Novel flow cytometry-based assay for HIV-infected cells

描述(申请人提供)：艾滋病毒感染的抗逆转录病毒疗法(ART)是医学史上最重要和最迅速的进步之一。然而，虽然ART在限制病毒复制和减缓病程方面有效，但如果病毒不迅速反弹和疾病发展，就无法阻止ART。此外，虽然接受抗逆转录病毒治疗的患者可以预防艾滋病，但与普通人群相比，接受抗逆转录病毒治疗的艾滋病毒感染者发生心血管疾病和神经认知能力下降等重大非艾滋病事件的频率更高，年龄也更早。最后，疫情太大，推广的障碍太大。 太高了，无法有效地治疗每个受感染的人来结束疫情。因此，我们必须了解艾滋病毒如何在感染者体内持续存在的机制，以设计新的疗法来根除病毒或从功能上治愈患者。了解TIS的第一步是开发检测病毒持久性水平的方法。有几种分析方法，但在提供多少信息、执行它们的速度有多快、它们的成本有多高以及测量的可变性方面都有严重的限制。我们在这里提出了一种新的方法，利用流式细胞术在单细胞分辨率下检测HIV感染的细胞。这种方法一旦优化，将提供更多的信息，更快地进行，成本更低，与其他用于测量艾滋病毒宿主的分析方法相比，变异性更小。我们将优化这一检测方法，然后将其应用于了解不同HIV感染人群中HIV储存库的状态，以评估一些临床不同的受试者是否具有不同的HIV储存库的大小、活性或重新激活潜力，并开发一种高通量筛选方法来评估重新激活或杀死潜在储存库的新疗法。这一过程将证明这种分析的实用性，这是一种突出的情况，它可以在其中应用，并传播该分析，供其他人在他们认为合适的情况下使用。

DESCRIPTION (provided by applicant): Antiretroviral therapy (ART) for HIV infection is one of the most important and rapid advances in the history of medicine. However, while effective at limiting virus replication and slowing the course of disease, ART cannot be stopped without rapid rebound of virus and progression of disease. In addition, while AIDS is prevented in patients on ART, significant non-AIDS events such as cardiovascular disease and neurocognitive decline occur more frequently and at an earlier age in HIV-infected subjects on ART than in the general population. Finally, the epidemic is too large and the barriers to roll-out too high to effectively treat every infected individual to end the epidemic. It is therefore imperative that we understand the mechanisms of how HIV persists in the infected individual to design novel therapies to eradicate the virus or functionally cure the patient. The first step in tis understanding is the development of assays to measure the level of virus persistence. Several assays exist, but have serious limitations in terms of how much information is provided, how quickly they can be performed, how expensive they are, and how variable the measurement is. We propose here a novel method to detect HIV-infected cells at single cell resolution using flow cytometry. This method, once optimized, will provide more information, be performed more rapidly, cost less, and have less variability than other assays for the measurement of HIV reservoirs. We will optimize this assay and then apply it to understand the state of the HIV reservoir in various groups of HIV-infected subjects to evaluate whether some clinically different subjects may have different sizes, activities, or reactivation potentials of their HIV reservoirs ad also to develop a high-throughput screening assay for the evaluation of novel therapeutics to reactivate or kill the latent reservoir. This process will demonstrate the utility of this assay an highlight situations in which it can be applied as well as disseminate the assay for use by others to apply as they see fit.