Zinc Finger Proteins to Modify Heart Function

锌指蛋白可改变心脏功能

• 批准号: 7150124
• 负责人: FRANK J GIORDANO
• 金额: $ 41.25万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7150124
• 关键词: DNA binding protein; RNA splicing; angiogenesis; biotechnology; chromatin; disease /disorder model; gene delivery system; gene expression; gene induction /repression; gene targeting; gene therapy; genetically modified animals; heart function; ischemia; laboratory mouse; laboratory rat; leg; myocardial infarction; phospholamban; protein engineering; protein structure; stoichiometry; transcription factor; vascular endothelial growth factors

DESCRIPTION (provided by applicant): This proposal is based on the novel approach of designing an engineering novel zinc finger protein transcription factors (ZFPs) to regulate the expression of endogenous genes for the treatment of cardiovascular disease. We have previously validated this approach in vivo and in vitro with engineered ZFPs that activate expression of the VEGF-A gene. We now propose further studies to more vigorously investigate the mechanism of action of our ZFP constructs and to delineate the differences in the biological effects of endogenous gene activation vs. cDNA-based transgene expression. In the context of these experiments we study the determinants of ZFP half-life and the effects of differential ZFP positional DNA- binding on the stoichiometry of VEGF splice variant expression. We will also investigate ZFP-induced gene repression using a ZFP construct engineered to target and repress the phospholamban (PLB) gene. PLB is a critical determinant of cardiac contractility, and the ability to downregulate the expression of PLB has significant clinical implications. In the context of these experiments we will also study the relationship between the physical half-life of the PLB repressor ZFP and the functional half-life of PLB repression. These experiments represent the first time an engineered transcription repressor will have been used in a clinically relevant in vivo context. Finally, we will further build on work we have recently completed demonstrating the feasibility of building cell- permeable ZFPs that are capable of crossing cell membranes, entering the nucleus and regulating the expression of specifically targeted genes. This represents a novel new therapeutic approach that also has significant clinical implications.

描述（由申请人提供）：该建议基于设计工程新型锌指蛋白转录因子（ZFP）的新方法，以调节内源基因的表达以治疗心血管疾病。我们先前曾在体内和体外验证了这种激活VEGF-A基因表达的工程ZFP。现在，我们提出进一步的研究，以更积极地研究ZFP构建体的作用机理，并描述内源基因激活与基于cDNA的转基因表达的生物学作用的差异。在这些实验的背景下，我们研究了ZFP半衰期的决定因素以及差异ZFP位置DNA结合对VEGF剪接变体表达的化学计量的影响。我们还将使用设计用于靶向和抑制Phospholamban（PLB）基因的ZFP构造的ZFP诱导的基因抑制。 PLB是心脏收缩性的关键决定因素，下调PLB表达的能力具有显着的临床意义。在这些实验的背景下，我们还将研究PLB阻遏物ZFP的物理半衰期与PLB抑制的功能半衰期之间的关系。这些实验代表了工程转录阻遏物首次在体内与临床相关的情况下使用。最后，我们将进一步建立我们最近完成的工作，证明了能够跨越细胞膜，进入细胞核并调节特定靶向基因的表达的可渗透性ZFP的可行性。这代表了一种新型的新治疗方法，它也具有重要的临床意义。