Human Studies of Attenuated Salmonella Vectors

说明(申请人提供)：接种疫苗是一种有效和经济的预防疾病的方法。人们希望口服疫苗有最广泛的应用，并诱导粘膜表面的免疫。活的、口服的减毒细菌载体在动物模型中非常有效。本项目的目的是通过在人类志愿者身上研究表达HIV-1抗原的鼠伤寒沙门氏菌载体，获得重要的安全性和免疫原性数据。该提案描述了一个综合实验室和临床项目。该申请建议完成正在进行的表达HIV-1 Gag的鼠伤寒沙门氏菌载体(CKS257)的第一阶段剂量递增研究，并描述了旨在增强人类对HIV-1 Gag的免疫反应的后续研究。这些实验包括多次剂量，经皮增强，以及研究“粘膜主要/全身增强”策略，如果可能的话。基于前期实验室和动物实验，我们选择了Phop/PhoQ/AroA缺失的细菌载体和表达系统，表明该菌株是安全的，并能对载体抗原产生细胞免疫应答。Phop/PhoQ和aroA基因对哺乳动物体内沙门氏菌的持久性/毒力是重要的；这些基因的缺失会导致沙门氏菌的主要衰减。一个合理设计的HIV-1 Gag抗原从一个平衡致死的质粒中表达，并通过III型分泌系统(TTSS)从沙门氏菌载体中分泌出来。在小鼠中，通过TTSS分泌的抗原比载体相关抗原产生更好的免疫反应和更好的疫苗效力。仔细评估临床安全性和疫苗脱落，并测量对载体和HIV-Gag的体液、粘膜和细胞免疫反应。此外，还将在实验室中产生新的结构，用于临床测试一种有前景的方法，该方法在动物模型中增强沙门氏菌载体抗原的免疫原性，但从未在人类身上进行测试：从Phop激活的启动子驱动抗原。这将需要使用一种不同于特征良好的Phop/PhoQ零菌株的媒介衰减策略，因此需要大量的临床前工作。该实验室在活细菌载体的翻译研究方面拥有丰富的经验。该项目的主要目标是提供实用的人体安全性和免疫原性数据，这些数据将有助于有效地指导活细菌载体的开发。

DESCRIPTION (provided by applicant): Vaccination is an effective and economical method of preventing disease. It is hoped that orally administered vaccines will have the widest application and induce immunity at mucosal surfaces. Live, orally administered attenuated bacterial vectors have been very effective in animal models. The purpose of this project is to gain important safety and immunogenicity data on Salmonella typhimurium vectors expressing a clinically relevant HIV-1 antigen by studying this prototype vaccine organism in human volunteers. The proposal describes an integrated laboratory and clinical project. The application proposes completion of an ongoing Phase I dose escalation study of a Salmonella typhimurium vector expressing HIV-1 Gag (CKS257), and describes follow-up studies designed to enhance immune responses to HIV-1 Gag in humans. These experiments include multi-dosing, transcutaneous boosting, and study of a "mucosal prime/systemic boost" strategy, if possible. The phoP/phoQ/aroA-deleted bacterial vector and expression system were chosen based upon pre-clinical laboratory and animal studies which suggest that this strain will be safe and engender cellular immune responses to the vectored antigen. The phoP/phoQ and aroA genes are important for Salmonella persistence/virulence within mammals; deletion of these genes results in major attenuation. A rationally-engineeered HIV-1 Gag antigen is expressed from a balanced-lethal plasmid and secreted from the Salmonella vector via the Type III Secretion System(TTSS). In mice, antigens secreted via the TTSS results in superior immune responses and greater vaccine efficacy than vector associated antigens. Clinical safety and vaccine shedding is carefully evaluated and humoral, mucosal and cellular immune responses to the vector and HIV-Gag are measured. In addition, new constructs will be generated in the laboratory to clinically test a promising approach which enhances the immunogenicity of Salmonella-vectored antigens in animal models, but has never been tested in humans: driving antigens from PhoP-activated promoters. This will necessitate using a vector attenuation strategy different from the well-characterized PhoP/PhoQ null strains, and therefore require significant preclinical work. The laboratory has significant experience in translational investigation of live bacterial vectors. The major goal of the project is to provide practical safety and immunogenicity data in humans which will help efficiently direct development of live bacterial vectors.