Spatial and Temporal Control of Transfection Using Ferrocene-Containing Lipids

使用含二茂铁的脂质进行转染的空间和时间控制

基本信息

批准号：
7385610
负责人：
DAVID M LYNN
金额：
$ 21.39万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-09-01 至 2009-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): This exploratory/developmental proposal seeks to develop methods for achieving active control over the transfection of cells using a ferrocene-containing cationic lipid that can be transformed electrochemically. The approach is founded on preliminary observations that lipid/DNA complexes prepared from a ferrocene- containing cationic lipid lead to high levels of cell transfection when the lipid is in a reduced redox state, but very low levels of transfection when the lipid is in the oxidized state. This critical observation suggests the basis of a general and facile approach that could be used to achieve active control over the transfection of cells in vitro or in vivo though the in situ electrochemical activation of lipoplexes. The proposed research seeks to test the hypothesis that a redox-active, ferrocene-containing cationic lipid can be used to formulate lipoplexes that are either active or inactive toward cell transfection, and that the activation of `inactive' formulations can be can be controlled externally and actively through the application of low electrical potentials. The following Specific Aims are designed to evaluate this hypothesis and determine the feasibility of this approach. They are: 1) To characterize levels of transfection and cytotoxicity in a panel of cell lines using lipoplexes formed from plasmid DNA and either the completely oxidized or completely reduced forms of a model ferrocene-containing cationic lipid (BFDMA). A second goal of this Aim seeks to evaluate differences in the ability of reduced and oxidized BFDMA to mediate transgene expression in vivo when administered to mice by tail vein injection; 2) To measure the rates at which changes in the oxidation state of BFDMA can be effected within lipoplexes and to characterize the rates of reorganization of lipoplexes upon transformation. A second goal of this Aim seeks to characterize the integrity of DNA within lipoplexes upon exposure to the electrochemical potentials used to transform the lipid; and 3) To demonstrate that electrochemical transformation of DNA/BFDMA lipoplexes in situ leads to changes in the extent of transfection of cells in vitro and in vivo. We envisage the potential outcomes of this exploratory/developmental research as having substantial fundamental and applied impacts in at least two ways: first, through the introduction of new tools for basic biological and biomedical research, and, secondly, through the introduction of methods that could ultimately be used to achieve spatial and temporal control of transfection in therapeutic or clinical contexts.

描述（由申请人提供）：该探索性/开发提案旨在开发使用可电化学转化的含二茂铁的阳离子脂质来实现对细胞转染的主动控制的方法。该方法基于初步观察，即当脂质处于还原氧化还原状态时，由含二茂铁的阳离子脂质制备的脂质/DNA复合物导致高水平的细胞转染，但当脂质处于氧化状态时，转染水平非常低状态。这一重要的观察结果表明了一种通用且简便的方法的基础，该方法可用于通过脂质复合物的原位电化学激活来实现对体外或体内细胞转染的主动控制。拟议的研究旨在检验以下假设：氧化还原活性、含二茂铁的阳离子脂质可用于配制对细胞转染有活性或无活性的脂质复合物，并且“无活性”制剂的激活可以从外部控制并积极地通过应用低电位。以下具体目标旨在评估该假设并确定该方法的可行性。它们是： 1) 使用由质粒 DNA 和完全氧化或完全还原形式的含二茂铁阳离子脂质模型 (BFDMA) 形成的脂质复合物来表征一组细胞系中的转染和细胞毒性水平。该目标的第二个目标是评估通过尾静脉注射给予小鼠时还原和氧化的 BFDMA 介导体内转基因表达的能力的差异； 2)测量脂质复合物内BFDMA氧化态变化的速率，并表征脂质复合物在转化时的重组速率。该目标的第二个目标是在暴露于用于转化脂质的电化学势时表征脂质复合物内 DNA 的完整性； 3) 证明DNA/BFDMA lipoplex的原位电化学转化会导致体外和体内细胞转染程度的变化。我们预计这项探索性/开发性研究的潜在成果至少会在两个方面产生重大的基础和应用影响：首先，通过引入用于基础生物学和生物医学研究的新工具，其次，通过引入可以最终用于在治疗或临床环境中实现转染的空间和时间控制。