构建纳米管阵列高效光穿孔系统可控递送mRNA疫苗进入树突状细胞的研究

CD8+T细胞是清除肿瘤的关键，其诱导依赖于抗原在树突状细胞（DC）中的高效递呈。将编码抗原的mRNA疫苗递送入DC实现抗原的内源性表达被认为是最有效的手段。然而，因其强大的天然防御机制，DC难以被转染，物理递送可能优于化学转染和病毒载体。申请人前期系统性比较研究了碳基光热材料在光穿孔中的性能，开发了优于传统电穿孔的氧化石墨烯（rGO）光穿孔系统，可实现反复穿孔和胞内递送。为进一步调控穿孔孔径，实现mRNA向穿孔部位同步定向迁移，避免游离rGO在临床中的潜在毒性，本项目拟构建rGO包覆纳米管阵列光穿孔递送系统。利用激光激发rGO包覆的氧化铝纳米管产生穿孔，同时利用纳米管实现mRNA的同步注射。探索激光和纳米管参数对光穿孔孔径的调节，实现对不同长度mRNA穿孔条件的精确匹配，并探讨光热效应对DC细胞抗原递呈功能的潜在促进作用，最终将DC回输体内，激发CD8+T细胞反应，促进癌症免疫治疗。

CD8+ T cell is the key cell that could eliminate the tumor, which could only be activated by the presentation from the dendritic cell (DC). It is considered as an efficient way to deliver the antigen encoded mRNA into the DC to get an endogenous expression. However, due to the natural defense system, DC is hard to be transfected. Physical delivery methods could be a better choice than the biochemical transfection methods and viral vectors. The applicant has systematically studied the carbon-based photothermal materials used for laser-induced photoporation and developed a rGO-photoporation system, which is much better than traditional electroporation. A repeated photoporation is achieved for intracellular delivery. To further regulate the size of the pore and to control the migration of mRNA into the pore simultaneously, as well well without endocytosis of rGO, we propose to develop a rGO coated nanostraw array assisted photoporation delivery system. By illuminating the rGO coated nanostraw to porate the cell membrane, the mRNA is injected into the cells spontaneously. We will investigate the parameters of laser and nanostraw to regulate the pore size to match with different sizes of mRNA. We will also explore if the photothermal effect could induce better presentation function of DCs. The loaded DCs will be injected back to the body to activate the CD8+ T cell and induce cancer immunotherapy.