C 端结构域缺失增强了负载 cpa/cpb 的固体脂质纳米粒对 BALB/c 小鼠感染利什曼原虫的保护作用。

BACKGROUND: We have demonstrated that vaccination with pDNA encoding cysteine proteinase Type II (CPA) and Type I (CPB) with its unusual C-terminal extension (CTE) can partially protect BALB/c mice against cutaneous leishmanial infection. Unfortunately, this protection is insufficient to completely control infection without booster injection. Furthermore, in developing vaccines for leishmaniasis, it is necessary to consider a proper adjuvant and/or delivery system to promote an antigen specific immune response. Solid lipid nanoparticles have found their way in drug delivery system development against intracellular infections and cancer, but not Leishmania DNA vaccination. Therefore, undefined effect of cationic solid lipid nanoparticles (cSLN) as an adjuvant in enhancing the immune response toward leishmanial antigens led us to refocus our vaccine development projects. METHODOLOGY/PRINCIPAL FINDINGS: Three pDNAs encoding L. major cysteine proteinase type I and II (with or without CTE) were formulated by cSLN. BALB/c mice were immunized twice by 3-week interval, with cSLN-pcDNA-cpa/b, pcDNA-cpa/b, cSLN-pcDNA-cpa/b(-CTE), pcDNA-cpa/b(-CTE), cSLN, cSLN-pcDNA and PBS. Mice vaccinated with cSLN-pcDNA-cpa/b(-CTE) showed significantly higher levels of parasite inhibition related to protection with specific Th1 immune response development, compared to other groups. Parasite inhibition was determined by different techniques currently available in exploration vacciation efficacy, i.e., flowcytometry on footpad and lymph node, footpad caliper based measurements and imaging as well as lymph node microtitration assay. Among these techniques, lymph node flowcytometry was found to be the most rapid, sensitive and easily reproducible method for discrimination between the efficacy of vaccination strategies. CONCLUSIONS/SIGNIFICANCE: This report demonstrates cSLN's ability to boost immune response magnitude of cpa/cpb(-CTE) cocktail vaccination against leishmaniasis so that the average parasite inhibition percent could be increased significantly. Hence, cSLNs can be considered as suitable adjuvant and/or delivery systems for designing third generation cocktail vaccines.

背景：我们已经证明，用编码半胱氨酸蛋白酶 II（CPA）和 I 型（CPB）及其不寻常的 C 末端延伸（CTE）的 pDNA 进行疫苗接种可以部分保护 BALB/c 小鼠免受皮肤利什曼病感染。不幸的是，这种保护不足以在没有加强注射的情况下完全控制感染。此外，在开发利什曼病疫苗时，有必要考虑适当的佐剂和/或递药系统，以促进针对抗原的特异性免疫反应。固体脂质纳米粒已在针对细胞内感染和癌症的药物递药系统开发中找到了自己的道路，但在利什曼病 DNA 疫苗接种中却没有。因此，阳离子固体脂质纳米粒（cSLN）作为佐剂增强针对利什曼抗原的免疫反应的效果尚不清楚，这导致我们重新关注我们的疫苗开发项目。

方法/主要发现：用 cSLN 对编码 L. major 半胱氨酸蛋白酶 I 和 II（带或不带 CTE）的三种 pDNA 进行了配方。BALB/c 小鼠每隔 3 周用 cSLN-pcDNA-cpa/b、pcDNA-cpa/b、cSLN-pcDNA-cpa/b(-CTE)、pcDNA-cpa/b(-CTE)、cSLN、cSLN-pcDNA 和 PBS 进行两次免疫。与其他组相比，用 cSLN-pcDNA-cpa/b(-CTE) 疫苗接种的小鼠显示出更高水平的寄生虫抑制，与保护相关，并且产生了特定的 Th1 免疫反应。寄生虫抑制通过当前可用于探索疫苗接种疗效的不同技术来确定，即足部和淋巴结的流式细胞术、足部卡尺测量和成像以及淋巴结微量滴定测定。在这些技术中，发现淋巴结流式细胞术是区分疫苗接种策略疗效的最快、最敏感和最容易重现的方法。

结论/意义：本报告证明了 cSLN 能够增强 cpa/cpb(-CTE) 鸡尾酒疫苗接种对利什曼病的免疫反应幅度，从而可以显著提高平均寄生虫抑制百分比。因此，cSLNs 可以被认为是设计第三代鸡尾酒疫苗的合适佐剂和/或递药系统。