Gene Silencing and Gene Editing in Phototransduction

光转导中的基因沉默和基因编辑

基本信息

批准号：
10437001
负责人：
Stephen H Tsang
金额：
$ 39.04万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10437001
关键词：
Ablation Address Adverse effects Affect Alleles American Base Sequence Biological Models CRISPR interference CRISPR therapeutics CRISPR/Cas technology Cell Culture Techniques Cell Cycle Clinical Clustered Regularly Interspaced Short Palindromic Repeats Codon Nucleotides DNA Sequence Defect Development Disease FDA approved Feasibility Studies Funding Future G2 Phase Gene Expression Regulation Gene Silencing Gene therapy trial Genes Genomics Goals Guide RNA Human Inherited Investigational New Drug Application Knock-in Link Longevity Mediating Methods Modeling Modification Molecular Mus Mutation Pathology Pharmaceutical Preparations Phase Phenocopy Phototransduction Pre-Clinical Model Preclinical Testing Proteins RNA Interference RNA Interference Therapy Research Resources Retina Retinitis Pigmentosa Rhodopsin S phase Safety Supplementation Technology Testing Therapeutic Toxic effect Transgenic Mice Translational Research Viral Vector adeno-associated viral vector base blind clinically relevant experimental study gain of function mutation gene augmentation therapy gene replacement gene therapy humanized mouse in vivo Model individualized medicine innovation loss of function mouse model mutant novel pre-clinical precision medicine preclinical evaluation repaired restoration therapeutic gene therapeutic genome editing treatment response vector

项目摘要

Project Summary Gene-augmentation therapy cannot address autosomal-dominant (ad) disorders, in which a defective protein is expressed. To treat these mutations, such as the dominant form of rhodopsin (RHO)-related retinitis pigmentosa RP (adRP), the only cure is to silence (RNAi) or edit the mutant allele. Precise CRISPR therapeutic editing is predicted to be the best means of treating several dominant disorders, but molecular methods harnessing CRISPR have been slow to develop because homology-directed repair (HDR) occurs only during the S and G2 phases of the cell cycle. To overcome these limitations, we have established a versatile editing technology that uses 2 guide RNAs (gRNAs) to target all 150 dominant RHO mutations at any phase of the cell cycle. To conduct preclinical evaluation of this species-specific technology, we have replaced the naïve mouse chromosomal Rho with human RHO mutant sequences (humanized RHO). If successful, the viral vectors used in our therapeutic editing strategy could be directly reused in human trials for adRP without further modification, thus avoiding FDA hurdles and accelerating translation of this research to a clinical setting. Our preclinical adRP model could also be useful for testing of future gene- and drug-based therapies. The proposed research is innovative, as it introduces new methods and model systems as the initial steps of a precision medicine approach toward developing adRP treatment. These methods are also potentially adaptable to therapies based on targeting DNA sequences of genes linked to other dominant disorders.

项目概要基因增强疗法无法解决常染色体显性（AD）疾病，其中表达有缺陷的蛋白质来治疗这些突变，例如视紫红质的主要形式。 (RHO) 相关视网膜色素变性 RP (adRP)，唯一的治疗方法是沉默 (RNAi) 或编辑突变体精确的 CRISPR 治疗编辑预计将是治疗多种等位基因的最佳方法。显性疾病，但利用 CRISPR 的分子方法发展缓慢因为同源定向修复 (HDR) 仅发生在细胞的 S 期和 G2 期为了克服这些限制，我们建立了一种使用多功能编辑技术。 2 引导 RNA (gRNA) 在细胞周期的任何阶段靶向所有 150 个显性 RHO 突变。为了对这种物种特异性技术进行临床前评估，我们已经取代了朴素的技术具有人 RHO 突变序列的小鼠染色体 Rho（人源化 RHO）如果成功，我们的治疗编辑策略中使用的病毒载体可以直接在人体试验中重复使用无需进一步修改即可用于 adRP，从而避免 FDA 障碍并加速转化我们的临床前 adRP 模型也可用于测试临床环境。未来的基于基因和药物的疗法是创新的，因为它介绍了。新方法和模型系统作为精准医学方法的第一步这些方法也可能适用于基于 AdRP 的治疗。靶向与其他显性疾病相关的基因 DNA 序列。