iPSC Modeling of the Role of NKX2-1 in Human Lung Development and Disease

描述(申请人提供)：最近发现的诱导多能干细胞(IPSCs)为体外模拟人类疾病和更好地了解人类发育提供了前所未有的机会。IPSC体外模型系统的临床翻译是该项目的目标，其目的是了解人类发育的最早阶段，重点是已知的最早的肺命运标志物-关键转录调控因子Nkx2.1在发育和人类疾病中的作用。对于这些研究，我们将使用我们开发的重新编程技术，该技术使我们能够建立一个全面的IPSCs库，这些IPSCs来自患有肺部疾病的儿童，这些疾病是由基因突变(如Nkx2.1)引起的，这些基因从肺部谱系指定的最早时刻起就活跃。这些疾病特异性细胞系提供了自然发生的人类模型，研究人类低形态、突变或显性负基因表达的影响，为理解控制正常和异常早期肺发育的机制提供了令人兴奋的机会。最终，对肺发育生物学的详细了解应该会导致对肺部疾病患者的更好治疗，以本项目中要研究的儿童为例，即4个因Nkx2.1编码区突变而导致临床表型不同的人。我们将把我们的体外研究和模型与从这些儿童的肺组织建立的微阵列相关联，我们将检验我们的假设：体外IPSC系统可以用于建模和理解遗传性肺部疾病，例如因Nkx2.1活性降低或增强而导致肺-甲状腺-脑临床表型不同的患者。目的1建议定义人类肺上皮祖细胞分化的正常和疾病特异性遗传特征，并评估这些体外模型系统与在这些细胞来源的患者中观察到的临床表型的比较。目的2评估Nkx2.1结合或失活改变等导致这些患者疾病发病的分子机制，目的3测试改善Nkx2.1相关人类疾病表型的机械驱动潜在疗法。

DESCRIPTION (provided by applicant): The recent discovery of induced pluripotent stem cells (iPSCs) has provided unprecedented opportunities to model human disease in vitro and better understand human development. The clinical translation of the iPSC in vitro model system is the goal of this project, whose aim is to learn about the earliest stages of human development, focused on the role in development and human disease of the earliest known marker of lung fate, the key transcriptional regulator NKX2.1. For these studies we will utilize reprogramming technology we have developed that has allowed us to establish a comprehensive bank of iPSCs generated from children with lung diseases caused by mutations in genes (such as NKX2.1) that are active from the earliest moments of lung lineage specification. These disease-specific cell lines provide naturally occurring human models of the effects of hypomorphic, mutant, or dominant negative gene expression in human beings, providing exciting opportunities to understand mechanisms that control normal vs. aberrant early lung development. Ultimately a detailed understanding of lung developmental biology should result in better treatments for patients with lung disease, exemplified by the children to be studied in this project, namely 4 individuals with divergent clinical phenotypes resulting from mutations in the coding region of NKX2.1. We will correlate our in vitro studies and models with microarrays established from the lung tissue of these children and we will test our hypothesis: the in vitro iPSC system can be applied to model and understand genetic lung diseases, exemplified by patients with divergent lung-thyroid-brain clinical phenotypes arising from reduced or augmented activity of NKX2.1. Aim 1 proposes to define the normal vs. disease-specific genetic signatures of differentiating human lung epithelial progenitors and to assess how these in vitro model systems compare to the clinical phenotypes observed in the patients from whom these cells are derived. Aim 2 evaluates the molecular mechanism, such as binding or inactivating alterations of NKX2.1 responsible for disease pathogenesis in these patients, and Aim 3 tests mechanistically-driven potential therapies for ameliorating the NKX2.1-related human disease phenotypes.