Pathophysiology and Gene Replacement Strategies for Arginase Deficiency

描述（由申请人提供）：尿素循环是哺乳动物氨解毒的主要途径。精氨酸酶I缺乏症被认为是尿素循环障碍中最不常见的，可导致高精氨酸血症。在人类中，缺乏这种酶的临床特征是进行性精神损害、痉挛、生长迟缓和周期性高氨血症发作。本研究有两方面的目的：1)利用病毒载体对精氨酸酶缺乏症进行基因校正，并对校正后的动物进行行为学和生化检查；2)评价精氨酸酶缺乏症患者精氨酸水平升高及相关代谢物（除氨外）对脑发育和智力发育迟滞的影响。初步资料：课课组构建并表征了精氨酸酶I敲除小鼠；2)用重组辅助物依赖腺病毒载体和腺相关病毒载体证明至少短期（如果不是更长）的纠正和挽救；3)敲除小鼠神经元中处于细胞周期S期的细胞比例较高；4)基因敲除小鼠分化后的神经元形态更成熟；5)基因敲除神经元可诱导与氧化损伤保护相关的基因。在目标1中，重组病毒载体将用于拯救小鼠免于死亡，并更好地了解新生儿基因治疗面临的挑战，包括细胞快速增殖和单独载体与整合载体的潜在损失。在Aim 2中，高精氨酸血症对神经系统的影响将在体外与GABA和谷氨酰胺合成/释放一起进行检查，并进行广泛的微阵列分析。在Aim 3中，高精氨酸血症对神经系统的影响将在体内与GABA和谷氨酰胺的合成/释放、胍类化合物的测定以及一氧化氮及其代谢物可能对神经系统损伤的作用一起进行研究。这些研究的成功完成将为精氨酸酶I缺乏症和相关疾病的脑损伤机制提供分子理解。此外，预计通过这些研究将开发出精氨酸酶缺乏症的基因替代策略，并且在治疗具有快速增殖发育组织的新生儿中获得的信息将适用于其他先天性代谢错误。

DESCRIPTION (provided by applicant): The urea cycle is the major pathway for detoxification of ammonia in mammals. Arginase I 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 develop gene-based correction of arginase deficiency with viral vectors and to closely examine corrected animals behaviorally and biochemically; and 2) to evaluate the role elevated arginine and related metabolites (beyond just ammonia) have on the developing brain and development of mental retardation in arginase deficiency. Preliminary data: Our research group has: 1) constructed and characterized the arginase I knockout mouse; 2) demonstrated at least short-term (if not longer) correction and rescue with recombinant helper-dependent adenoviral vectors and adeno-associated viral vectors; 3) demonstrated that knockout mouse neurons have a higher percentage of cells in the S phase of the cell cycle; 4) shown that differentiated neurons from knockout mice have a more mature morphology; and 5) shown that knockout neurons demonstrate induction of genes related to protection from oxidative damage. In Aim 1, recombinant viral vectors will be used to rescue mice from lethality and to obtain a better understanding of the challenges facing neonatal gene therapy that involve rapid cellular proliferation and potential loss with episomal vs. integrated vectors. In Aim 2, the effect of hyperargininemia on the nervous system will be examined in vitro along with GABA and glutamine synthesis/release, and extensive microarray analysis. In Aim 3, the effect of hyperargininemia on the nervous system will be examined in vivo along with GABA and glutamine synthesis/release, guanidino compound determination, and the role that nitric oxide and its metabolites may have on nervous system injury. Successful completion of the proposed studies will provide a molecular understanding of the mechanism of injury to the brain in arginase I deficiency and related disorders. In addition, it is expected that a gene replacement strategy will have been developed for arginase deficiency through these studies and that the information obtained in treating neonates that have developing tissues undergoing rapid proliferation will be applicable to other inborn errors of metabolism.