Dendritic Cell Targeting of Lassa Fever Vaccine

描述(由申请人提供)：我们的目标是使用新的生物有机化合物来中介递送来提高DNA疫苗的效力。在临床试验中，活疫苗一直比DNA疫苗更有效，但最近允许将DNA输送到特定细胞的技术为DNA疫苗提供了新的可能性。在这里，我们建议使用新的糖多胺作为递送载体来测试定向递送到树突状细胞(DC)的DNA疫苗。DC是主要的抗原提呈细胞，表达丰富的细胞表面碳水化合物受体，如甘露糖受体和DC-SIGN。我们的研究中使用的基于环糊精的糖多胺由两个功能区组成：与DNA结合并形成复合物的多个氨基，以及结合细胞表面凝集素并将DNA复合物靶向DC的糖配体。增强DC对DNA的摄取将增强DNA疫苗的免疫原性。细胞培养研究已经证明，甘露糖化多胺能显著增强DC对质粒DNA的摄取。在这里，我们建议测试体内靶向递送的效果。我们假设，针对树突状细胞表面碳水化合物受体的DNA疫苗将引起增强的细胞介导性反应。为了验证这一假设，我们首先将使用LCMV感染的小鼠模型系统，该模型系统已知会引发强烈的细胞介导的免疫反应。我们将有两个具体目标。在目标1中，我们将合成新型糖基化环糊精多胺(CDPA)，并检测报告基因在树突状细胞培养中的摄取和表达。将甘露糖和低聚甘露糖配体引入CDPA，分别靶向于甘露糖受体和树突状细胞表面的DC-sLGN。在目标2中，我们将测试口服不同剂型的DNA疫苗后细胞免疫的大小。编码LCMV gp基因的质粒DNA将以裸露DNA或与CDPA或糖基化CDPA络合的DNA形式口服给小鼠。一旦我们找到了最适合诱导细胞免疫的配方，我们将用与拉沙热保护性免疫相关的DNA编码抗原(Lassa NP和GP基因)取代LCMV-GP基因。我们将在小鼠系统中确定与我们的化合物络合的DNA是否仍能引发强大的细胞介导的免疫反应。一旦我们在小鼠模型系统中优化了疫苗配方，我们将把靶向递送系统应用于猴子模型和人类临床试验。这些研究将导致针对拉沙热的改进疫苗和对细胞特异性递送有用的新化合物。

DESCRIPTION (provided by applicant): Our goal is to enhance the efficacy of DNA vaccines using new bio-organic compounds to mediate delivery. Live vaccines have always been more effective than DNA vaccines in clinical trials, but recent technology allowing DNA delivery to specific cells provides new possibilities for DNA vaccines. Here we propose to test DNA vaccines for targeted delivery to dendritic cells (DC) using novel glyco-polyamines as the delivery vehicle. DC are the primary antigen-presenting cells and they express abundant cell surface carbohydrate receptors, such as mannose receptor and DC-SIGN. The cyclodextrin-based glyco-polyamines used in our studies consist of two functional domains: the multiple amino groups that bind and form complexes with DNA, and the sugar ligands that bind cell surface lectins and target the DNA complexes to DC. Enhancing the uptake of DNA by DC will enhance the immunogenicity of DNA vaccines. Cell culture studies have already demonstrated that mannosylated polyamines significantly enhance the uptake of plasmid DNA into DC. Here we propose to test the efficacy of targeted delivery in vivo. We hypothesize that DNA vaccines targeted to the cell surface carbohydrate receptors of dendritic cells will elicit enhanced cell-mediated responses. To test this hypothesis, we will initially employ the LCMV-infected murine model system that is known to elicit strong cell-mediated immune responses. We will have two specific aims. In aim 1, we will synthesize novel glycosylated cyclodextrin-based polyamines (CDPA) and test the uptake and expression of reporter genes in dendritic cell culture. Mannose and oligomannose ligands will be introduced into CDPA for targeting to mannose receptors and DC-SlGN on dendritic cells, respectively. In aim 2, we will test the magnitude of cell mediated immunity after oral inoculation with DNA vaccines in various formulations. Plasmid DNA encoding the LCMV GP gene will be orally delivered to mice as naked DNA or as DNA complexed with CDPA or glycosylated CDPA. Once we find a formulation that is optimum for eliciting cell-mediated immunity, we will replace the LCMV-GP gene with DNA encoding antigens associated with protective immunity in Lassa fever (Lassa NP and GP genes). We will determine, in the murine system, whether DNA complexed with our compound can still elicit strong cell-mediated immune responses. Once we have optimized the vaccine formulation in the murine model system, we will apply the targeted delivery system to monkey models and human clinic trials. These studies will lead to improved vaccines against Lassa fever and new compounds that will be useful for cell-specific delivery.