Core B: Genome Engineering Core

核心 B：基因组工程核心

• 批准号: 10617345
• 负责人: Frederic D Bushman
• 金额: $ 16.03万
• 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/10617345
• 关键词: Adverse event; Aliquot; Behavior; Categories; Cell physiology; Cells; Chromosomal translocation; Clinical; Clonal Expansion; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; DNA; Data; Event; Evolution; Frequencies; Gene Modified; Gene therapy trial; Genes; Genetic; Genetic Transcription; Genome; Genome engineering; Genomics; Growth; HIV; HIV-1; HMGA2 gene; Human; Human Genome; IL2RG gene; 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; Therapeutic Effect; Time; 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; 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.

摘要核心 B. 量化基因组修饰的结果 Core B将追踪基因组修改的性质和后果 项目1、3和4，并利用这些数据设计针对CART基因组的改进试剂 修改。核心 B 将分析几种类型的基因组修饰，如 每个项目的需求。慢病毒载体的整合必然会破坏宿主 整合位点的细胞位点。我们和其他人发现插入 用于传递 CAR 的载体的诱变可能与改变的细胞有关 生长，这导致了临床不良事件，但也产生了临床上有利的克隆 增强治疗效果的扩展。核心将使用这些数据 确定可操作的基因和途径以优化 CART 扩展和 坚持。载体整合还独特地标记每个转导的细胞，从而允许 追踪后代细胞的分配，可用于追踪结果的信息 并了解可能的不良事件的机制。宿主的裂解 利用靶向核酸酶进行基因组测序也是实现基因组测序的有效手段 修改;然而，脱靶切割和相关缺失是安全的 的担忧。核心将使用我们的 iGUIDE 技术监控脱靶裂解。 最后，在多个 sgRNA/CAS9 复合物切割多个靶标的方案中 单细胞中，多个位点的核酸酶切割可导致 DNA 末端重新连接 “交换”时尚，导致染色体易位。核心B将监测分子 每个类别的结果可以优化试剂、评估 安全性和细胞功能评分。