A mouse model Niemann-Pick type C disease to test proteostasis therapies

C型尼曼-皮克病是一种致命性常染色体隐性遗传性脂肪堆积症。 患者出现临床上的异质性表型，包括进行性 神经退化和过早死亡。疾病通常由功能丧失突变引起。 在NPC1基因(95%的病例)中，编码一个多通道跨膜糖蛋白需要 用于将未酯化的胆固醇从晚期的内切体和溶酶体中输出。最常见的 致病突变(约20%的病例)是异亮氨酸到苏氨酸的位置替代 1061(I1061T)。I1061T NPC1在内质网中错误折叠并被迅速降解 通过蛋白酶体和内质网自噬。重要的是，I1061T在体外瞬时过表达 兰尼定受体拮抗剂驱动对I1061T NPC1患者成纤维细胞的作用 将突变的NPC1运输到溶酶体，在那里它仍然起作用。这些观察结果已经 激发了人们对开发蛋白质平衡调节剂治疗疾病的兴趣。这些努力依赖于 在小鼠NPC1基因中插入I1061T突变的基因靶向小鼠 测试治疗策略。然而，我们的初步数据表明，有标记的 人和小鼠I1061T NPC1蛋白在细胞内处理方式的差异 质量控制机械。这些以前未被认识到的差异突显了 需要开发一种新的模型系统，可靠地复制人类I1061T NPC1蛋白稳态。 这个项目的基本原理是，开发人源化的I1061T NPC1小鼠将能够在体内 Niemann-Pick C.的蛋白抑制疗法测试以实现这一总体目标 在应用方面，我们将使用遗传和生化方法来实现以下具体目标： 人源化I1061T NPC1小鼠的建立和鉴定。我们预计以鼠标为目标 表达人类I1061T NPC1蛋白的NPC1基因将产生一个强大的疾病模型，在 突变的NPC1蛋白的哪种错误折叠和运输与人类的行为非常相似 I1061T NPC1 AS发生在Niemann-Pick C患者中。

Niemann-Pick disease type C is an invariably fatal autosomal recessive lipid storage disease. Patients develop a clinically heterogeneous phenotype that includes progressive neurodegeneration and early death. Disease is commonly caused by loss-of-function mutations in the NPC1 gene (95% of cases), encoding a multipass transmembrane glycoprotein required for exporting unesterified cholesterol from late endosomes and lysosomes. The most common disease-causing mutation (~20% of cases) is an isoleucine to threonine substitution at position 1061 (I1061T). I1061T NPC1 misfolds in the endoplasmic reticulum (ER) and is rapidly degraded by the proteasome and ER-autophagy. Importantly, transient over-expression of I1061T in vitro or treatment of I1061T NPC1 patient fibroblasts with ryanodine receptor antagonists drives trafficking of mutant NPC1 to the lysosome where it is still functional. These observations have spurred interest in developing proteostasis modulators to treat disease. These efforts have relied on gene targeted mice in which the I1061T mutation was inserted into the mouse Npc1 gene to test therapeutic strategies. However, our preliminary data indicate that there are marked differences in how the human and mouse I1061T NPC1 proteins are handled by the cellular quality control machinery. These previously unappreciated differences underscore the critical need to develop a new model system that reliably reproduces human I1061T NPC1 proteostasis. The rationale for this project is that developing humanized I1061T NPC1 mice will enable in vivo testing of proteostatic therapeutics for Niemann-Pick C. To attain the overall objective of this application, we will use genetic and biochemical approaches to pursue the following specific aim: Develop and characterize humanized I1061T NPC1 mice. We expect that targeting the mouse Npc1 gene to express human I1061T NPC1 protein will generate a robust model of disease, in which misfolding and trafficking of the mutant NPC1 protein closely mimics the behavior of human I1061T NPC1 as occurs in Niemann-Pick C patients.