Characterization and Early Assays Development in FOXG1 Deficient Neurons

FOXG1 缺陷神经元的表征和早期检测开发

• 批准号: 10727503
• 负责人: JED L HUBBS
• 金额: $ 17.7万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10727503
• 关键词: Biological Assay; Boston; CRISPR/Cas technology; Cell Differentiation process; Cell Line; Cells; Clinical; Development; Developmental Delay Disorders; Disease; Dyes; Ethics; Fluorescent Dyes; Future; Gene Proteins; Genes; Genetic; Genotype; Goals; Heterozygote; Image; Individual; Interneurons; Ligands; Measures; Methods; Microcephaly; Movement; Muscle hypotonia; Mutation; Neurodevelopmental Disorder; Neurologic Symptoms; Neurons; Parents; Pathway interactions; Patients; Pediatric Hospitals; Pharmaceutical Preparations; Pharmacotherapy; Pluripotent Stem Cells; Protein Deficiency; Proteins; Reporter; Reporter Genes; Reporting; Research; Seizures; Somatic Cell; Stereotyping; Symptoms; Syndrome; System; Telencephalon; Validation; Work; assay development; drug discovery; gene therapy; genomic locus; induced pluripotent stem cell; interest; mutant; nerve stem cell; nervous system disorder; postnatal development; pre-clinical; programs; protein expression; screening; severe intellectual disability; small molecule; small molecule therapeutics; stem cell proliferation; symptomatic improvement; therapeutic development; therapy development; transcription factor

PROJECT SUMMARY FOXG1 syndrome is a rare neurological disorder that often causes severe intellectual disability, microcephaly and neurological symptoms including hypotonia, seizures, and stereotypic movements. FOXG1 syndrome is caused by heterozygous mutation or deletion of the forkhead box 1 protein (FOXG1), a transcription factor that controls expression of key cortical development proteins. FOXG1 is specifically important for telencephalon neural progenitor cell (NPC) proliferation and differentiation to GABAergic interneurons. We have prioritized FOXG1 syndrome based on a framework we developed to evaluate monogenetic neurodevelopmental disorders for therapeutic development based on generic, preclinical validation, clinical and ethical considerations. We have also created three pluripotent stem cell (PSC) lines from FOXG1 syndrome patients and used CRISPR/Cas9 gene editing to create isogenic control lines. We hypothesize that a drug-like small molecule that increases FOXG1 expression in patients during early postnatal development would improve symptoms in these individuals. In the proposed research, we will use FOXG1 PSC lines to create an endogenous locus gene reporter line by using CRISPR/Cas9 to insert a HaloTag reporter protein at the C-terminus of the FOXG1 gene. A method will be developed whereby these modified FOXG1 PSCs with be differentiated to cortical neural progenitor cells (NPCs) and treated with HaloTag ligand tethered dyes to measure accurately measure FOXG1 expression. The primary goal of this project will be to demonstrate that this FOXG1 reporter system is suitable for screening small molecules for their ability to increase FOXG1 expression.

项目总结 FOXG1综合征是一种罕见的神经疾病，通常会导致严重的智力残疾， 小头畸形和神经症状，包括低眼压、癫痫发作和刻板印象 动静。FOXG1综合征是由叉头杂合突变或缺失引起的 盒1蛋白(FOXG1)，一种控制关键皮质发育表达的转录因子 蛋白质。FOXG1对端脑神经前体细胞(NPC)具有特别重要的作用 GABA能中间神经元的增殖和分化。我们已经优先考虑FOXG1综合征 基于我们开发的评估单基因神经发育障碍的框架 基于仿制药、临床前验证、临床和伦理的治疗进展 考虑因素。我们还从FOXG1中创造了三个多能干细胞(PSC)系 并使用CRISPR/Cas9基因编辑创建等基因对照系。我们 假设一种增加患者体内FOXG1表达的类药物小分子 出生后早期发育会改善这些人的症状。在建议的 研究表明，我们将利用FOXG1PSC系通过以下途径创建一个内源基因座报告系 利用CRISPR/Cas9在FOXG1基因的C末端插入HaloTag报告蛋白。 将开发一种方法，通过该方法将这些修改的FOXG1 PSCs区分为 皮质神经前体细胞及HaloTag配体系带染料处理后的检测 精确测量FOXG1的表达。该项目的主要目标将是演示 这种FOXG1报告系统适合于筛选小分子的能力 增加FOXG1的表达。