减毒鼠伤寒沙门氏菌介导的抗原递呈系统在肿瘤治疗性口服DNA疫苗及蛋白质疫苗制备中的应用

发布时间：2018-04-09 08:55

  本文选题：治疗性疫苗　切入点：减毒鼠伤寒沙门氏菌　出处：《复旦大学》2010年博士论文

【摘要】： 治疗性疫苗能够打破免疫耐受并引起针对癌症细胞的长时程免疫反应,由于疫苗给药途径与构建有效的抗原递呈系统是疫苗制备需要解决的关键性问题。减毒鼠伤寒沙门氏菌作为癌症疫苗的载体能够将DNA从原核细胞传递至真核细胞,并且能够选择性积聚在肿瘤组织。热休克蛋白70能够通过其多肽结合结构域与抗原结合并与抗原递呈细胞表面的Hsp70受体作用,将抗原传递至抗原递呈细胞。本论文的第一部分,我们构建了一种新型的低拷贝DNA疫苗,该疫苗基于Hsp70-TAA复合物构建,并由减毒鼠伤寒沙门氏菌SL3261菌株作为载体将DNA运送至小鼠体内。口服该重组细菌疫苗能够诱发针对黑色素肿瘤细胞的CTL细胞毒性细胞杀伤作用并显著激活T细胞反应。在肿瘤预防实验组C57BL/6J小鼠模型中,该治疗性口服疫苗能够诱发57.1%的肿瘤预防免疫反应,在肿瘤治疗组中,该疫苗能够完全消除62.5%的C57BL/6J小鼠中B16F10肿瘤的生长,显著延长小鼠的生存期。 口服减毒沙门氏菌疫苗载体通常定位在吞噬性细胞中,限制了其所携带的抗原蛋白进入抗原递呈途径,论文的第二部分我们利用沙门氏菌Ⅲ型分泌系统对蛋白质的转运功能来克服这种限制。Ⅲ型分泌蛋白能够将沙门氏菌编码的抗原蛋白直接运送至抗原递呈细胞胞质。抗原递呈细胞表面存在Hsp70的特异性受体,故Hsp70能够以Hsp70-TAA复合物的形式向APC细胞递呈抗原。本研究利用Ⅲ型分泌系统来实现对Hsp70-TRP2融合抗原蛋白的转运和递呈。口服该重组细菌疫苗能够实现对肿瘤特异性抗原的有效递呈,打破免疫耐受,诱导产生肿瘤细胞特异性的CTL反应和NK细胞杀伤作用,并在C57BL/6J小鼠上建立的B16F10黑色素肿瘤模型中获得了75%的肿瘤预防效果和62.5%的治疗效果。我们设计了以减毒鼠伤寒沙门氏菌作为肿瘤DNA疫苗及蛋白疫苗的传递工具的两套抗原递呈系统,为肿瘤的预防和治疗提供了有效的新方法。
[Abstract]:Therapeutic vaccine can break the immune tolerance and induce long-term immune response to cancer cells. Because of the vaccine delivery pathway and the construction of an effective antigen presentation system is the key problem to be solved in vaccine preparation.Attenuated Salmonella typhimurium can transfer DNA from prokaryotic cells to eukaryotic cells as a vector of cancer vaccine and can selectively accumulate in tumor tissues.Heat shock protein 70 (HSP70) can transfer antigens to antigen-presenting cells through its polypeptide binding domain which binds to antigens and acts with Hsp70 receptors on the surface of antigen-presenting cells.In the first part of this thesis, we constructed a novel low copy DNA vaccine, which was constructed based on Hsp70-TAA complex and was transported to mice by attenuated Salmonella typhimurium SL3261 strain as a vector.Oral administration of the recombinant bacterial vaccine could induce cytotoxicity of CTL cells against melanoma cells and activate T cell response significantly.In the C57BL/6J mice model of tumor prevention group, the therapeutic oral vaccine could induce 57.1% tumor preventive immune response. In the tumor treatment group, the vaccine could completely eliminate the growth of B16F10 tumor in 62.5% of C57BL/6J mice.The survival time of mice was significantly prolonged.Oral attenuated Salmonella vaccine vectors are usually located in phagocytic cells, limiting the entry of antigen proteins into antigen presentation.In the second part of this paper, we use the protein transport function of Salmonella type 鈪，

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