FLOW CYTOMETRY STUDY OF T CELL RESPONSES TO HIV VACCINES

T 细胞对 HIV 疫苗反应的流式细胞术研究

• 批准号: 6080187
• 负责人: JOSEPH M MCCUNE
• 金额: $ 37.74万
• 依托单位: J. DAVID GLADSTONE INSTITUTES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6080187
• 关键词: AIDS vaccines; HIV infections; T lymphocyte; active immunization; cellular immunity; clinical research; drug screening /evaluation; flow cytometry; gag protein; human immunodeficiency virus 1; human subject; immunologic assay /test; leukocyte activation /transformation; technology /technique development; vaccine development

The evaluation of HIV-1 vaccines will be facilitated by the availability of better assays for HIV-1-specific human CD4+ and CD8+ T cell immunity. Such assays should be robust, reproducible, and amenable to high throughput analysis of trans-shipped clinical specimens. We have developed and optimized a flow cytometry-based assay to detect specific CD4+ and CD8+ human T cell responses to cytomegalovirus (CMV) in cohorts of HIV-1-infected patients. This "cytokine flow cytometry" (CFC) assay has shown that the presence of CD4+ T cell responses against CMV is correlated with the absence or resolution of CMV-associated end organ disease. More recently, results from this assay have been shown to correlate with results obtained using major histocompatibility complex (MHC) Class I tetramers bearing specific epitopes of CMV. Since the CFC assay appears to reliably and specifically detect such antigen-specific responses, we have also developed related assays capable of detecting specific human CD4+ and CD8+ T cell responses to other AIDS-related opportunistic infections, including those caused by Mycobacterium tuberculosis, the Mycobacterium avium complex, cryptococcus, and human papilloma virus. We now propose to devise a similar CFC assay for the detection and quantitation of CD4+ and CD8+ human T cell responses against epitopes of HIV-1. Preliminary experiments indicate that it is possible to detect such responses against a whole virus vaccine preparation and against defined epitopes of HIV-1 Gag. To develop, optimize, and standardize this CFC assay, and to correlate it with results obtained using MHC Class I/HIV-1 peptide tetramers (Specific Aim 1), we have formed a collaboration with Becton Dickinson Biosciences, a leading flow cytometry reagent and equipment manufacturer and distributor. To further evaluate this CFC assay, we have made plans to analyze HIV-1-specific immune responses in patients who have been exposed to but not infected by HIV-1, who are in varying stages of HIV-1 disease progression, or who are HIV-1-infected nonprogressors (Specific Aim 2). Finally, to determine whether the results from this assay may facilitate the design and development of HIV-1 vaccines, we have established collaborations with several research groups that are testing HIV-1 vaccines (including The Immune Response Corporation, Chiron, VaxGen, and the laboratory of Dr. David Weiner at University of Pennsylvania) and we have made plans to analyze HIV-1-specific T cell responses in both HIV- 1-seropositive and -seronegative vaccinees (Specific Aim 3).

更好的HIV-1特异性人CD 4+和CD 8 + T细胞免疫测定的可用性将促进HIV-1疫苗的评价。 此类测定应是稳健的、可再现的，并且适合于转运临床标本的高通量分析。 我们开发并优化了一种基于流式细胞术的检测方法，用于检测HIV-1感染患者队列中特异性CD 4+和CD 8+人类T细胞对巨细胞病毒（CMV）的反应。 这种“细胞因子流式细胞术”（CFC）测定表明，针对CMV的CD 4 + T细胞应答的存在与CMV相关终末器官疾病的不存在或消退相关。 最近，该测定的结果已显示与使用携带CMV特异性表位的主要组织相容性复合体（MHC）I类四聚体获得的结果相关。 由于CFC检测似乎可靠和特异性地检测这种抗原特异性反应，我们还开发了相关的检测方法，能够检测特异性的人CD 4+和CD 8 + T细胞对其他艾滋病相关机会性感染的反应，包括由结核分枝杆菌，鸟分枝杆菌复合体，隐球菌和人乳头瘤病毒引起的感染。我们现在提出设计一种类似的CFC检测方法，用于检测和定量针对HIV-1表位的CD 4+和CD 8+人类T细胞应答。 初步实验表明，有可能检测到针对全病毒疫苗制剂和针对HIV-1 Gag的确定表位的这种应答。 为了开发、优化和标准化这种CFC检测，并将其与使用MHC I类/HIV-1肽四聚体（特定目标1）获得的结果相关联，我们与Becton Dickinson Biosciences（一家领先的流式细胞术试剂和设备制造商和分销商）合作。 为了进一步评估这种CFC检测，我们计划分析暴露于但未感染HIV-1的患者的HIV-1特异性免疫应答，这些患者处于HIV-1疾病进展的不同阶段，或者是HIV-1感染的非进展者（具体目标2）。 最后，为了确定该试验的结果是否有助于HIV-1疫苗的设计和开发，我们与几个正在测试HIV-1疫苗的研究小组建立了合作关系。（包括The Immune Response Corporation，Chiron，VaxGen，和宾夕法尼亚大学的大卫韦纳博士的实验室），我们已经制定了分析HIV-1特异性T细胞反应的计划，血清阳性和血清阴性疫苗接种者（具体目标3）。