Mechanisms of Substrate Reduction Therapy for Niemann-Pick C Disease

描述（由申请人提供）：尼曼-皮克C型（NPC）病是一种胆固醇-糖鞘脂（GSL）溶酶体贮积障碍，最常见的原因是NPC1的缺陷，NPC1是一种跨膜蛋白，被认为对溶酶体的底物后吞运输至关重要。大多数受影响的儿童在出生时表现正常，在早期发展为进行性神经系统疾病，并在第二个十年死亡。我们率先开发了两种治疗这种疾病的化合物。第一种，n -丁基脱氧诺吉霉素（NB-DNJ）或米卢司他是GSL合成的抑制剂，而第二种，羟丙基-环糊精（HPBCD）是fda批准的用于药物溶解的赋形剂。在NPC1疾病小鼠模型中，这两种化合物都能有效延缓神经系统疾病的发作和延长寿命（分别为25%和100%）。然而，这两种药物都不知道其有效的确切机制。对于米卢司他，口服Npc1小鼠后神经节苷脂储存持续减少的证据缺乏。同样，对于HPBCD，虽然在Npc1小鼠中治疗后胆固醇和GSL储存都大大减少，但这种益处的机制完全未知，甚至对其穿越血脑屏障的能力也存在争议。本项目将开展一系列互补的体内和体外研究，采用现有的和新型的试剂和动物模型，定量高分辨率成像，生化和遗传评估，每一个都针对鼻咽癌疾病的治疗机制。我们的前两个目标是精确定义HPBCD在减少神经元中胆固醇/GSL储存方面的作用机制，并批判性地重新检查和评估米卢司他减少GSL合成的能力，以此作为其对神经元存活有益影响的基础。我们的第三个目标是使用无偏倚的基因分析方法来探索每种药物影响的全范围代谢途径。利用这些目标的经验教训，新的组合治疗策略将在第四个目标中进行测试，作为大幅度改善鼻咽癌儿童治疗的一种手段。

DESCRIPTION (provided by applicant): Niemann-Pick type C (NPC) disease is a cholesterol-glycosphingolipid (GSL) lysosomal storage disorder caused most commonly by defects in NPC1, a transmembrane protein believed critical in retroendocytic trafficking of substrates from lysosomes. Most affected children appear normal at birth, develop progressive neurological disease in their early years and die in their second decade. We have pioneered the development of two compounds for this disorder. The first, N-butyldeoxynojirimycin (NB-DNJ) or miglustat is a documented inhibitor of GSL synthesis, whereas the second, hydroxypropyl ¿-cyclodextrin (HPBCD), is an FDA-approved excipient used for drug solublization. Both compounds are efficacious in delaying onset of neurological disease and prolonging life (by 25% and 100%, respectively) in the mouse model of NPC1 disease. Yet neither drug is understood in terms of the precise mechanism responsible for its effectiveness. For miglustat, evidence for sustained reductions in ganglioside storage following oral administration to Npc1 mice is lacking. Similarly, for HPBCD, while both cholesterol and GSL storage are substantially reduced following treatment in Npc1 mice, the mechanism underlying this benefit is completely unknown, and indeed controversy continues even over its ability to cross the blood brain barrier. This proposal will carry out a series of complementary in vivo and in vitro studies employing current and novel reagents and animal models, and quantitative high-resolution imaging, biochemical and genetic evaluations, each directed at treatment mechanisms for NPC disease. Our first two aims are to precisely define HPBCD's mechanism of action in reducing cholesterol/GSL storage in neurons and to critically re-examine and assess miglustat's ability to reduce GSL synthesis as a basis for its beneficial impact on neuron survival. Our third aim uses an unbiased gene analysis approach to explore the full range of metabolic pathways impacted by each drug. Capitalizing on lessons learned in these aims, new combinatorial treatment strategies will be tested in the fourth aim as a means to substantially improve therapy for children with NPC disease.