新型脂质-多肽杂交纳米疫苗介导的多靶点可视化免疫治疗黑色素瘤研究

Tumor vaccine has great potential in the treatment of metastatic melanoma. Recent studies have shown that dendritic cells (DCs) and CD169+ macrophages（Mφ）are important antigen presenting cells (APCs).We have reported that synchronous in vivo targeted delivery of antigen to APCs and activation of APCs can significantly improve the T-cell activation efficacy. However, there is a lack of vaccine that can efficiently activate DC and CD169+ macrophages cells, targeting deliver antigens to these cells, and directly kill tumor cells. Previously, we have developed a lipid-melittin hybrid nanoparticle, which possess the ability to efficiently kill melanoma cells and in vitro stimulate DC maturation. Moreover, we reported that magnetic pull force can facilitate DC migration and resulted in enhanced antitumor efficacy. Taking advantage of the high expression of SR-B1 in melanoma cells, mDCs, and macrophages, the project aims to establish a nanovaccine with simultaneous loading of magnetic resonance and optical imaging contrast agents, melanoma antigen peptide as well as melittin. In combination with magnetic pull force, this study aims to achieve: 1) efficient targeting of melanoma cells and APCs in vivo; 2) targeting delivery of antigens to DCs and CD169+ macrophages; 3) effective activation of APCs；4) direct killing of melanoma cells. 5)Another advantage of this study is the ability to use the dual-modality imaging technique (near infrared and magnetic resonance imaging) to in vivo dynamically evaluate the anticancer efficiency of the developed nanovaccines. Taken together, the project establishes a new method of integration of multi APC-targeted immunotherapy, anti-tumor peptide, magnetic pull force, as well as dula-modality imaging, thus providing a novel strategy for treating metastatic melanoma.

肿瘤疫苗治疗转移性黑色素瘤极具潜力。研究证实树突细胞（DC）和CD169+巨噬细胞（Mφ）均为重要的抗原递呈细胞（APCs），我们前期采用同步活体靶向抗原递送和促APC激活可提高T细胞激活效率，但仍缺乏能同时作用DC和CD169+Mφ并直接杀灭肿瘤细胞的疫苗。我们发现脂质-蜂毒肽杂交材料能高效杀伤肿瘤细胞并刺激DC成熟，采用磁力牵引能提高DC归巢效率而增强抗癌功效，且肿瘤细胞、成熟DC和Mφ均高表达清道夫受体（SR-B1）。据此本项目拟进一步研制同步负载黑色素瘤抗原肽、蜂毒肽和光学/核磁探针的纳米疫苗，并结合磁力牵引以实现：①在体高效经SR-B1靶向肿瘤细胞；②抗原靶向递送至DC和CD169+ Mφ；③激活APC；④直接杀伤肿瘤；⑤使用近红外/磁共振多模成像活体动态评价疫苗功效。本项目创新性地结合了多靶点免疫治疗、抗癌多肽、磁力牵引和多模成像手段，可为高效治疗转移性黑色素瘤提供可视化策略。

基于恶性黑色素瘤免疫治疗的现状，肿瘤疫苗治疗转移性黑色素瘤极具潜力。本课题组一直致力于探索制备新型纳米疫苗，以期达到既能在体高效激活T细胞，又能直接杀伤黑色素瘤细胞的目的。在前期采用蜂毒肽（melittin）多肽和抗原多肽纳米材料可同步活体靶向抗原递送和促进APC激活而提高T细胞激活效率的基础上，本项目研制了自组装成水凝胶的肿瘤杀伤性多肽MR（Melittin-RADA-32），具有制备简单、生物相容性好、负载效率高的特点，并进一步采用MR负载化疗药物、肿瘤细胞裂解液、免疫激活剂等制备一系列新型纳米肿瘤疫苗。其中负载阿霉素的纳米水凝胶MRD可实现药物缓释、化疗联合免疫一体化直接杀伤肿瘤细胞；负载肿瘤细胞裂解液的新型纳米疫苗MCL具有极佳的刺激DC成熟及预防肿瘤发生的功效；负载α肽和MnO2的肿瘤细胞膜磁性纳米疫苗具有HDL类似物功能，可在体经高表达清道夫受体SR-B1途径高效靶向成熟APC，并可同时实现磁共振/活体成像动态评价疫苗功效。实验结果显示，这一系列新型纳米疫苗均可引流至淋巴结完成APC摄取、递呈、活化，诱导未成熟DC成熟，促进Mφ极化，产生大量活化的CD8+T细胞杀伤肿瘤细胞，有效改善肿瘤免疫微环境，在体内、外均可发挥良好的治疗和预防黑色素瘤的功效，并且初步阐明杂交纳米疫苗所携带的肿瘤杀伤性多肽MR和肿瘤相关抗原激活APC的机制，明确了APC细胞在杂交疫苗作用过程中的关键作用，不同类型的DC以及Mφ的极化状态对抗肿瘤免疫的影响。另外，我们还探索了临床黑色素瘤免疫治疗的疗效预测指标。本项目创新性地结合了免疫治疗、化疗治疗、自身抗原以及多模成像手段，为高效治疗转移性黑色素瘤提供了可视化新策略。