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High throughput RNAi based functional genomics in primary cells and in vivo

原代细胞和体内基于高通量 RNAi 的功能基因组学

基本信息

批准号：
8870950
负责人：
Alexey Wolfson
金额：
$ 72.24万
依托单位：
ADVANCED RNA TECHNOLOGIES, LLC
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-01 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8870950
关键词：
Algorithms Area Automation Base Sequence Bioinformatics Biomedical Research Cells Chemistry Collaborations Communities Complex Custom Data Set Databases Development Drug Formulations Elements Environment Equipment and supply inventories Funding Gene Silencing Genes Genetic Transcription Genomics Goals Human Genome Immunology In Vitro Inflammation Inflammatory Libraries Maps Messenger RNA Methods Neurobiology Organ Model Pathway interactions Pharmacologic Substance Phase Process RNA RNA Interference Reagent Reporter Research Research Infrastructure Research Methodology Research Personnel Small Interfering RNA Stem cells Structure System Technology Time Tissue Model Tissues Transcript Transcriptional Regulation Transfection Validation base blastomere structure cell type commercialization cost design established cell line functional genomics gene function high throughput screening improved in vivo in vivo Model novel oncology parallel processing programs scaffold screening success text searching tool uptake web based interface

项目摘要

DESCRIPTION (provided by applicant): Completion of the HGP (human genome program) has opened the 'Genomic Era' in biomedical research and enables functional genomics analysis of various gene elements. The long-term goal of the ENCODE Project is to identify and functionally characterize all of the sequence-based genomic elements. Functional elements that have been studied in ENCODE include transcribed sequences, regulators of transcription, and regulators of RNA transcripts themselves. The function of more than 50% of transcribed gene sequences is unknown. Most of them are expressed in specialized cells and thus their functions can only be studied in a biologically relevant context (i.e. primary cells, in vivo). In this propoal we plan to develop and commercialize a new RNAi platform that enables gene studies in a biologically relevant context with a long term goal of building a functional map of human genome. Introduction of small interfering RNAs (siRNAs) into cells with transfection reagents results in efficient gene silencing. siRNA-based functional genomics is widely used in established cell lines in vitro, but its applicability to primary cells and in vivo target validatin has been limited because of lack of efficient and non-toxic delivery systems. In collaboration with RXi pharmaceuticals we have developed a novel class of RNAi compounds - self deliverable RNAs. These small, asymmetric, hydrophobically modified RNA compounds enter cells and tissues without requirement for delivery formulation and efficiently silence genes in vitro and in vivo, enabling functional genomics studies in primary cells, embryonic cells, tissues, ex vivo, and in vivo. The major technical hurdle, which impedes wide spread use of this platform by scientific community is the complex and costly process of compound identification, synthesis and validation. The focus of this fast-track proposal is optimization of compound discovery process (Phase I) and development of a panel of 200-1000 functionally validated sdRNAs (Phase II), Completion of this proposal will build a commercially available product platform that will revolutionize functional genomics studies in biologically relevant systems such as primary cells, stem cells, tissue and organ models and eventually in vivo.

描述（由申请人提供）：HGP（人类基因组计划）的完成开启了生物医学研究的“基因组时代”，并使各种基因元件的功能基因组学分析成为可能。 ENCODE 项目的长期目标是识别所有基于序列的基因组元件并对其进行功能表征。 ENCODE 中研究的功能元件包括转录序列、转录调节因子以及 RNA 转录本本身的调节因子。超过 50% 的转录基因序列的功能未知。它们中的大多数在专门的细胞中表达，因此它们的功能只能在生物学相关的背景下（即体内的原代细胞）进行研究。在此提案中，我们计划开发并商业化一个新的 RNAi 平台，该平台能够在生物学相关的背景下进行基因研究，长期目标是构建人类基因组的功能图谱。使用转染试剂将小干扰 RNA (siRNA) 引入细胞中，可实现有效的基因沉默。基于siRNA的功能基因组学广泛应用于体外建立的细胞系，但由于缺乏高效且无毒的递送系统，其对原代细胞和体内靶点验证的适用性受到限制。我们与 RXi Pharmaceuticals 合作开发了一类新型 RNAi 化合物——可自我递送的 RNA。这些小型、不对称、疏水修饰的 RNA 化合物无需递送制剂即可进入细胞和组织，并在体外和体内有效沉默基因，从而能够在原代细胞、胚胎细胞、组织、离体和体内进行功能基因组学研究。阻碍科学界广泛使用该平台的主要技术障碍是化合物鉴定、合成和验证过程复杂且成本高昂。该快速通道提案的重点是优化化合物发现过程（第一阶段）和开发一组 200-1000 个经过功能验证的 sdRNA（第二阶段）。该提案的完成将构建一个商业可用的产品平台，该平台将彻底改变生物相关系统（如原代细胞、干细胞、组织和器官模型）以及最终体内的功能基因组学研究。