Core B: Genome Engineering Core

核心 B：基因组工程核心

• 批准号: 9891734
• 负责人: Frederic D Bushman
• 金额: $ 16.2万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-15 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9891734
• 关键词: Adverse event; Aliquot; Behavior; Categories; Cell physiology; Cells; Chromosomal translocation; Cleaved cell; Clinical; Clonal Expansion; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; DNA; Data; Event; Evolution; Frequencies; Gene therapy trial; Gene-Modified; Genes; Genetic; Genetic Transcription; Genome; Genome engineering; Genomics; Growth; HIV; HIV-1; HMGA2 gene; Human; Human Genome; Immunotherapy; Infusion procedures; Insertional Mutagenesis; Lentivirus Vector; Methods; Modification; Molecular; Monitor; Nature; Outcome; Pathway interactions; Patients; Procedures; Protocols documentation; Reagent; Recurrence; Safety; Site; Specificity; T-Cell Proliferation; T-Lymphocyte; Technology; Therapeutic Effect; Time; base; beta Thalassemia; cell growth; cellular transduction; clinical efficacy; design; engineered T cells; experience; experimental study; gene therapy; genotoxicity; human subject; improved; integration site; lentiviral integration; nuclease; safety assessment; targeted nucleases; therapeutic cloning; vector

Abstract Core B. Quantifying outcomes of genome modification Core B will track the nature and consequences of genome modifications carried out in Projects 1, 3 and 4, and use these data to design improved reagents for CART genome modification. Core B will analyze several types of genome modification as dictated by the needs of each project. Integration of lentiviral vectors necessarily disrupts the host cell locus at the site of integration. We and others have found that insertional mutagenesis by vectors used to deliver CARs can be associated with altered cell growth, which has resulted in clinical adverse events, but also clinically favorable clonal expansions that have bolstered therapeutic effects. The core will use these data to identify genes and pathways to manipulate to optimize CART expansion and persistence. Vector integration also marks each transduced cell uniquely, allowing tracing of the partitioning of descendant cells, information useful in tracking outcomes and understanding mechanisms of possible adverse events. Cleavage of the host genome using targeted nucleases is also an effective means of achieving genome modification; however, off-target cleavage and associated deletions are safety concerns. The core will monitor off-target cleavage using our iGUIDE technology. Lastly, in protocols where multiple sgRNA/CAS9 complexes cleave multiple targets in a single cell, nuclease cleavage at multiple sites can result in rejoining of DNA ends in a “swapped” fashion, causing chromosomal translocations. Core B will monitor molecular outcomes in each of these categories to allow optimization of reagents, assessment of safety, and scoring of cell function.

抽象核心B。量化基因组修饰的结果 核心B将跟踪基因组修饰的性质和后果， 项目1、3和4，并使用这些数据设计用于CART基因组的改进试剂 改性核心B将分析几种类型的基因组修饰， 每个项目的需要。慢病毒载体的整合必然会破坏宿主 在整合位点的细胞位点。我们和其他人发现， 通过用于递送汽车的载体的诱变可以与改变的细胞特性相关。 生长，这导致了临床不良事件，但也有临床有利的克隆 扩大了治疗效果。核心将使用这些数据， 鉴定基因和途径以进行操作以优化CART扩增， 坚持不懈载体整合还独特地标记每个转导的细胞， 追踪后代细胞的划分，在追踪结果中有用的信息 并了解可能的不良事件的机制。宿主分裂 使用靶向核酸酶的基因组也是实现基因组测序的有效手段。 然而，脱靶切割和相关缺失是安全的， 性问题核心将使用我们的iGUIDE技术监测脱靶裂解。 最后，在其中多个sgRNA/CAS9复合物以多个靶切割多个靶的方案中， 单个细胞，核酸酶在多个位点的切割可导致DNA末端以 “交换”的方式导致染色体易位核心B将监测分子 这些类别中的每一个的结果，以允许优化试剂，评估 安全性和细胞功能评分。