Activity of novel drug candidates in primary cell models of HIV latency

潜伏感染的静止CD 4 T细胞是根除HIV-1的主要障碍。主要的障碍是外周血中潜伏感染细胞的频率低，以及缺乏已知的表型标记，可以将它们与未感染的细胞区分开来。这些障碍促进了潜伏期体外细胞模型的发展。这种模型允许操纵细胞和病毒特征，以获得对潜伏期如何建立和调节的机械理解。然而，没有一个HIV潜伏期的实验系统被认为是完全概括了体内潜伏病毒库的生物学。这主要是因为（a）HIV建立潜伏期的机制是多种多样的;（B）潜伏感染的细胞类型也是多种多样的。关于特定刺激（例如活化/分化、稳态增殖、细胞因子）和病毒因子（例如整合位点、Tat驱动的表达）如何影响实验系统中潜伏储库的动态，已经获得了丰富的知识。项目2.2将对新的小分子化合物和化合物组合进行“二级”表征，这些化合物已通过项目2.1中Hazuda博士小组的初步筛选被确定为诱导HIV转录。在目标1和2中，我们提出使用一组充分表征的原代细胞系统和一个基于Jurkat的细胞系克隆来验证候选药物化合物。第三个目标将是开发额外的原代细胞模型，以研究其他相关细胞类型中的HIV潜伏期，这些细胞类型以前未在体外探索过（例如，过渡记忆T细胞和巨噬细胞）。在至少一种原代培养系统中具有活性且未显示明显毒性或诱导细胞活化/增殖的处理将在Delaney Collaborative项目中转移至后续表征和开发阶段。这将包括详细的机制研究（目标1），在人源化小鼠和非人灵长类动物（目标3）以及离体患者细胞（目标4）中检测活性。我们的最终目标是确定新的药物和药物组合，当应用于患者的治疗策略时，将有很高的成功概率激活HIV潜伏病毒。

Latently infected, quiescent CD4 T cells represent a central obstacle to eradication of HIV-1. Major hurdles are the low frequencies of latently infected cells in peripheral blood and the lack of known phenotypic markers that can distinguish them from uninfected ones. These impediments have prompted the development of in vitro cell models of latency. Such models allow manipulation of cellular and viral characteristics to gain a mechanistic understanding of how latency is established and regulated. However, no single experimental system of HIV latency is perceived to completely recapitulate the biology of the latent viral reservoir in vivo. This is mainly because (a) the mechanisms for establishment of latency by HIV are multiple; and (b) the types of cells harboring latent infection are also multiple. A wealth of knowledge has been gained regarding how specific stimuli (e.g. activation/differentiation, homeostatic proliferation, cytokines) and viral factors (e.g. integration site, Tat-driven expression) influence the dynamics of latent reservoirs in experimental systems. Project 2.2 will perform "secondary level" characterization of novel small molecule compounds and combinations of compounds that have been identified to induce HIV transcription through primary screening by Dr. Hazuda's group in Project 2.1. In Aims 1 and 2, we propose to use a panel of well-characterized primary cell systems and one Jurkat-based cell line clone to validate candidate drug compounds. A third Aim will be the development of additional primary cell models to enable the study of HIV latency in other relevant cell types, not previously explored ex vivo (e.g. transitional memory T cells and macrophages). Treatments that are active in at least one of the primary culture systems, without showing overt toxicity or induction of cellular activation/proliferation, will be moved to subsequent phases of characterization and development within the Delaney Collaborative program. This will include detailed mechanistic studies (Objective 1), testing for activity in humanized mice and non-human primates (Objective 3) and in patient cells ex vivo (Objective 4). Our ultimate goal is to identify novel drugs and drug combinations that will have a high probability for success in activating HIV latent viruses when applied to treatment strategies in patients.