Protein Tranduction for Treatment of Krabbe Disease

描述(由申请人提供)： Krabbe病是一种退行性神经疾病，主要影响婴儿和幼儿，尽管已描述了罕见的成人发病病例。受影响的人通常在生命的头几个月出现症状。疾病进展通常很快，导致在1-2年内死亡。这种疾病是由半乳脑苷酶(GALC)基因突变引起的常染色体隐性遗传，这种突变严重影响酶的活性。明显的治疗方法包括通过基因或蛋白质疗法将活性GALC酶输送到大脑。尽管多年来在基因治疗方面取得了重大进展，但蛋白质在大脑中的稳定表达尚未实现。酶替代疗法提供了另一种可能的治疗方法，已被证明对另一种溶酶体储存障碍-I型高谢病的外周临床表现是安全有效的。然而，对于像Krabbe病这样的疾病，治疗性酶必须被输送到大脑才能产生重大的临床影响。最近，Steven Dowdy和他的同事在将大分子输送到大脑方面取得了重大进展(Schwarze等人，1999)。他们已经证明，当与来自Irv Tat蛋白的11个氨基酸的蛋白质转导结构域结合时，即使是非常大的蛋白质也可以通过血脑屏障以生物活性的形式进入大脑。在这项应用中，我们提出了产生TAT PTD/GALC融合蛋白的实验，并检查这些融合蛋白是否会到达大脑，恢复正常功能并提高Krabbe病动物模型的存活率。从这些实验中学到的经验教训可能对其他需要提供治疗性蛋白质的疾病有用，例如阿尔茨海默病和帕金森氏症。

DESCRIPTION (provided by applicant): Krabbe disease is a degenerative neurological disorder primarily affecting infants and young children although rare cases of adult onset have been described. Affected individuals typically present with symptoms in the first few months of life. Disease progression is generally rapid, leading to death within 1-2 years. The disease is inherited as an autosomal recessive trait caused by mutations in the galactocerebrosidase (GALC) gene that severely affect the activity of the enzyme. Obvious therapeutic approaches include delivery of active GALC enzyme to the brain through either gene or protein therapy. While significant advances have been made in gene therapy over the years, stable expression of proteins in the brain has not yet been achieved. Enzyme replacement therapy offers another possible therapeutic approach and has been shown to be safe and effective for the treatment of peripheral clinical manifestations in another lysosomal storage disorder, Type I Gaucher's disease. For diseases such as Krabbe disease, however, the therapeutic enzyme must be delivered to the brain to have a significant clinical impact. Recently, Steven Dowdy and colleagues have made a significant advance in the delivery of macromolecules to the brain (Schwarze et al., 1999). They have shown that that even very large proteins can cross the blood-brain barrier to enter into the brain in biologically active form when coupled to an 11 amino acids protein transduction domain derived from the IRV TAT protein. In this application we propose experiments to generate TAT PTD/GALC fusion proteins and examine whether these will get to the brain and restore normal function and enhance survivability in an animal model of Krabbe disease. The lessons learned from these experiments may be useful for other diseases where delivery of a therapeutic protein may be desired, such as Alzheimer's disease and Parkinson's.