Efficient and Selective Delivery of Oligonucleotides

通过基因修饰调节细胞行为的能力作为一种治疗策略具有巨大的潜力，同时也为阐明基因功能（所谓的功能基因组学）提供了强大的工具。反义寡核苷酸是最常见的长度为15-25个核苷酸的单链DNA分子，通过与靶基因mRNA上的互补片段结合来调节基因表达。虽然反义技术正在成为功能基因组学的一种可行的治疗实体和平台，但其广泛应用的主要障碍仍然存在：遗传物质（多核酸）以生物有效的数量和功能完整且无毒的形式传递到细胞。我们的总体目标是选择性地将反义分子传递到目标细胞类型（肝细胞），从而低剂量抑制感兴趣基因的表达。为了实现这一目标，我们将使用组合合成方法开发一种新的多功能DNA传递载体（multiplexes）。这些载体将具有浓缩DNA的仿生聚合物，破坏细胞膜稳定的阳离子肽，以及靶向肝细胞（原代肝细胞）的半乳糖部分。载体将以大小、稳定性和细胞毒性为特征。我们将研究这些材料对靶细胞和非靶细胞的吸附，并在胶体化学数学模型的框架下解释结果，该模型将用于改进和优化载体的组成。这些载体传递基因表达调节反义寡核苷酸的有效性将被评估和评估，以进一步完善该方法。我们期望长期结果是一种选择性和有效的方法，用于治疗和功能基因组学应用的寡核苷酸递送。

The ability to modulate cell behavior through genetic modification has great potential as a therapeutic strategy, as well as providing a powerful tool for elucidating gene function (so-called functional genomics). Antisense oligonucleotides, which are most commonly single-stranded DNA molecules 15-25 nucleotides in length, modulate gene expression by binding to a complementary segment on the mRNA from the target gene. While antisense technology is becoming a viable therapeutic entity and platform for functional genomics, a major barrier to its widespread practice still exists: the delivery of the genetic material (polynucleic acid) to cells in a quantity that is biologically effective and in a form that is functionally intact, yet non-toxic. Our overall goal is to deliver antisense molecules selectively to a target cell type (hepatocytes), resulting in low-dose inhibition of expression of genes of interest. To achieve this goal, we will develop a new family of multifunctional DNA delivery vectors (multiplexes) using a combinatorial synthesis approach. These vectors will possess biomimetic polymers that condense DNA, cationic peptides that destabilize cellular membranes, and galactose moieties that target them to hepatocytes (primary liver cells). Vectors will be characterized for size, stability, and cytotoxicity. We will study the adsorption of these materials to target vs. non-target cells, and interpret the results in the framework of a colloid-chemical mathematical model, which will be used to refine and optimize the composition of the vectors. The effectiveness of these vectors to deliver gene expression-modulating antisense oligonucleotides will be evaluated and assessed to further refine the approach. W expect the long-term outcome to be a selective and efficient method for oligonucleotide delivery for therapeutic and functional genomics applications.