Novel Oligonucleotide Delivery Vehicles for Gene Tharapy

用于基因治疗的新型寡核苷酸递送载体

• 批准号: 6630988
• 负责人: TJ THOMAS
• 金额: $ 31.21万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-01-15 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6630988
• 关键词: MCF7 cell; antisense nucleic acid; atomic force microscopy; breast neoplasms; circular dichroism; electron microscopy; fluorescence spectrometry; gene expression; gene therapy; genetic transcription; genetically modified animals; high performance liquid chromatography; laboratory mouse; light scattering; messenger RNA; neoplasm /cancer therapy; neoplastic cell; northern blottings; nucleic acid metabolism; nucleic acid structure; oligonucleotides; polyamines; protooncogene; transfection; western blottings

DESCRIPTION (provided by applicant): The overall goal of this project is to elucidate the mechanism of action of polyamine analogues in facilitating the uptake and function of antisense oligonucleotides (ODNs). The hypothesis is that the selectivity of polyamine analogues in DNA condensation and/or stabilization of RNA.DNA hybrids can be utilized to facilitate the cellular uptake and function antisense ODNs for breast cancer therapy. Antisense ODNs targeted to c-myc and HER-2 genes will be studied. To evaluate the structural specificity effects of polyamine analogues on RNA.DNA hybrid stabilization, association constants will be determined using a molecular beacon technique. The role of ODN nanoparticles in facilitating cellular uptake of ODNs will be quantified by dynamic light scattering as well as electron, scanning force and polarizing spectroscopy. Cellular uptake and function of ODNs will be measured using MCF-7 and SK-BR-3 breast cancer cells. ODN uptake will be determined using radioactive and fluorescent ODNs. Effects of polyamine analogue/ODN combinations on target gene expression, cell growth arrest, and apoptosis will be examined to demonstrate enhanced function of ODNs. Subsequently, HER-2 gene targeted ODN and selected polyamine analogues will be examined for in vivo ODN delivery and function using a transgenic mouse model of breast cancer. The effects of polyamine analogues and ODNs on natural polyamines will be quantified by HPLC. These studies will advance our knowledge of the mechanism of cellular uptake of ODNs, and help to develop novel approaches for breast cancer therapy.

描述（由申请人提供）：该项目的总体目标是阐明多胺类似物促进反义寡核苷酸（ODN）的摄取和功能的作用机制。假设是，多胺类似物在 DNA 缩合和/或 RNA.DNA 杂交体稳定中的选择性可用于促进细胞摄取和发挥反义 ODN 的功能，用于乳腺癌治疗。将研究针对 c-myc 和 HER-2 基因的反义 ODN。为了评估多胺类似物对 RNA.DNA 杂交稳定性的结构特异性影响，将使用分子信标技术确定关联常数。 ODN 纳米颗粒在促进细胞摄取 ODN 方面的作用将通过动态光散射以及电子、扫描力和偏振光谱进行量化。将使用 MCF-7 和 SK-BR-3 乳腺癌细胞测量 ODN 的细胞摄取和功能。 ODN 摄取量将使用放射性和荧光 ODN 来确定。将检查多胺类似物/ODN 组合对靶基因表达、细胞生长停滞和细胞凋亡的影响，以证明 ODN 的增强功能。随后，将使用乳腺癌转基因小鼠模型检查 HER-2 基因靶向 ODN 和选定的多胺类似物的体内 ODN 递送和功能。多胺类似物和 ODN 对天然多胺的影响将通过 HPLC 进行定量。这些研究将增进我们对 ODN 细胞摄取机制的了解，并有助于开发乳腺癌治疗的新方法。