CORE--RNA VECTOR

核心--RNA载体

• 批准号: 6668360
• 负责人: MICHEL SADELAIN
• 金额: $ 19.94万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6668360
• 关键词: Lentivirus; Retroviridae; biomedical facility; biotechnology; cooperative study; genetic transduction; microorganism mass culture; transfection /expression vector

Core B is designed to support oncoretroviral- and lentiviral-mediated gene transfer studies in Projects 1-5. These projects need to achieve efficient gene transfer into cells of hematopoietic origin, including dendritic cells and CD34+ hematopoietic cells. One clinical trial (in Project 5) also requires the production of a clinical grade retroviral vector. Thus, projects 1-5 require well-characterized and periodically tested high- titer producers cell lines and cell free retroviral stocks, as well as detection of replication-competent retroviral (RCR). The centralized generation of high-quality producer cell lines, retroviral stocks and biosafety testing is cost effective permits standardization of procedures, and facilitates the exchange of information and expertise between the five projects. In the pre-clinical phase of investigation, the specific aims of the RNA Core are: 1. construction of recombinant oncoretroviral or lentiviral vectors; 2. transfection of vector DNA in packaging cells and selection of producer cell lines; 3. characterization of vector transmission in standard cell lines; 4. expansion of independent clones and identification of the best clone for the intended target cells; 5. titration of cell-free viral stocks; 6. detection of replication-competent oncoretroviral in cell-free viral stocks and transduced target cells. In the clinical phase of investigation, the specific aims of the RNA Core are to carry out and/or coordinate: 1. the generation of high-titer producers, master cell banks and viral stocks for clinical studies; 2. the development of scaled up procedures for production of 5 to 15 liter batches of clinical viral stocks in semi-closed systems; 3. the development of a scaled procedure for the transduction of hematopoietic progenitor cells in semi-closed systems, in collaboration with investigators of projects 5; 4. The ability to carry out both production and transduction in the Gene Transfer and Cell Engineering Facility at MSKCC decreases the cost of generating materials and therefore the cost of clinical investigation. M. Sadelain, M.D., Ph.D., is the Director of Core B and I. Riviere, Ph.D, the Co-Director. Within the Gene Transfer and Cell Engineering Facility at MSKCC, they are currently assisted by two senior Research Assistants with considerable experience in cell culture, virology and molecular techniques and by Quality Assurance/Quality Control specialist who will review and assess all quality control issues related to the manufacturing and the release of the clinical vector and patient products.

Core B 旨在支持项目 1-5 中的肿瘤逆转录病毒和慢病毒介导的基因转移研究。这些项目需要实现有效的基因转移到造血来源的细胞中，包括树突状细胞和CD34+造血细胞。一项临床试验（项目 5）还需要生产临床级逆转录病毒载体。因此，项目 1-5 需要充分表征并定期测试的高滴度生产细胞系和无细胞逆转录病毒储液，以及复制能力逆转录病毒 (RCR) 的检测。高质量生产细胞系、逆转录病毒储备和生物安全测试的集中生成具有成本效益，允许程序标准化，并促进五个项目之间的信息和专业知识交流。在临床前研究阶段，RNA Core的具体目标是： 1.构建重组肿瘤逆转录病毒或慢病毒载体； 2.包装细胞中载体DNA的转染以及生产细胞系的选择； 3. 标准细胞系中载体传播的表征； 4. 独立克隆的扩增并鉴定针对预期靶细胞的最佳克隆； 5.无细胞病毒原液的滴定； 6.检测无细胞病毒储液和转导的靶细胞中具有复制能力的肿瘤逆转录病毒。在临床研究阶段，RNA Core 的具体目标是执行和/或协调： 1. 产生用于临床研究的高滴度生产者、主细胞库和病毒储液； 2. 开发在半封闭系统中生产 5 至 15 升批次临床病毒原液的放大程序； 3. 与项目 5 的研究人员合作，开发用于在半封闭系统中转导造血祖细胞的规模化程序； 4. 在 MSKCC 的基因转移和细胞工程设施中进行生产和转导的能力降低了生成材料的成本，从而降低了临床研究的成本。 M. Sadelain 医学博士、博士是 Core B 的主任，I. Riviere 博士是联合主任。在 MSKCC 的基因转移和细胞工程设施内，他们目前得到两名在细胞培养、病毒学和分子技术方面拥有丰富经验的高级研究助理以及质量保证/质量控制专家的协助，他们将审查和评估与临床载体和患者产品的制造和发布。