新型核心蛋白聚糖靶向性重组腺相关病毒的制备与鉴定

核心蛋白聚糖(decorin, DCN)是广泛存在于细胞基质中的一种富含亮氨酸的蛋白多糖, 属于蛋白聚糖家族中的小分子类. DCN可作为多种细胞因子的配体, 发挥多种生物学功能. DCN在一些肿瘤组织中高水平表达，调控恶性肿瘤的生长和迁移. 腺相关病毒(AAV)是肿瘤基因治疗中常用的基因工程载体, 利用重组技术可以实现对病毒衣壳蛋白的改造, 使其感染具有靶向性. 而针对DCN高表达细胞的转导可能成为肿瘤基因治疗应用中定向导入治疗基因的有效策略. 本研究在对多种DCN结合蛋白序列保守区的分析基础上, 筛选出具有较高活性的DCN结合功能域(DB1), 并将其融合至AAV衣壳蛋白VP2编码序列的N端; 继而利用AAV的嵌合包装技术, 成功制备了衣壳展示DB1表位的重组AAV. 在过表达DCN细胞的感染实验中, 该病毒表现出针对DCN较强的靶向性. 本研究所制备的DCN靶向性腺相关病毒不仅为肿瘤治疗的应用提供了一种新型载体, 同时可作为一类特殊的基因导入工具为研究DCN在肿瘤发生发展中的作用提供帮助.

Preparation and Characterization of Novel Adeno-associated Viral Vectors Targeting to Decorin-expressing Cells

.Decorin (DCN) is a core protein shared by a great variety of small molecular weight family members of proteoglycans. DCN is leucine rich in the amino acid composition, and is abundantly expressed in the extracellular matrix in numerous forms of polysaccharides. DCN serves as ligands of many types of cytokines and exerts important biological functions to influence cell growth, especially in cancers where high levels of DCN expression has been observed. Adeno associated virus (AAV) is a common vector being applied in cancer therapies. Targeted infection of DCN expressing tissues with AAV could be an attractive approach to selectively deliver therapeutic genes for cancer gene therapy. We have identified a DCN-binding motif (DB1) based on the consensus region among DCN-binding proteins (DBPs). By fusion of DB1 to the N-terminus of the VP2 capsid protein, we have successfully obtained the DB1-displaying recombinant AAV2 through a chimera pseudo packaging strategy. The obtained viral vector with DB1 motif in the capsid was able to infect DCN-overexpressing cells with much increased efficiency. Our novel DCN-targeting AAV not only can be a useful vector for the consideration in cancer gene therapy applications, but also can be a helpful gene delivery tool in studies to investigate the functions of DCN during cancer development.