Impact of HIV-1 Fitness on Disease Progression

描述(由申请人提供)：本建议书的基本目标为 BE将测量HIV-1感染者分离株的体外适合性 B分支和非B分支HIV-1，然后将这种适应性与 疾病进展的预测因子。一项初步研究显示，两者之间存在很强的相关性 在体外HIV-1适合度和病毒载量之间。为了扩展这项研究，多个 从大约80名艾滋病毒感染者中分离出的艾滋病毒-1已经被 从瑞典卡罗林斯卡研究所和 比利时安特卫普的热带医学。在具体的AIM1中，我们将各自进行竞争 用四种对照HIV-1毒株确定患者的相对体外试验 健身价值。然后将这些适应值与不同的临床数据进行比较 参数，如CD4细胞计数和病毒载量。这一目标的目标是 延续之前的一项初步研究，该研究表明艾滋病毒-1与 健康和疾病进展。最终，住院期间的发病率 进展可能在HIV-1分离株或亚型的分布中起作用 这场流行病。具体的目标2是比较内部和外部的差异 亚型间适合度。来自分支A、B、C和C的多个分离物的适合度 D通过外周血单核细胞的配对竞争。初步 数据显示，不同亚型的分离株之间的差异比 属于相同的亚型。因此，在具体的AIM 3中，我们将尽可能地调查 亚型适合度的差异可能会影响非B分支的疾病进展 感染。如目标1所示，HIV-1初级分离株的体外适合性来自 大约150名感染艾滋病毒的乌干达人和津巴布韦人(参加 NICHD资助的研究)将与疾病的各种预测因素进行比较 进步。因为病毒首先会从血液和阴道中获得 在初次感染期间的分泌物，我们也会进行亚型间的检测 朗格罕细胞的竞争实验。病毒传播健身 研究是一种费力的技术，可以通过实验来绕过 在特定的目标4中提出。使用一种分析来观察 在HIV-1复制周期双重病毒感染期间，我们现在已经初步 数据表明，HIV-1的进入可能控制了竞争。我们会 在特定的目标4中确认这一初步观察，然后使用一本小说 HIV-1NL4-3包膜基因假型克隆的克隆策略 直接从病人样本中提取。这些包膜伪型HIV-1之间的竞争 然后将克隆与两个传播的HIV-1之间的竞争进行比较 从相同的两个患者样本中分离出来。

DESCRIPTION (provided by applicant): The basic objective of this proposal will be to measure the ex vivo fitness of HIV-1 isolates from individuals infected with clade B and non-clade B HIV-1 and then compare this fitness to various predictors of disease progression. A pilot study revealed a strong correlation between ex vivo HIV-1 fitness and viral load. To extend this study, multiple HIV- 1 isolates from approximately 80 HIV-infected individuals have been obtained from the Karolinska Insititute in Sweden and at the Institute of Tropical Medicine in Antwerp, Belgium. In SPECIFIC AIM1, we will compete each patient isolate with four control HIV-1 strains to determine a relative ex vivo fitness value. These fitness values will then be compared to various clinical parameters such as CD4 cell counts and viral load. The objective of this aim is to extend a previous pilot study showing a strong correlation between HIV-1 fitness and disease progression. Ultimately the rate of intrapatient disease progression may play a role in distribution of HIV-1 isolates or subtypes in the epidemic. SPECIFIC AIM 2 is to compare differences in intra- and intersubtype fitness. The fitness of multiple isolates from clades A, B, C, and D via pair-wise competitions in peripheral blood mononuclear cells. Preliminary data suggests a greater difference between isolates of different subtypes than of the same subtype. Thus, in SPECIFIC AIM 3, we will investigate if possible differences in subtype fitness may affect disease progression in non-clade B infections. As in aim 1, the ex vivo fitness of primary HIV-1 isolates from approximately 150 HIV-infected Ugandans and Zimbabweans (participating in an NICHD-funded study) will be compared to various predictors of disease progression. Since virus will be first obtained from both blood and vaginal secretions during primary infection, we will also perform intersubtype competition experiments using Langerhan cells. Viral propagation for fitness studies is a laborious technique that may be circumvented with the experiments proposed in SPECIFIC AIM 4. Using an assay to look at multiple steps in the HIV-1 replication cycle during dual virus infections, we now have preliminary data to suggest that HIV-1 entry may be controlling the competition. We will confirm this preliminary observation in SPECIFIC AIM 4 and then use a novel cloning strategy to produce HIV- 1 NL4-3 clones pseudotyped with env genes directly from patient samples. Competitions between these env pseudotyped HIV-1 clones will then be compared to competitions between two propagated HIV-1 isolates from the same two patient samples.