Calpain inhibitors in models of Parkinson's disease

描述（由申请人提供）：帕金森病是一种神经退行性疾病，特别影响黑质中的多巴胺能神经元。尽管已经提出了几种假设来解释该疾病神经退行性特征的特异性，但该疾病的确切原因仍有待阐明。偶然发现的神经毒素 1-甲基-4-苯基-1,2,3,6-四氢吡啶 (MPTP) 在人类和实验动物中引起与帕金森病患者相同的神经退行性特征，这为我们对该疾病的可能原因的理解提供了重大进展。预防帕金森病神经变性的潜在靶点是钙依赖性蛋白酶钙蛋白酶。帕金森病患者死后黑质中的钙蛋白酶水平升高，颗粒细胞培养物中的MPP神经毒性与钙蛋白酶激活有关并被钙蛋白酶抑制剂阻断，并且钙蛋白酶与多种神经退行性疾病有关。我们最近获得了一系列新型有效的钙蛋白酶抑制剂，并证明了它们在预防培养海马切片中 NMDA 诱导的钙蛋白酶激活方面的功效。目前的提议旨在检验以下假设：钙蛋白酶激活在帕金森病动物模型中发挥关键作用，并且钙蛋白酶抑制剂在这些模型中具有神经保护作用。我们将首先确定钙蛋白酶抑制剂预防大鼠中脑培养切片中 MPTP 毒性的效力和功效。然后，我们将使用结构活性关系与其他测定相结合，以确定要在体内模型中测试的最佳抑制剂。最后，我们将测试以下假设：钙蛋白酶被激活，并且钙蛋白酶抑制剂对 C57BI/6 小鼠体内 MPTP 介导的神经毒性和行为损伤以及大鼠体内鱼藤酮介导的神经毒性具有神经保护作用。将测试促凋亡因子 Bid 向其活性截短形式 tBid 的转化，作为钙蛋白酶激活诱导细胞死亡的机制的一部分。这些研究将检验这样的假设：钙蛋白酶抑制剂不仅可以预防帕金森病的神经变性，还可以预防因暴露于环境毒素而导致的各种疾病。最后，由于钙蛋白酶也与肌萎缩侧索硬化症（ALS）的机制有关，因此我们的建议也可能导致这种神经退行性疾病的治疗取得重大进展。

DESCRIPTION (provided by applicant): Parkinson' s disease is a neurodegenerative disease that specifically affects dopaminergic neurons in the substantia nigra. Although several hypotheses have been proposed to account for the specificity of the neurodegenerative features of the disease, the exact cause of the disease remains to be elucidated. Significant advances in our understanding of the possible causes of the disease were provided by the serendipitous discovery that a neurotoxin, 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP), elicits a pattern of neurodegenerative features in humans and experimental animals identical to that seen in patients with Parkinson' s disease. A potential target to prevent neurodegeneration in Parkinson' s disease is the calcium-dependent protease calpain. Calpain levels are elevated in post-mortem substantia nigra of patients with Parkinson' s disease, MPP+ neurotoxicity in granule cell cultures is associated with calpain activation and blocked by calpain inhibitors, and calpain has been implicated in several neurodegenerative diseases. We have recently obtained a series of novel and potent calpain inhibitors and have demonstrated their potency in preventing NMDA-induced calpain activation in cultured hippocampal slices. The current proposal is aimed at testing the hypothesis that calpain activation plays a critical role in animal models of PD and that calpain inhibitors are neuroprotective in these models. We will first determine the potency and efficacy of calpain inhibitors to prevent MPTP toxicity in cultured slices from rat mesencephalon. We will then use structure activity relationship in conjunction with additional assays to identify the best inhibitors to be tested in in vivo models. Finally, we will test the hypothesis that calpain is activated and that calpain inhibitors are neuroprotective against MPTP-mediated neurotoxicity and behavioral impairments in vivo in C57BI/6 mice, and against rotenone-mediated neurotoxicity in rats. Conversion of the pro-apoptotic factor Bid to its active, truncated form tBid will be tested as part of the mechanisms by which calpain activation induces cell death. These studies will test the hypothesis that calpain inhibitors might prevent neurodegeneration not only in Parkinson' s disease but also in a variety of conditions resulting from exposure to environmental toxins. Finally, because calpain has also been implicated in the mechanisms underlying Amyotrophic Lateral Sclerosis (ALS), our proposal could lead to significant advances in the treatment of this neurodegenerative disease as well.