoskeletal protein, and screen for compounds that normalize this phenotype. Together, the screens for phenotypes and for corrective chemical agents should set the stage for use of this assay as a stepping stone for drug discovery and development for MR disorders. Public health relevance: Our long-term goal is to treat mental retardation disorders with drugs that fix the underlying abnormalities of brain development. The proposed experiments are an important step toward that goal.

描述（由申请人提供）： 精神发育迟滞（MR）是大脑发育障碍的常见表现，目前几乎没有有效的治疗方法。该提议基于这样的假设：通过药物纠正神经元形态或可塑性缺陷将增强 MR 患者认知技能的获得。许多不同基因的突变都可能导致 MR，并且许多 MR 基因是相互关联的遗传途径的成员，这些遗传途径在动物物种中高度保守。这种保护提供了在临床测试之前使用遗传模型系统探索新的治疗策略的机会。我们开发了一种原代细胞培养测定法，用于鉴定具有 MR 基因突变的脑神经元的异常形态或可塑性，以及鉴定使缺陷正常化的化合物。这些神经元来自果蝇（果蝇）蘑菇体，这是昆虫大脑中参与学习和记忆的最典型区域，是在类固醇激素依赖性发育可塑性的高峰阶段收获的。当野生型神经元在体外培养时，它们会形成具有高度定型形态的乔木。该检测揭示了正常的性别二态性以及单基因突变引起的表型。在某些情况下，体外培养系统具有高度敏感性，可以揭示其体内表现过于微妙而无法通过传统显微镜检测到的缺陷。为了提高筛选速度，我们正在开发 NeuronMorphometrics，这是一组用于 ImageJ 的 Java 插件，可以执行半自动数据分析。大约 75% 的人类 MR 基因在果蝇中有一个对应基因，其序列足够相似，似乎可以在大脑中执行类似的功能。我们将筛选果蝇 MR 基因的一个子集，我们相信这些基因是在体外揭示神经元形态和可塑性缺陷的主要候选基因。在平行研究中，我们将重点关注“丝状”表型，这是一种由于细胞骨架蛋白遗传缺失而导致的体外神经突生长的异常模式，并筛选使该表型正常化的化合物。表型和校正化学试剂的筛选共同为使用该测定作为 MR 疾病药物发现和开发的垫脚石奠定了基础。公共卫生相关性：我们的长期目标是用药物治疗精神发育迟滞疾病，以修复大脑发育的潜在异常。拟议的实验是实现这一目标的重要一步。