Wang Shijie, Huang Weiwei, Li Kui, Yao Yufeng, Yang Xu, Bai Hongmei, Sun Wenjia, Liu Cunbao, Ma Yanbing
Laboratory of Molecular Immunology, Institute of Medical Biology, Chinese Academy of Medical Sciences & Peking Union Medical College.
Yunnan Key Laboratory of Vaccine Research & Development on Severe Infectious Diseases.
Int J Nanomedicine. 2017 Sep 12;12:6813-6825. doi: 10.2147/IJN.S143264. eCollection 2017.
Currently, therapeutic tumor vaccines under development generally lack significant effects in human clinical trials. Exploring a powerful antigen delivery system is a potential approach to improve vaccine efficacy. We sought to explore engineered bacterial outer membrane vesicles (OMVs) as a new vaccine carrier for efficiently delivering tumor antigens and provoking robust antitumor immune responses.
First, the tumoral antigen human papillomavirus type 16 early protein E7 (HPV16E7) was presented on -derived OMVs by genetic engineering methods, acquiring the recombinant OMV vaccine. Second, the ability of recombinant OMVs delivering their components and the model antigen green fluorescent protein to antigen-presenting cells was investigated in the macrophage Raw264.7 cells and in bone marrow-derived dendritic cells in vitro. Third, it was evaluated in TC-1 graft tumor model in mice that the recombinant OMVs displaying HPV16E7 stimulated specific cellular immune response and intervened the growth of established tumor.
DH5α-derived OMVs could be taken up rapidly by dendritic cells, for which vesicle structure has been proven to be important. OMVs significantly stimulated the expression of dendritic cellmaturation markers CD80, CD86, CD83 and CD40. The HPV16E7 was successfully embedded in engineered OMVs through gene recombinant techniques. Subcutaneous immunization with the engineered OMVs induced E7 antigen-specific cellular immune responses, as shown by the increased numbers of interferon-gamma-expressing splenocytes by enzyme-linked immunospot assay and interferon-gamma-expressing CD4 and CD8 cells by flow cytometry analyses. Furthermore, the growth of grafted TC-1 tumors in mice was significantly suppressed by therapeutic vaccination. The recombinant E7 proteins presented by OMVs were more potent than those mixed with wild-type OMVs or administered alone for inducing specific cellular immunity and suppressing tumor growth.
The results indicated that the nano-grade OMVs might be a useful vaccine platform for antigen delivery in cancer immunotherapy.
目前，正在研发的治疗性肿瘤疫苗在人体临床试验中普遍缺乏显著效果。探索一种强大的抗原递送系统是提高疫苗疗效的潜在途径。我们试图探索工程化细菌外膜囊泡（OMV）作为一种新型疫苗载体，以有效递送肿瘤抗原并激发强大的抗肿瘤免疫反应。
首先，通过基因工程方法将肿瘤抗原人乳头瘤病毒16型早期蛋白E7（HPV16E7）呈递在衍生的OMV上，获得重组OMV疫苗。其次，在巨噬细胞Raw264.7细胞和体外骨髓来源的树突状细胞中研究重组OMV将其成分和模型抗原绿色荧光蛋白递送至抗原呈递细胞的能力。第三，在小鼠的TC-1移植瘤模型中评估展示HPV16E7的重组OMV刺激特异性细胞免疫反应并干预已建立肿瘤生长的情况。
DH5α衍生的OMV可被树突状细胞迅速摄取，已证明囊泡结构对此很重要。OMV显著刺激树突状细胞成熟标志物CD80、CD86、CD83和CD40的表达。通过基因重组技术，HPV16E7成功嵌入工程化OMV中。用工程化OMV进行皮下免疫诱导了E7抗原特异性细胞免疫反应，酶联免疫斑点试验显示表达干扰素-γ的脾细胞数量增加，流式细胞术分析显示表达干扰素-γ的CD4和CD8细胞数量增加。此外，治疗性疫苗接种显著抑制了小鼠体内移植的TC-1肿瘤的生长。OMV呈递的重组E7蛋白在诱导特异性细胞免疫和抑制肿瘤生长方面比与野生型OMV混合或单独给药的重组E7蛋白更有效。
结果表明，纳米级OMV可能是癌症免疫治疗中用于抗原递送的有用疫苗平台。
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