基于纳米氧化石墨烯核酸递送系统的大肠癌靶向基因-病毒治疗及其机制研究

Targeting Gene-Viro-Therapy strategy for colorectal cancer has exhibited the excellent antitumor effect in preclinical studies. However, the immune response and side effect after intravenous injection hamper its clinical application. Development of novel nano-carrier loaded with viral genome DNA can effectively improve the antitumor effect and safety. Based on the systematic study of nano-graphene oxide carrier and oncolytic adenovirus, we firstly develop the nano-graphene oxide carrier CPP-NGO-PEG-PEI targeting colorectal cancer, which can carry adenovirus genome DNA with ST13-IETD-Endo dual functional gene. After the internalization of complex, the cancer cells act as "virus factory" to package and amplify a large number of virus particles, and are lysed. The viruses are released from the lytic cancer cells and infect surrounding cancer cells to achieve the targeting cascade anticancer effect. Next, the evaluation of in vivo behavior of biological and safety for nano-carrier and virus plasmid complex CPP-NGO-PEG-PEI-pAd5/F11-CEAp-ST13-IETD-Endostatin is performed. The dynamic process of nano-carrier virus plasmid complexes internalization, the release of DNA, virus amplification and tumor lysis is observed. Finally, the antitumor effects and mechanism of the complex on PDTT xenografts derived from different colorectal cancer individual are evaluated. This study will effectively solve the problems of low targeting gene virus therapy effect and poor safety through intravenous administration, which can provide a new strategy for the clinical therapy of colorectal cancer.

大肠癌靶向基因-病毒治疗在临床前研究中效果良好，但仍存在静脉注射的免疫反应和毒副效应。寻求新型纳米载体荷载溶瘤病毒基因组可有效提高静脉给药的治疗效果和安全性。本项目拟在前期对纳米氧化石墨烯和溶瘤腺病毒研究基础上，首次制备大肠癌靶向纳米氧化石墨烯载体CPP-NGO-PEG-PEI，通过其运载携带ST13-IETD-Endo双基因的溶瘤腺病毒基因组DNA ，以肿瘤细胞为“病毒工厂”包装扩增病毒，溶瘤释放后感染周边肿瘤细胞实现级联抗癌效果；评估纳米载体-病毒质粒复合物CPP-NGO-PEG-PEI-pAd5/F11-CEAp-ST13-IETD-Endostatin的生物学行为和安全性，观察其入胞、DNA释放、病毒扩增和溶瘤的动态过程；最后，利用大肠癌PDTT模型分析纳米复合物的治疗效果及机制。该项目的完成将解决经静脉全身给药时靶向基因-病毒治疗效果不佳和安全性难题，为大肠癌临床治疗提供新思路。

大肠癌靶向基因-病毒治疗在临床前研究中效果良好，但仍存在静脉注射的免疫反应和毒副效应。寻求新型纳米载体荷载溶瘤病毒基因组可有效提高静脉给药的治疗效果和安全性。本项目通过制备纳米氧化石墨烯运载携带ST13和Endostatin双基因的Ad5/F11溶瘤腺病毒质粒，获得纳米载体-病毒质粒复合物NGO-PEG-PEI-pAd5/F11-CEAp-ST13-IETD-Endo；通过对纳米氧化石墨烯材料NGO-PEG-PEI递送病毒质粒进入细胞及病毒质粒自组装形成病毒颗粒等过程的监测，掌握纳米载体-病毒质粒复合物靶向治疗大肠癌的关键信息，实现了以肿瘤细胞为“病毒工厂”包装扩增病毒，溶瘤释放后感染周边肿瘤细胞的级联抗癌效果；同时验证了纳米载体-病毒质粒复合物的生物安全性，明确纳米载体-病毒质粒复合物靶向治疗大肠癌的可行性；构建SW480裸鼠大肠癌动物肿瘤模型，体内评估纳米载体-病毒质粒复合物对肿瘤的趋向性及杀伤作用，实验结果表明纳米载体-病毒质粒复合物协同光热治疗大肠癌具有显著效果，并在此基础上对纳米载体-病毒质粒复合物协同治疗作用机制进行了初步探讨。此外，基于纳米石墨烯复合系统及溶瘤病毒，我们分别进行了纳米石墨烯-二氧化锰复合物改善肿瘤微环境、增强放疗敏感性及IL-12基因修饰的溶瘤痘苗病毒协同PD-1单抗治疗大肠癌的研究，获得了较好的实验结果。本项目的一系列研究结果为大肠癌的治疗提供了一种新的途径，为肿瘤的靶向双基因病毒治疗策略提供技术手段，为以NGO-PEG-PEI为载体的基因-病毒靶向治疗提供理论依据，并为肿瘤的联合治疗奠定基础。本项目执行期间已发表SCI论文14篇，培养研究生2名，并对本项目主持人在本领域的学术提升和相关交叉科学研究提供及宝贵的支持。