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: 6895006
• 负责人: MICHAEL Shannon MCGRATH
• 金额: $ 33.88万
• 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/6895006
• 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储库和隔室中的特定HIV-1基因型是否与HIV相关性痴呆（HAD）的发病机制相关。目前的研究将评估克隆的全长HIV-1基因组，这些基因组来自HAART前后死于早期、中期和晚期HAD的患者以及对照组的大脑。新开发的分子分析工具将使我们能够确定在HAD患者的CMS中是否存在特定的HIV进化途径。此外，我们将系统地解决正在进行的争论是否有一个特定的“签名序列”在HAD发病机制的演变。将祖先HAD和非HAD序列以及含有HAD特异性特征模式的序列（如果发现）分子取代到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，并允许组织化学鉴定大脑病变区域内的修饰的巨噬细胞。在初步研究中，我们发现HAD患者中HIV gp 120-V3序列的原始/最早形式位于脑膜中。这个序列显然是所有其他序列的起源，包括大脑其他四个区域的序列，以及周围的序列，特别是精囊和淋巴结。在大脑中，HIV的最进化形式是在皮质下白色物质中，这是与HAD相关的经典组织学变化最强烈参与的区域。初步结果部分已经验证了我们将在当前应用中采用的新的系统发育方法。具体目标包括：1）定量HIV DMA水平，并从与对照相比死于HIV疾病的患者的脑的解剖学上不同的区域对全长HIV基因组进行测序。将使用一种新的软件HIVbase对20多个尸检标本（通过ACSR）中多达100，000个HIV序列进行评估，HIVbase是一种序列特异性关系数据库，可以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AIM 1中测序的每个组织中病毒群的传播动力学。这一特定的目标将测试病毒序列是否与疾病发病机制平行进化，并可能识别疾病特异性“特征序列”。3)测试CMS内的HIV病毒序列迁移是否以程序性和疾病特异性方式发生。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，并尝试在HAD患者的大脑中体内定位这些细胞。评估早期感染中涉及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。最终的目标是确定特定的HIV序列是否有助于HAD的发病机制，以及这些序列是否可以在HAD风险患者的血液中识别。