Development of a DNA Vaccine against Respiratory Syncytial Virus

拟议的项目目标是开发和研究一种针对呼吸道合胞病毒(RSV)的DNA疫苗。RSV感染了几乎所有两岁以下的儿童，目前还没有疫苗可用。本项目旨在设计一种含有RSV F基因免疫重要区域和修饰霍乱毒素基因的重组DNA载体。DNA免疫原肌肉注射可诱导局部和全身免疫反应。由于DNA质粒诱导Th1细胞反应，这种方法特别适合对抗RSV，在RSV中Th1免疫反应在诱导保护性免疫中非常关键。该项目吸引人的方面包括DNA质粒设计和开发的简单性，以及诱导细胞、体液和粘膜免疫反应的潜力。DNA质粒将使用DNA载体(来自基因治疗系统的phCMV1)构建，其中将克隆RSV F区和修饰的霍乱毒素基因。PCMV1DNA载体含有CMV启动子、内含子A和强终止子。DNA将被提纯并在小鼠体内传递 肌肉内。此外，为了测试Th1细胞应答的建立，将DNA疫苗和重组RSV F蛋白联合免疫小鼠。将分析免疫后产生的体液和细胞免疫反应。免疫原保护小鼠免受RSV感染的能力将通过用活的RSV攻击小鼠来评估。上呼吸道和下呼吸道感染将通过研究上粘膜和肺部的病毒载量进行评估。这项研究将有助于了解DNA疫苗接种和粘膜免疫反应，并有助于设计和开发更安全、更有效的疫苗来对抗RSV和其他感染黏膜的病毒。

The Proposed project goal is to develop and study a DNA vaccine against Respiratory Syncytial Virus (RSV). RSV infects almost all children under two years of age and there is no vaccine available. This project aims to engineer a recombinant DNA vector containing immunologically important regions of the RSV F gene and modified cholera toxin gene. It is expected that the DNA immunogen will induce local and systemic immune response when administered intramuscularly. Since DNA plasmids induce a Th1 cell response, this approach is particularly well suited against RSV where Th1 immune response is very critical in inducing protective immunity. Attractive aspects of the project include the simplicity of the design and development of the DNA plasmid, potential to induce cellular, humoral and the mucosal immune responses. The DNA plasmid will be constructed using a DNA vector (phCMV1 from Gene Therapy Systems) into which regions of RSV F and modified cholera toxin genes will be cloned. The phCMV1 DNA vector contains a CMV promoter, intron A and a strong terminator. DNA will be purified and delivered in mice intramuscularly. Additionally, to test the establishment of Th1 cell response, mice will be immunized with combination of DNA vaccine and recombinant RSV F protein. The humoral and cellular immune responses resulting from immunizations will be analyzed. The ability of the immunogen to protect mice from RSV infection will be evaluated by challenging mice with live RSV. The upper and lower respiratory tract infections will be assessed by studying virus load in upper mucosa and the lungs. This study will lead to an understanding of the DNA vaccination and mucosal immune response and contribute to design and development of a safer and more effective vaccine against RSV and other viruses infecting the mucosa.