Investigate stability of RNA interference in human lymphocytes

研究人类淋巴细胞中 RNA 干扰的稳定性

• 批准号: 7290306
• 负责人: Dong Sung An
• 金额: $ 33.75万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-30 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7290306
• 关键词: Address; Adverse effects; Affect; Animal Model; Animals; Apoptosis; Basic Science; Biological Models; CCR5 gene; Cells; Chemokine (C-C Motif) Receptor 5; Chronic Disease; Cytoprotection; Data; Development; Down-Regulation; Follow-Up Studies; Future; Gene Expression; Genetic; Genome; Goals; HIV-1; Hematopoietic stem cells; Human; In Vitro; Lentivirus Vector; Lymphocyte; Lymphoid Cell; Maintenance; Mammalian Cell; Mediating; Modeling; Pathway interactions; Population; Process; Public Health; RNA Interference; Relative (related person); Research; Research Personnel; Retroviral Vector; Small Interfering RNA; Stem cell transplant; T-Lymphocyte; Technology; Testing; Therapeutic; Therapeutic Intervention; Time; Toxic effect; base; cancer therapy; cellular transduction; cytotoxic; cytotoxicity; desire; experience; gene therapy; immunoregulation; in vivo; mouse model; novel; peripheral blood; programs; reconstitution; small hairpin RNA; therapeutic gene; transgene expression; vector

DESCRIPTION (provided by applicant): Our overall hypothesis is that RNA interference (RNAi) can be developed to reduce specific gene expression over long-term without toxic effects in human lymphocytes. RNAi using siRNA to inhibit specific gene expression is a powerful and promising technology for both basic research and therapeutic intervention. However, our results and several recent studies have shown unintended nonspecific cytotoxic effects associated with this technology in mammalian cells raising a concern for the long term use of the RNAi in therapeutic applications in humans. The cytotoxic effects were attributed at least in part to apoptosis, although other mechanisms are also possible. The cytotoxicity was associated with higher levels of shRNA expression. Based upon these results, we hypothesized that that the levels of shRNA are important for determining whether a given shRNA will be cytotoxic or not and that optimization of those levels can yield shRNAs that are both effective and not or minimally cytotoxic. We will also address whether the cell localization of the shRNA and/or its pathway for processing can affect the extent of cytotoxicity within cells. Through our proposed studies we will understand the effects of shRNA expression in mature lymphoid cells in vitro as well as the effects on the differentiation of the lymphocyte populations in an in vivo setting. We will minimize the cytotoxic effect and maximize stable and effective function of RNAi in primary T-lymphocytes. The relevance of this research to public health. RNA interference is a powerful and promising technology to inhibit specific gene expression for both basic research and therapeutic intervention. The inhibition of specific gene expression needs to be effective without unintended side effects. This proposal propose to study the effects of RNAi in human T lymphocytes to maximize the RNAi effect and minimize side effects.

描述（由申请人提供）：我们的总体假设是可以开发RNA干扰（RNAi）以减少长期的特定基因表达，而没有人类淋巴细胞的毒性作用。使用siRNA抑制特定基因表达的RNAi是基础研究和治疗干预的强大而有前途的技术。但是，我们的结果和最近的一些研究表明，与该技术在哺乳动物细胞中相关的非特异性细胞毒性作用引起了人们长期使用RNAi在人类治疗应用中的关注。细胞毒性作用至少部分归因于细胞凋亡，尽管其他机制也是可能的。细胞毒性与较高水平的shRNA表达有关。基于这些结果，我们假设shRNA的水平对于确定给定的shRNA是否具有细胞毒性很重要，并且这些水平的优化可以产生既有效又不微的细胞毒性的shRNA。我们还将解决shRNA的细胞定位和/或其加工途径是否会影响细胞内细胞毒性的程度。通过我们提出的研究，我们将了解体外淋巴样细胞中shRNA表达的影响，以及对体内环境中淋巴细胞种群分化的影响。我们将最大程度地减少细胞毒性效应，并最大程度地提高RNAi在原代T淋巴细胞中的稳定和有效功能。这项研究与公共卫生的相关性。 RNA干扰是一种强大而有希望的技术，可抑制基础研究和治疗干预的特定基因表达。在没有意外副作用的情况下，对特定基因表达的抑制需要有效。该建议建议研究RNAi在人T淋巴细胞中的影响，以最大程度地提高RNAi效应并最大程度地减少副作用。