载SirT1 siRNA免疫纳米粒协同介导肝癌放疗増敏的作用机制研究

Radiation resistance is an important restriction factor in the treatment of liver cancer. Our previous studies have found that high levels of silent information regulator 1 (SirT1) expression is one of the key factors for the radiation resistance of hepatoma cells. Tumor associated macrophages (TAMs) are important infiltrating cells in the tumor. The activated M1 type TAMs not only facilitate their antitumor immunity, but also enhance their ability to capture immunonanoparticles. Therefore, we aim at constructing a SirT1-siRNA/CpG DNAsome immunonanoparticle which could efficiently load siRNA and specifically promote the polarization of TAMs into M1 type. Then, in vivo and in vivo experiments will be conducted to study the molecular mechanism of the active uptake of this immunonanoparticle by TAMs and their high-efficiency delivery of SirT1 siRNA to the neighboring hepatoma cells to inhibit the expression of SirT1 gene. Meanwhile, the internal mechanism that the immunonanoparticle specifically stimulate TAMs to express and secret tumor-killing factors will be explored. Finally, the mechanism of SirT1-siRNA/CpG DNAsome cooperatively mediated radiosensitization in hepatocellular carcinoma will be elucidated. This project will develop a new nanocarrier for targeted RNA interference as well as immune stimulation, and will provide new strategies and ideas for targeted radiotherapy of hepatocellular carcinoma.

辐射抵抗是制约肝癌放疗疗效的重要因素。我们前期研究发现，沉默信息调节因子1（SirT1）高表达是肝癌细胞具备辐射抗性的关键因素之一。肿瘤相关巨噬细胞（TAMs）是肿瘤内的重要浸润细胞，激活的M1型TAMs既能促进免疫杀伤肿瘤，又能主动捕获免疫纳米粒。因此，我们以TAMs为靶点，构建能高效荷载siRNA并特异性靶向刺激TAMs向M1型极化的SirT1-siRNA/CpG DNAsome免疫纳米粒，通过体内外实验研究TAMs主动摄取纳米粒并将SirT1 siRNA高效递送到邻近肝癌细胞以抑制SirT1基因表达的分子机制，同时探索纳米粒特异性刺激TAMs表达和分泌肿瘤杀伤因子的内在机制，并阐明在此双重作用下SirT1-siRNA/CpG DNAsome协同介导肝癌放疗増敏的作用机制。本项目的实施有望开发靶向性RNA干扰的新型免疫纳米系统，为肝癌的靶向放疗提供新的策略与思路。

辐射诱导巨噬细胞在辐射肿瘤中的聚集和浸润是导致肿瘤复发和转移的关键因素之一。通过持续稳定的免疫反应识别和杀死放疗后残留存活的癌细胞是预防肿瘤复发和远处转移的有效策略。在此，我们构建了一种基于独特的互补碱基配对规则，由CpG ODN负载的Y型双链DNA载体自组装的CpG DNAsome。CpG DNAsome的结构使其在单个纳米结构中集中了超过8125.5±822.5个CpG ODNs拷贝。CpG DNAsome能够显著抵抗核酸酶降解，并有效地将巨噬细胞复极化为M1型表型。此外，随着放射治疗诱导巨噬细胞的积累，更多的CpG DNAsome被内吞到肿瘤组织的巨噬细胞中，然后再极化巨噬细胞以引起高效的免疫刺激作用。体外和体内实验也进一步证实，CpG DNAsome成功抑制了单次放疗后肝癌的生长。