Using iPSC variation to define HIV-1 regulatory networks

致病性逆转录病毒人类免疫缺陷病毒1型（HIV-1）引起了全球范围内获得性免疫缺陷综合征（艾滋病）的大流行。据估计，目前约有3700万人感染艾滋病毒-1，每年约有180万新感染病例，约有100万人死于艾滋病（2017年数字，来源：http://www.unaids.org/en/资源/情况介绍）。艾滋病目前被认为是一种可治疗的疾病，因为针对必需病毒蛋白的有效抗病毒药物已经开发出来。至关重要的是，没有预防新感染的疫苗，而且，一旦一个人被感染，就无法治愈（导致病毒根除的治疗）。此外，随着时间的推移，耐药菌株正在成为临床日益重要的问题。因此，有必要开发新的抗hiv -1治疗方法。尽管经过多年的深入研究，对细胞编码因子和调控HIV-1复制途径的复杂性（包括正性和负性）的理解仍然明显不完整。新的病毒调节因子的识别和定义将为未来的治疗干预提供新的目标，这除了有助于控制病毒生长和HIV-1相关疾病外，还可能影响寻求HIV-1治愈和/或疫苗接种的研究规划。Maraviroc是一种结合HIV-1进入受体CCR5阻断感染的小分子抑制剂，它证明了靶向细胞蛋白抑制HIV-1的可行性。在这个项目中，我们将开发和利用一种创新的跨学科策略来发现调节HIV-1复制的新的人类细胞因子和途径。我们将使用深度表征的诱导多能干细胞（iPSCs） HipSci文库（www.hipsci.org），其中已经存在详细的基因组，转录组学和蛋白质组学信息，并将HIV-1感染表型的变化与每个iPSC系固有的关键分子特征相关联。为了补充这种方法，我们还将对HIV-1感染高风险患者和已知疾病结局的患者的血液样本（多中心艾滋病队列研究- MACS； http://aidscohortstudy.org/）重新编程为iPSCs。通过使分子特征和感染数据与临床结果相关联，这些品系的协调特征将为我们的研究增加进一步的维度。对这些正交数据集进行深入的生物信息学研究将提示候选细胞编码的HIV-1调节因子，然后使用已建立的HIV-1感染实验室模型测试其功能。总之，我们的实验管道包括：(i)从两个来源筛选iPSC细胞系以发现极端的HIV-1感染表型；（ii）使用生物信息学工具和全细胞分子特征来发现新的调节因子；（iii）在细胞培养系统中验证我们的发现，并在分子水平上表征新型HIV-1调节因子的作用机制。我们的目标是发现新的人类蛋白质和控制HIV-1复制的途径，并为旨在设计克服HIV-1感染的新治疗策略的举措提供信息。最后，重要的是，我们致力于分享我们将从HIV-1暴露个体中获得的iPSC系：这些可以作为研究HIV-1生物学的独特资源，因为iPSC实际上是取之不尽的，并且具有分化为任何人类细胞类型的潜力，包括HIV-1的自然细胞靶标。

The pathogenic retrovirus human immunodeficiency virus type 1 (HIV-1) has caused the worldwide acquired immunodeficiency syndrome (AIDS) pandemic. It is estimated that ~37 million people currently live with HIV-1, with ~1.8 million new cases of infection and ~1 million AIDS deaths occurring every year (2017 figures, source: http://www.unaids.org/en/ resources/fact-sheet). AIDS is currently considered a treatable disease as a battery of effective anti-viral drugs that target essential viral proteins have been developed. Critically, there is no vaccine to prevent new infections and, once a person is infected, there is no cure (treatment leading to viral eradication). Moreover, with the passage of time, drug-resistant strains are becoming an increasingly significant problem in the clinic. It is therefore necessary to develop new anti-HIV-1 therapeutic approaches. Despite many years of intense investigation, the understanding of the complexity of cell-encoded factors and pathways that regulate HIV-1 replication (both positively and negatively) remains significantly incomplete. The identification and definition of new viral regulatory factors will suggest novel targets for future therapeutic intervention which, in addition to helping control viral growth and HIV-1-associated disease, may also impact research programmes seeking HIV-1 cure and/or vaccination. The feasibility of therapeutically targeting a cellular protein to inhibit HIV-1 is demonstrated by Maraviroc, a small molecule inhibitor that binds the HIV-1 entry receptor CCR5 to block infection. In this programme, we will develop and exploit an innovative inter-disciplinary strategy to discover novel human cell factors and pathways that regulate HIV-1 replication. We will use the deeply characterised HipSci library (www.hipsci.org) of induced pluripotent stem cells (iPSCs), for which detailed genomic, transcriptomic and proteomic information already exists, and correlate variations in HIV-1 infection phenotypes with key molecular signatures that are intrinsic to each iPSC line. To complement this approach, we will also reprogramme blood samples from patients at high risk of HIV-1 infection, and with known disease outcomes (the Multicenter AIDS Cohort Study - MACS; http://aidscohortstudy.org/), into iPSCs. Co-ordinated characterisation of these lines will add a further dimension to our study by also enabling molecular signatures and infection data to be correlated with clinical outcomes. In depth bioinformatic interrogation of these orthogonal datasets will suggest candidate cell-encoded HIV-1 regulatory factors, which will then be tested for function using established laboratory models of HIV-1 infection.In sum, our experimental pipeline comprises: (i) screening iPSC cell lines from two sources to find extreme HIV-1 infection phenotypes, (ii) using bioinformatic tools and whole cell molecular signatures to discover new regulatory factors and (iii) validating our findings in cell culture systems and characterising the mechanisms of action of novel HIV-1 regulators at the molecular level. Our goal is to uncover novel human proteins and pathways controlling HIV-1 replication, and to inform initiatives aiming to devise fresh therapeutic strategies for overcoming HIV-1 infection. Finally, and importantly, we are committed to sharing the iPSC lines that we will derive from HIV-1 exposed individuals: these can serve as a unique resource to study HIV-1 biology since iPSCs are virtually inexhaustible and have the potential to be differentiated into any human cell type, including the natural cellular targets of HIV-1.