Vaccine with altered HIV antigens

HIV 抗原发生改变的疫苗

• 批准号: 6591367
• 负责人: Danuta B Kozbor
• 金额: $ 23.37万
• 依托单位: ROSWELL PARK CANCER INSTITUTE CORP
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-06-01 至 2004-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6591367
• 关键词: AIDS vaccines; MHC class I antigen; antibody titering; cell line; cellular immunity; cytotoxic T lymphocyte; dendritic cells; endoplasmic reticulum; enzyme linked immunosorbent assay; gag protein; genetically modified animals; helper T lymphocyte; human immunodeficiency virus; humoral immunity; immunization; laboratory mouse; neuronal transport; secretory protein; vaccine development; vaccinia virus; vector vaccine; virus antigen; virus protein

DESCRIPTION: (Provided by Applicant) HIV Gag-Pol produces the Gag precursor protein and the Gag-Pol fusion protein in infected cells by frameshifting in a 20:1 ratio. This unequal production of Gag versus Pol proteins was suggested to be responsible for the low immunogenicity of Pol compared with that of Gag in numerous nonhuman primates and human studies using DNA vaccines or a live recombinant vector containing Gag-Pol constructs. In fact, recent experiments have shown that deletion of a frameshift site resulted in production of a Gag-Pol fusion protein, designated hGag-Pol FS Pr, with improved ability to induce Pol-specific immune responses. We prepared Gag-Pol vectors that encode immunogenic regions of Gag (p17 and p24) and Pol (p51) from a single continuous open reading frame as cytoplasmic (GP-92) and secretory (S-GP-92) fusion proteins. The GP-92 fusion protein contains 183 CTL epitopes with multiple HLA-binding motifs and 49 Th epitopes. We showed that immunization of the HLA-A2/Kb transgenic mice with the GP-92 construct, expressed in recombinant vaccinia viruses (rVV), induced increases in immune responses to Gag and Pol gene products compared with responses elicited by a Gag-Pol pseudoparticle. In this study, we propose to compare immunogenicity of the cellular and secretory form of Gag-Pol FS Pr and GP-92 fusion proteins expressed in the context of rVV and DNA vectors. We hypothesize that targeting the constructs to the endoplasmic reticulum (ER) will increase the level of specific CTL responses by augmenting the delivery of antigenic peptides to the MHC class I molecules through I) the retrograde transport of the secretory protein from the ER to the cytosol, and ii) the exogenous antigen uptake. Because antigen processing and presentation may augment immunogenicity of subdominant epitopes, it is possible that immunization with the modified fusion proteins can induce qualitative changes in the immune repertoire and improve vaccine efficacy. The Specific Aims are: I) Using HLA-A2/Kb transgenic mice, we will perform a head-to-head analysis of the level of immune responses induced by immunization with the cellular and secretory form of Gag-Pol FS Pr and GP-92 fusion proteins expressed in the context of rVV and DNA vectors. II) We will investigate differences in presentation of the cellular and secretory form of Gag-Pol FS Pr and GP-92 on MHC class I molecules. III) We will analyze protection levels achieved by DNA immunization with the modified Gag-Pol fusion constructs against intrarectal challenge with rVV expressing Gag and Pol proteins.

描述：（由申请人提供）HIV Gag-Pol 产生 Gag 前体 蛋白质和 Gag-Pol 融合蛋白在感染细胞中通过移码 20：1的比例。 Gag 与 Pol 蛋白的这种不平等的产生被认为是 与 Gag 相比，Pol 的免疫原性较低。 大量使用 DNA 疫苗或活体疫苗的非人类灵长类动物和人类研究 含有 Gag-Pol 构建体的重组载体。事实上，最近的实验 已经表明，移码位点的删除会导致产生 Gag-Pol 融合蛋白，命名为 hGag-Pol FS Pr，具有改进的能力 诱导 Pol 特异性免疫反应。 我们制备了编码 Gag 免疫原性区域（p17 和 p17）的 Gag-Pol 载体。 p24) 和 Pol (p51) 来自单个连续开放阅读框作为细胞质 (GP-92) 和分泌性 (S-GP-92) 融合蛋白。 GP-92融合蛋白 包含 183 个具有多个 HLA 结合基序的 CTL 表位和 49 个 Th 表位。 我们证明用 GP-92 免疫 HLA-A2/Kb 转基因小鼠 在重组痘苗病毒（rVV）中表达的构建体，诱导增加 与反应相比，对 Gag 和 Pol 基因产物的免疫反应 由 Gag-Pol 赝粒子引发。在这项研究中，我们建议比较 Gag-Pol FS Pr 和 GP-92 的细胞和分泌形式的免疫原性 在 rVV 和 DNA 载体中表达的融合蛋白。我们假设 靶向内质网 (ER) 的构建体将会增加 通过增强抗原的递送来提高特异性 CTL 反应的水平 通过 I) 逆行转运将肽连接至 MHC I 类分子 从 ER 到细胞质的分泌蛋白，以及 ii) 外源抗原 吸收。因为抗原加工和呈递可能会增强免疫原性 亚优势表位，有可能用修饰的免疫接种 融合蛋白可以引起免疫组库的质变 提高疫苗功效。 具体目标是： I) 使用 HLA-A2/Kb 转基因小鼠，我们将进行 免疫诱导的免疫反应水平的头对头分析 具有细胞和分泌形式的 Gag-Pol FS Pr 和 GP-92 融合蛋白 在 rVV 和 DNA 载体中表达。 II) 我们将调查 Gag-Pol FS Pr 的细胞和分泌形式存在差异 和 GP-92 作用于 MHC I 类分子。 III) 我们将分析保护级别 通过使用改良的 Gag-Pol 融合构建体进行 DNA 免疫来实现 对抗表达 Gag 和 Pol 蛋白的 rVV 的直肠内攻击。