肿瘤微环境响应自促释放型基因载体的构建及性能研究

Gene therapy is widely promising in treating serious diseases of human. However, high performance vectors are always in need due to the degradability of nucleic acids (NAs) in body. Stimuli-responsive gene vectors, which can accelerate the release of NAs in the unique tumor microenvironment to produce well therapeutic results, have been attracting much attention. In this proposal, heparin with high negative charge density is selected and modified with functional molecules. The modified heparin will be crosslinked into “unlockable” nanoparticles by dynamic covalent bonds. Then, polycations will be assembled to the heparin nanoparticles to produce gene vectors with core-shell structures. Crosslinked negatively charged heparin core can be breakdown under the stimulation of tumor microenvironment. The unlocked heparin would interact with the polycation shell and replace the condensed NA, which could further facilitate the NA release. Various biological tests will be performed to evaluate the properties of vectors, such as toxicity and therapeutic effects. Their structure-property relationship in response to various tumor microenvironment will also be studied systematically. This proposal focuses on the construction of high-performance gene vectors based on self-accelerating release of NA in response to the tumor microenvironment, which is of great significance for the development of new delivery systems.

基因治疗在攻克人类危重大疾病方面有着非常广阔的临床应用前景，但由于基因治疗药物“核酸”的降解性，基因治疗的实现离不开性能优异的递送载体。刺激响应性载体因其可以在独特的微环境下促进核酸释放，提高基因治疗效果得到了广泛关注。本项目以负电性肝素多糖为基础，通过不同功能小分子修饰后，利用动态共价键将其交联成纳米粒，进而将得到的纳米粒与阳离子聚合物组装得到核-壳结构的基因载体用于核酸递送。在肿瘤微环境中，交联的负电性肝素纳米粒核心可响应解锁，与壳层中的阳离子聚合物相互作用，实现对运载核酸的自促释放，并进一步评价递送载体的安全性、肿瘤治疗效果及响应不同肿瘤微环境的构效关系等。本项目通过构建和系统地研究系列可肿瘤微环境响应解锁的核酸自促释放型基因载体，以实现更好的治疗效果，为研发新型递送体系奠定基础。

基因、蛋白等生物活性分子在疾病治疗中的作用得到了广泛关注。而为了提高其治疗效率，合适的递送载体必不可少。近期研究表明，胺化的聚甲基丙烯酸缩水甘油酯（PGMA）可有效用于构建低毒高效的阳离子基因载体。申请人通过对载体进行拓扑结构、电荷分布、动态共价键交联等不同设计，基于开环反应构建了主客体组装型淀粉多糖基阳离子基因载体、可解锁的核壳结构肝素多糖基阳离子基因载体、低毒长循环阳离子载体，实现了载体微环境响应、电荷反转自促释放负载核酸、体内长循环等性能的调控。项目组对载体的理化性能、体外生物学性能、体内富集及滞留时间、疾病治疗效果等进行了系统性研究，系列聚合物载体在CRISPR/Cas9系统、MST1质粒、siMST1等不同功能核酸递送中表现出优异性能，证实了载体系统在多种疾病中的应用前景。本项目实施有望为安全高效的新型载体构建提供新思路。