核酸是生物体内最重要的生物大分子之一。可控的核酸功能调控对维持正常细胞和器官功能的动态平衡以及基因相关疾病（特别是肿瘤）的治疗具有重要的意义。本项目拟设计、合成一系列针对肿瘤细胞过表达的蛋白酶（如MMP-2和Cathepsin B）响应的发夹结构反义核酸。将反义链核酸序列通过对上述蛋白酶响应的多肽序列连接另一段正义链核酸序列的抑制链来构建发夹结构的反义核酸。优化抑制正义链的不同序列和配对碱基，筛选出结构稳定且暂时不具有活性的发夹结构功能反义核酸。在溶液和特定肿瘤细胞水平上，系统地评价其在特定蛋白酶存在下，反义核酸的活化和对靶基因的特异性调控，阐明酶响应反义核酸的结构、热稳定性与酶激活的基因调控之间的相关关联，为在动物水平上的特定肿瘤的酶响应性反义核酸药物前药的研究奠定基础。

Nucleic acid is one of the most important biomolecules. Controllable regulation of a specific nucleic acid function is extremely important to maintain the dynamic balance of cellular and organ functions as well as disease therapy (esp. cancers). The proposal is aimed to design and synthesize a new series of enzyme-responsive antisense oligonucleotides with hairpin structure and peptide loops that could be cleaved by specific proteinases (such as MMP-2 and Cathepsin B). By varying the paired positions and complementary sequences of inhibitory sense strand oligonucleotides, the enzyme-responsive antisense oligonucleotides are optimized and their antisense activities are temporally masked until the proteinases cleaved the peptide substrates to release the antisense oligonucleotides. We will evaluate the enzyme-responsive antisense oligonucleotides in solution and cancer cells, and demonstrate the relationship among the structures, thermodynamic stability and enzyme activated gene knockdown. These studies will lay the foundation of future research of enzyme-responsive nucleic acid prodrugs in model animals.