Cell and Gene Replacement Strategies for Arginase Deficiency

项目摘要/摘要 尿素循环是哺乳动物体内氨解毒的主要途径。精氨酸酶1缺乏被认为是 是尿素循环障碍中最不常见的一种，会导致高精氨酸血症。在人类中，缺乏 这种酶的临床特征是进行性智力障碍、痉挛、生长迟缓和 周期性的高氨血症。这一建议有两个方面：1)继续推进以基因为基础的 利用适当的小鼠模型治疗精氨酸酶缺乏症；病毒载体和基因组校正 将应用技术来检查动物是否可以在行为和生物化学上得到纠正；以及2) 评价将肝细胞放置在生物活性支架上以提供尿素的IPSC衍生细胞治疗方法 循环函数。初步数据：我们的研究小组(除其他发现外)：1)构建和 鉴定了精氨酸酶1基因敲除小鼠；2)证明了重组人的长期存活和挽救 腺相关病毒载体；3)证明只有低水平的尿失禁才是长期治疗的必要条件。 存活；4)通过一系列的行为学测试表明，经过精氨酸酶基因敲除处理的动物缺乏神经 系统异常，与产仔犬相比，在学习或行为上没有差异；5)如图所示 外周代谢可导致循环血浆精氨酸的控制；以及6)精氨酸酶基因的丢失 表达导致神经元的内在兴奋性和树突状突起的异常。在目标1中，长期 通过病毒载体基因治疗在肌肉中表达精氨酸酶2，以检测生化和 精氨酸酶缺乏症小鼠模型的表型纠正。这种方法可以避免中和免疫。 零突变患者的反应。在目标2中，研究将检查在支架上生长的辅助肝脏 可以提供最小的尿素循环功能，以导致表型纠正高精氨酸血症。这 这种方法可能对其他尿素循环障碍也是成功的。随着拟议研究的圆满完成， 预计基因和细胞替代的新疗法将使患者更近一步 精氨酸酶缺乏症。

Project Summary/Abstract The urea cycle is the major pathway for detoxification of ammonia in mammals. Arginase 1 deficiency is thought to be the least common of the urea cycle disorders and results in hyperargininemia. In humans, deficiency of this enzyme is characterized clinically by progressive mental impairment, spasticity, growth retardation, and periodic episodes of hyperammonemia. This proposal is two-fold: 1) to continue to advance gene-based therapies for arginase deficiency utilizing appropriate murine models; viral vectors and genomic correction technology will be applied to examine if animals can be corrected behaviorally and biochemically; and 2) to evaluate an iPSC-derived cell therapy approach with hepatocytes placed on bioactive scaffolds to supply urea cycle function. Preliminary data: Our research group has (amongst other findings): 1) constructed and characterized the arginase 1 knockout mouse; 2) demonstrated long-term survival and rescue with recombinant adeno-associated viral vectors; 3) demonstrated that only low-level ureagenesis is necessary for long-term survival; 4) shown that, using an array of behavioral tests, that treated arginase knockout animals lack nervous system abnormalities and there is no difference in learning or behavior when compared to littermates; 5) shown that peripheral metabolism can result in control of circulating plasma arginine; and 6) loss of arginase gene expression results in abnormalities of intrinsic excitability and the dendritic arbor of neurons. In Aim 1, long-term expression of arginase 2 in muscle by viral vector gene therapy will be performed to examine for biochemical and phenotypic correction in a murine model of arginase deficiency. This approach may avoid neutralizing immune responses in patients with null mutations. In Aim 2, studies will examine if an auxiliary liver grown on scaffolds can supply the minimal urea cycle function necessary to lead to phenotypic correction of hyperargininemia. This approach may be successful for other urea cycle disorders. With successful completion of the proposed studies it is expected that a new therapy with gene and cell replacement will be one step closer for patients afflicted with arginase deficiency.