Design of Gene Delivery System to Target Hepatocytes

描述(由申请人提供)： 非病毒(合成)核酸递送系统有可能提供基于核酸的疗法的实际应用。我们设计并创造了一种基于环糊精聚合物(CDP)的可调、自组装、非病毒的核酸递送系统，该递送系统已被证明在动物中具有非常低的毒性，并通过静脉给药在靶向组织中提供基因表达。 我们建议评估这样一种假设，即我们精心设计和设计的、合成的、基于半乳糖的靶向部分的非病毒载体可以有效地将核酸输送到小鼠的肝细胞，前提是颗粒直径不超过70 nm，表面电荷在+/-10 mV范围内。 具体目标： 1.用直径约70 nm或更小的基于半乳糖的靶向配体、表面电荷在+/-10 mV范围内、可被肝细胞上的去唾液酸糖蛋白受体识别和处理的含有siRNA的CDP颗粒，并确定在小鼠全身给药后最大限度地下调肝脏基因的最佳物理化学性质。 2.利用特定目的1中的研究结果，制备含PDNA(PDNA)的CDP为基础的颗粒，以指导组装和给药程序，并利用标记基因和治疗基因测定基因在小鼠肝细胞中的基因传递效率和基因表达的时程。 意义：以肝细胞为靶点的有效的合成递送载体的开发将为治疗多种疾病提供一种通用的方法。 长期目标：使用基于核酸的疗法，即所谓的基因疗法，提供新的疾病治疗的潜力受到安全和有效输送核酸的限制。我们建议的长期目标是设计和设计一个通用的、合成的系统，最终将基于核酸的治疗药物输送到人类的肝细胞。

DESCRIPTION (provided by applicant): Non-viral (synthetic) nucleic acid delivery systems have the potential to provide for the practical application of nucleic acid-based therapeutics. We have designed and created a tunable, self-assembling, non-viral nucleic acid delivery system that is based on cyclodextrin-containing polymers (CDP) and this delivery system has been shown to have very low toxicity in animals and give gene expression in targeted tissues from an intravenous administration. We propose to evaluate the hypothesis that our properly designed and engineered, synthetic, non-viral delivery vehicle bearing galactose-based targeting moieties can effectively deliver nucleic acids to hepatocytes in mice if the particle diameters do not exceed 70 nm and the surface charges are in the range of +/-10 mV. Specific Aims: 1. Formulate siRNA-containing, CDP-based particles with galactose-based targeting ligands of ca. 70 nm diameter or less with surface charges within the range of +/-10mV that can be recognized and processed by the asialoglycoprotein receptor on hepatocytes and determine the optimal physicochemical properties for maximum downregulation time and efficiency of a hepatic gene following systemic administration to mice. 2. Formulate plasmid DNA (pDNA)-containing, CDP-based particles using the conclusions obtained in Specific Aim 1 to guide the assembly and administration protocols and determine the gene delivery efficiency and the time course of gene expression in hepatocytes of mice using both marker and therapeutic genes. Significance: The development of effective, synthetic delivery vehicles for targeting hepatocytes would provide a generalized methodology for treating numerous diseases. Long-term goals: The potential for providing new disease treatments using nucleic acid-based therapies, the so-called gene therapies, has been restricted by limitations in the safe and effective delivery of nucleic acids. The long-term objective of our proposal is to design and engineer a generalized, synthetic system for ultimately delivering nucleic acid-based therapeutics to hepatocytes in humans.