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 CMS 储存库和区室中隐藏的特定 HIV-1 基因型是否与 HIV 相关痴呆 (HAD) 的发病机制有关。目前的研究将评估HAART时代前后死于早期、中期和晚期HAD的患者以及对照者大脑中克隆的全长HIV-1基因组。新开发的分子分析工具将使我们能够确定特定的 HIV 进化途径是否发生在患有 HAD 的患者的 CMS 中。此外，我们将系统地解决关于 HAD 发病过程中特定“特征序列”是否进化的持续争论。将祖先 HAD 和非 HAD 序列以及包含 HAD 特异性特征模式（如果找到）的序列分子替换到 HIV 分子克隆中，将使我们能够在体外测试基因组对巨噬细胞功能的影响，并允许对大脑患病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 患者的原始/最早形式的 HIV gp120-V3 序列位于脑膜中。该序列显然是所评估的所有其他序列的起源，包括大脑的其他四个区域以及外周的序列，尤其是精囊和淋巴结。在大脑内，HIV 进化程度最高的形式位于皮质下白质，这是与 HAD 相关的经典组织学变化最密切相关的区域。初步结果部分验证了我们将在当前应用程序中采用的新颖的系统发育方法。具体目标包括： 1) 与对照者相比，定量 HIV DMA 水平并对死于 HIV 疾病的患者大脑解剖学上不同区域的全长 HIV 基因组进行测序。使用新型软件 HIVbase（通过 ACSR）从 20 多个尸检样本中评估多达 100,000 个 HIV 序列，HIVbase 是一个序列特定关系数据库，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 Specific AIM1 中测序的每个组织中病毒群体的系统动力学。这一具体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异性的“特征序列”。 3) 测试 CMS 内的 HIV 病毒序列迁移是否以程序化和疾病特定的方式发生。 4) 利用 HAD 特异性“特征序列”进行分子替代研究，以测试对体外巨噬细胞功能的影响，并尝试将这些细胞定位在 HAD 患者大脑中的体内位置。评估参与早期感染的祖先序列的致病潜力。 5) 如果在上述特定目标中鉴定出 HAD 特异性“特征序列”，则将采用定量分析方法来测试与对照相比，是否可以在 HAD 患者的血液中鉴定出这些序列。最终目标是确定特定的 HIV 序列是否与 HAD 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出这些序列。