Electric Field - Forced Gene Transfer In Solid Tumors

电场 - 实体瘤中的强制基因转移

• 批准号: 6475237
• 负责人: FAN YUAN
• 金额: $ 32.58万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-11 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6475237
• 关键词: apoptosis; athymic mouse; biotechnology; cell line; collagen; electric field; electroporation; endopeptidases; experimental designs; extracellular matrix; gene delivery system; gene therapy; interleukin 12; intracellular transport; method development; mucopolysaccharides; neoplasm /cancer therapy; plasmids; skin transplantation; transfection; transposon /insertion element

Description (provided by applicant): The overall objective of this application is to improve electric field-forced gene transfer in solid tumors. Gene therapy has a great potential to improve treatment of different diseases, including solid tumors, but it has been impeded by the difficulties of gene delivery to specific cells. Recent reports by other investigators and ourselves have demonstrated that plasmid DNA delivery can be enhanced through the use of pulsed electric fields. However, present methods for optimizing electric field-forced gene transfer, in terms of electric pulse characteristics (e.g., field strength and pulse duration), are still empirical and incomplete. The fundamental hypothesis in this proposal is that electric field-forced DNA transport in the vicinity of cells determines the amount of gene transfer through the transient pores in the plasma membrane of cells created during in vivo electroporation. To test this hypothesis, we will systematically quantify the mechanisms of interstitial transport of DNA in response to different electric pulses both ex vivo (Aim 1) and in vivo (Aim 2). The quantification will be based on unique experimental methods developed in our labs, and it may result in an identification of effective pulse sequences for improving interstitial transport of DNA. The improved transport in the identified electric fields will be further increased through modifications in tumor tissue structures, using matrix enzymes, hypertonic solutions, or apoptotic agents (Aim 3). These chemical treatments transiently increase the interstitial space and thus decrease the resistance to DNA transport. Quantitative results from the transport studies in Specific Aims 1 through 3 will finally be used to improve transfection efficiency and therapeutic efficacy of interleukin- 12 gene in solid tumors transplanted in mice (Aim 4). The goal of the proposed study is to establish effective experimental protocols for improving interstitial transport of plasmid DNA which in turn may result in an increase in transfection efficiency in solid tumors without increasing toxicity.

描述（申请人提供）：本申请的总体目的 是为了改善实体瘤中的电场强制基因转移。基因治疗 具有改善不同疾病治疗的巨大潜力，包括 实体瘤，但它一直受到基因传递的困难， 特定细胞其他调查人员和我们自己最近的报告显示， 证明了质粒DNA递送可以通过使用 脉冲电场然而，目前用于优化电性能的方法 场致基因转移，就电脉冲特性而言（例如， 场强和脉冲持续时间）仍然是经验的和不完整的。的 这一提议的基本假设是电场强迫DNA 在细胞附近的转运决定了基因转移的量 通过细胞质膜上的瞬时孔， 体内电穿孔。为了验证这一假设，我们将系统地量化 不同的细胞因子对DNA间质转运的影响 体外（Aim 1）和体内（Aim 2）的电脉冲。量化 将基于我们实验室开发的独特实验方法， 导致有效脉冲序列的识别， DNA间质转运。经查明的 电场将通过肿瘤组织中的修饰而进一步增加 结构，使用基质酶、高渗溶液或凋亡剂 (Aim 3）。这些化学处理短暂地增加了间隙空间 从而降低对DNA转运的阻力。定量结果来自 具体目标1至3中的运输研究最终将用于 提高白细胞介素-12转染效率和治疗效果 基因在小鼠移植的实体瘤中的表达（目的4）。建议的目标 研究的目的是建立有效的实验方案， 质粒DNA的间质转运，这反过来可能导致 在不增加毒性的情况下提高实体瘤的转染效率。