IDENTIFICATION AND PRECLINICAL EVALUATION OF NOVEL THERAPEUTIC APPROACHES TO DYSKERATOSIS CONGENITA

项目摘要/摘要 先天性角化不良(DC)是一种罕见的疾病，它是由于在维持 端粒，这是保护染色体末端的结构。早产端粒缩短 导致广泛的器官病理，包括骨髓衰竭、肺(肺)纤维性瘢痕 肝纤维化)和肝(肝硬变)以及胃肠疾病。骨髓移植可以 成功地解决了骨髓衰竭，但其他病理没有得到有效治疗。在以下资助的研究中 美国国立卫生研究院R21拨款，主要研究人员最近证明，肠道病理涉及丢失 干细胞功能的正常支持是通过一种称为Wnt的细胞间通讯途径实现的，而药物 恢复Wnt通路信号(包括锂)可以改善这种病理。在此，我们建议 研究肺和肝脏的病理，特别是确定Wnt激活剂对这些组织是否有益 正如在肠道中观察到的那样。我们还将广泛调查其他受影响的途径，可能会提供 治疗干预要点。我们将使用人类诱导多能干细胞(IPSCs)和基因组 编辑以产生可在培养中研究的关键肺和肝细胞，以及端粒酶缺陷 小鼠模型，以研究这些机制并测试新的治疗方法，重点是使用 现有的FDA批准的药物。具体目标是： 1.鉴定培养的人DC-IPSC来源的II型肺泡上皮干细胞的缺陷，并检测 旨在逆转改变的通路以改善这些缺陷的药物操纵的能力。 2.鉴定培养的人DC-IPSC来源的肝细胞和星状细胞的缺陷，并检测 旨在逆转改变的通路以改善这些缺陷的药物操纵的能力。 3.使用小鼠模型研究药物操作拯救人类DC突变体的有效性 在活体背景下的IPSC来源的组织。这将包括最低血清锂的测定。 将挽救移植到老鼠体内的人体组织的浓度，以及锂和其他小分子的测试 拯救由端粒功能障碍驱动的肺纤维化和肝硬变的小鼠模型的分子。

Project Summary/Abstract The rare disease dyskeratosis congenita (DC) is caused by genetic deficiencies in the maintenance of telomeres, which are the structures that protect the ends of chromosomes. Premature telomere shortening leads to widespread organ pathology, including bone marrow failure, fibrotic scarring of the lungs (pulmonary fibrosis) and liver (hepatic cirrhosis), and gastrointestinal disorders. Bone marrow transplantation can successfully address marrow failure, but other pathologies are not treated effectively. In studies funded by an NIH R21 grant, the principal investigators demonstrated recently that intestinal pathology involves loss of normal support of stem cell function by an intercellular communication pathway called Wnt, and that drugs that restore Wnt pathway signaling (including lithium) can ameliorate this pathology. Here we propose to investigate lung and liver pathology, in particular to determine if Wnt activators are beneficial in these tissues as was observed in the intestine. We will also broadly investigate other affected pathways that might provide points of therapeutic intervention. We will use human induced pluripotent stem cells (iPSCs) and genome editing to generate key lung and liver cells that can be studied in culture, along with telomerase deficient mouse models, to investigate these mechanisms and test new therapeutic approaches focused on using existing FDA-approved drugs. The specific aims are: 1. Characterize defects in human DC iPSC-derived cultured type II alveolar epithelial stem cells, and test the capacity of pharmacologic manipulations aimed reversing the altered pathways to ameliorate these defects. 2. Characterize defects in human DC iPSC-derived cultured hepatocytes and stellate cells, and test the capacity of pharmacologic manipulations aimed at reversing the altered pathways to ameliorate these defects. 3. Use mouse models to address the efficacy of pharmacologic manipulations to rescue human DC mutant iPSC-derived tissues in an in vivo context. This will include determination of the minimum serum lithium concentration that will rescue human tissues transplanted into mice, and tests of lithium and other small molecules to rescue mouse models of pulmonary fibrosis and cirrhosis driven by telomere dysfunction.