SYNTHETIC VEHICLES FOR TARGETED GENE DELIVERY IN VIVO

利用细胞表面受体传递基因的能力 在体内的器官特异性靶向已经被证明。这种方法 拥有许多令人兴奋和有效的疗法的希望。这个 该项目的目标是开发化学定义的递送 将基因高效地送入细胞核的系统 首先是在体外，然后是在体内。 具体目标1是通过有机合成制备DNA结合 含有高亲和力、化学定义的碳水化合物的多肽 肝脏和肝脏去唾液酸糖蛋白受体的受体配体 骨骼肌中的甘露糖-6-磷酸受体。物理和 将使用化学方法来表征结构， DNA络合物的热力学和动力学性质。 具体目标2是确定DNA内吞后的配体 复合体及其释放到细胞质中，什么样的结构和 复合体中的组成变化是DNA移动所必需的 通过细胞质进入细胞核。DNA在细胞质中的运动， DNA在细胞核内的积累，以及可能的解离 细胞质中的复合体将使用生化方法和 数字成像荧光显微镜。 具体目标3是合成DNA结合配体，以促进 DNA通过细胞质进入细胞核的运输。SV-40 将测试DNA结合多肽上的核定位序列 作为DNA复合体的一部分。 具体目标4是证明构建在 在体外实验的基础上，对基因传递是有效的 活着。

The ability to deliver genes using cell surface receptors to achieve organ specific targeting in vivo has been demonstrated. This approach holds the promise of many exciting and effective therapies. The objective of this project is to develop chemically defined delivery systems that provide highly efficient delivery of genes into the nucleus of the targeted cells, first in vitro and then in vivo. Specific Aim 1 is to prepare, by organic synthesis, DNA binding polypeptides containing high affinity, chemically defined carbohydrate receptor ligands for the asialoglycoprotein receptor in the liver and for the mannose-6-phosphate receptor in skeletal muscle. Physical and chemical methods will be used to characterize the structural, thermodynamic, and kinetic properties of the DNA complexes. Specific Aim 2 is to determine, after endocytosis of the DNA:ligand complex and its release into the cytoplasm, what kind of structural and compositional changes in the complex are necessary for the DNA to move through the cytoplasm into the nucleus. DNA movement in the cytoplasm, DNA accumulation in the nucleus, and the probable dissociation of the complex in the cytoplasm will be measured using biochemical methods and digital imaging fluorescence microscopy. Specific Aim 3 is to synthesize DNA binding ligands that facilitate transport of the DNA through the cytoplasm into the nucleus. The SV-40 nuclear localization sequence on a DNA binding polypeptide will be tested as a component of the DNA complex. Specific Aim 4 is to demonstrate that the DNA complexes, constructed on the basis of in vitro experiments, are effective for gene delivery in vivo.