Mechanism of Selective Toxicity of SOD1 Mutants in ALS

描述（由申请人提供）：1993年，首次报道了 对编码Cu，Zn超氧化物歧化酶（SOD 1）的基因进行突变， 与一种家族性肌萎缩侧索硬化症（ALS）有关。以来 当时，已经发现SOD 1的70多个单一氨基酸突变， 导致人类的ALS。转几种人SOD 1基因中任何一种的小鼠 突变体在一个时间依赖性的过程中逐渐发展并最终致命的麻痹， 让人联想到人类ALS。对转基因小鼠的研究清楚地表明 SOD 1通过获得的功能是有毒的，因为突变体产生疾病， 即使在总SOD 1酶活性显著增加的情况下。尽管 有压倒性的证据表明， 功能仍然难以捉摸。同样令人困惑的是， 突变体仅对运动神经元有毒，尽管它们在所有神经元中都表达。 细胞类型。基于已发表的结果，SOD 1的体外表征 突变体，在培养的运动神经元中的研究，以及来自 转基因小鼠，我们已经制定了一个假设，可以解释为什么所有 ALS相关的SOD 1突变可以通过一种共同的机制产生毒性，为什么 毒性仅表现在运动神经元中。假设：我们建议， 缺锌（含铜）SOD 1是常见的毒性表型 ALS相关的SOD 1突变体，锌缺乏的SOD 1是有害的，通过其 利用抗坏血酸、氧气和一氧化氮催化 过亚硝酸根细胞毒素的形成， 蛋白质--它们强烈地与锌结合， 神经元-有助于形成锌缺乏SOD 1优先在 运动神经元本研究提出：具体目标1）测定缺锌 最广泛使用的ALS动物模型（G93 A转基因小鼠）中的SOD 1， 确定造成其积累的因素，具体目标2）， 确定导致SOD 1介导的氧化剂产生的条件及其 与毒性蛋白质聚集的潜在关系，以及具体目标3） 评价两类化合物的体内功效， 培养的运动神经元从锌缺乏SOD 1的毒性作用， 提高G93 A小鼠的存活率。

DESCRIPTION (provided by applicant): In 1993, it was first reported that a mutation to the gene coding for Cu,Zn superoxide dismutase (SOD1) was associated with a form of familial amyotrophic lateral sclerosis (ALS). Since that time, more than 70 single amino acid mutations to SOD1 have been found to cause ALS in humans. Mice transgenic for any one of several of the human SOD1 mutants develop progressive and ultimately lethal paralysis in a time-dependent manner reminiscent of human ALS. Studies in transgenic mice clearly indicate that SOD1 is toxic via a gained function because the mutant produces disease even in the presence of marked increases in total SOD1 enzyme activity. Despite the overwhelming evidence for a gained toxic function, the exact nature of that function has remained elusive. Equally puzzling has been the fact that SOD1 mutants are toxic only to motor neurons even though they are expressed in all cell types. Based on published results, in vitro characterizations of SOD1 mutants, studies in cultured motor neurons, and preliminary data from transgenic mice, we have formulated a hypothesis which may explain how all ALS-associated SOD1 mutations can be toxic via a common mechanism and why toxicity is manifested only in motor neurons. HYPOTHESIS: We are proposing that zinc-deficient (copper-containing) SOD1 is the common toxic phenotype of ALS-associated SOD1 mutants, that zinc-deficient SOD1 is injurious via its ability to utilize ascorbate, oxygen, and nitric oxide to catalyze the formation of the cytotoxin peroxynitrite, and that neurofilament proteins--which avidly bind zinc and are very abundant in motor neurons--contribute to the formation of zinc-deficient SOD1 preferentially in motor neurons. This study proposes: Specific Aim 1) to measure zinc-deficient SOD1 in the most widely used animal model of ALS (G93A transgenic mice) and determine the factors responsible for its accumulation, Specific Aim 2) to determine the conditions which lead to SOD1-mediated oxidant generation and its potential relationship to toxic protein aggregation, and Specific Aim 3) to evaluate the in vivo efficacy of two classes of compounds which protect cultured motor neurons from the toxic effects of zinc-deficient SOD1 and which enhance survival in G93A mice.