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介导递送来增强DNA疫苗的功效。在临床试验中，活疫苗一直比DNA疫苗更有效，但最近的技术允许将DNA递送到特定细胞，为DNA疫苗提供了新的可能性。在这里，我们建议测试DNA疫苗的靶向交付树突状细胞（DC）使用新的糖多胺作为运载工具。DC是主要的抗原提呈细胞，表达丰富的细胞表面糖受体，如甘露糖受体和DC-SIGN。在我们的研究中使用的基于环糊精的糖多胺由两个功能域组成：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基化多胺显著增强质粒DNA进入DC的摄取。在这里，我们建议测试体内靶向递送的功效。我们假设靶向树突状细胞的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的反应。为了验证这一假设，我们将首先采用LCMV感染的小鼠模型系统，该系统已知可引发强烈的细胞介导的免疫应答。我们将有两个具体目标。在目标1中，我们将合成新型的基于糖基化环糊精的多胺（CDPA），并在树突状细胞培养中测试报告基因的摄取和表达。将甘露糖和寡甘露糖配体引入CDPA中，分别靶向树突细胞上的甘露糖受体和DC-SlGN。在目标2中，我们将测试口服接种各种制剂的DNA疫苗后细胞介导的免疫力的大小。编码LCMV GP基因的质粒DNA将作为裸DNA或作为与CDPA或糖基化CDPA复合的DNA经口递送至小鼠。一旦我们找到最适合引发细胞介导的免疫的制剂，我们将用编码与拉沙热中的保护性免疫相关的抗原（拉沙NP和GP基因）的DNA替换LCMV-GP基因。我们将确定，在小鼠系统中，与我们的化合物复合的DNA是否仍然可以引起强烈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制剂，我们将把靶向递送系统应用于猴模型和人体临床试验。这些研究将导致针对拉沙热的改进疫苗和用于细胞特异性递送的新化合物。

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.