HIV Sequence Evolution in AIDS Dementia Pathogenesis

艾滋病痴呆发病机制中的HIV序列进化

• 批准号: 7017818
• 负责人: MICHAEL Shannon MCGRATH
• 金额: $ 29.44万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-02-14 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7017818
• 关键词: AIDS dementia complex; chemical substitution; clinical research; computer assisted sequence analysis; genetic markers; genotype; high throughput technology; human genetic material tag; human immunodeficiency virus 1; human subject; molecular cloning; neuropathology; nucleic acid sequence; psychoneuroimmunology; tissue /cell culture; virus DNA; virus genetics

DESCRIPTION (provided by applicant): The overall goal of this application is to determine if particular HIV-1 genotypes, harbored in CMS reservoirs and compartments, are linked to the pathogenesis of HIV-associated dementia (HAD). The current study will evaluate cloned full-length HIV-1 genomes from the brains of patients who died with early, intermediate, and late stage HAD in the pre and post HAART era as well as controls. Newly developed molecular analytical tools will allow us to determine if a specific HIV evolutionary pathway occurs in the CMS of patients that develop HAD. Additionally, we will systematically address the on-going dispute of whether or not a specific "signature sequence" evolves during HAD pathogenesis. Molecular substitution of both ancestral HAD and non-HAD sequences and sequences containing HAD-specific signature patterns (if found) into HIV molecular clones will allow us to test the effects that the genomes have on macrophage function in vitro and allow histochemical identification of modified macrophages within diseased areas of the brain. In preliminary studies we found that an original/earliest form of HIV gp120-V3 sequence in a patient with HAD was in the meninges. This sequence was clearly the origin of all of the other sequences evaluated, including those in four other areas of the brain, and in the periphery, notably the seminal vesicles and lymph nodes. Within the brain, the most evolved form of HIV was in the sub-cortical white matter, an area of most intense involvement with classical histological changes associated with HAD. The preliminary results section has validated the novel phylogenetic approach that we will employ in the current application. Specific aims include: 1) To quantitate HIV DMA levels and sequence full-length HIV genomes from anatomically distinct regions of the brains of patients who died with HIV disease as compared to controls. Up to 100,000 HIV sequences will be evaluated from more than 20 autopsy derived specimens (through the ACSR) using a novel software, HIVbase, a sequence specific relational database that allows large scale storage and rapid processing of DNA sequence data. 2) To compare the phylodynamics of viral populations in each tissue sequenced in Specific AIM1 for all patients. This specific aim will test whether viral sequences evolve in parallel with disease pathogenesis and potentially identify disease specific "signature sequences". 3) Test whether HIV viral sequence migration within the CMS occurs in a programmatic and disease specific manner. 4) Perform molecular substitution studies utilizing HAD specific "signature sequences" to test effects on in vitro macrophage function and attempt to localize those cells in vivo in brains from patients with HAD. Evaluate the pathogenic potential of ancestral sequences involved in early stage infection. 5) If HAD specific "signature sequences" are identified in the above specific aims, quantitative analytic approaches will be employed to test whether those sequences can be identified in the blood of patients with HAD as compared to controls. The ultimate goal will be to determine whether specific HIV sequences contribute to HAD pathogenesis and whether those sequences can be identified in the blood of patients at risk for HAD.

描述(由申请人提供)：本申请的总体目标是确定储存在CMS储藏室和隔间中的特定HIV-1基因类型是否与HIV相关性痴呆(HAD)的发病机制有关。目前的研究将评估从HAART前后死于早、中、晚期HAART时代的患者以及对照组的大脑中克隆的全长HIV-1基因组。新开发的分子分析工具将使我们能够确定在患有HAD的患者的CMS中是否存在特定的HIV进化途径。此外，我们将系统地解决在HAD发病过程中是否有特定的“签名序列”进化这一持续的争议。将祖先的HAD和非HAD的序列和包含HAD特异性签名模式的序列(如果发现)分子替换到HIV分子克隆中，将使我们能够测试基因组对体外巨噬细胞功能的影响，并允许对大脑病变区域内修饰的巨噬细胞进行组织化学鉴定。在初步研究中，我们发现HAD患者的HIV gp120-V3序列的原始/最早形式在脑膜中。这个序列显然是评估的所有其他序列的来源，包括大脑其他四个区域和外围的那些序列，特别是精囊和淋巴结节。在大脑内，HIV最进化的形式是在皮质下白质，这是与HAD相关的经典组织学变化最强烈的区域。初步结果部分验证了我们将在当前应用程序中采用的新的系统发育方法。具体目标包括：1)量化艾滋病毒DNA水平，并与对照相比，对死于艾滋病毒疾病的患者大脑中解剖上不同的区域的全长艾滋病毒基因组进行测序。将使用一种新的软件HIVbase从20多个尸检样本(通过ACSR)评估多达10万个艾滋病毒序列，HIVbase是一种特定于序列的关系数据库，允许大规模存储和快速处理DNA序列数据。2)比较所有患者在特定AIM1测序的每个组织中的病毒种群的系统动力学。这一特定目标将测试病毒序列是否与疾病发病机制平行进化，并有可能识别疾病特有的“特征序列”。3)测试CMS内的HIV病毒序列迁移是否以程序性和特定于疾病的方式发生。4)利用HAD特定的“签名序列”进行分子替代研究，以测试对体外巨噬细胞功能的影响，并尝试在体内定位HAD患者脑组织中的这些细胞。评估与早期感染有关的祖先序列的致病潜力。5)如果在上述特定目的中确定了HAD特定的“特征序列”，将使用定量分析方法来测试与对照相比，是否可以在HAD患者的血液中识别这些序列。最终目标将是确定特定的HIV序列是否与HAD的发病机制有关，以及是否可以在有HAD风险的患者的血液中识别这些序列。