Role of Potent Trophic Factors on Glia and Motor Neurons in ALS

描述（由申请人提供）：肌萎缩性侧索硬化症（ALS）是一种毁灭性的运动神经元疾病，可导致肌肉瘫痪和最终的呼吸衰竭和死亡。肌萎缩性侧索硬化症的根本原因尚不清楚，也无法治愈。许多报告，包括我们实验室的工作，已经证明了神经营养因子在家族性ALS （fALS）啮齿动物模型中的高度治疗潜力。事实上，胰岛素样生长因子-1 （IGF-1）、神经胶质源性神经营养因子（GDNF）和血管内皮生长因子（VEGF）在ALS啮齿类动物模型发病时传递，在延缓疾病进展方面具有深远的作用。最近的研究令人惊讶地表明，表达超氧化物歧化酶突变的星形胶质细胞和小胶质细胞可以加剧运动神经元的死亡，这支持了早期关于ALS是一种非细胞自主疾病的研究。具体来说，神经胶质细胞已被证明会产生异常活动，分泌有毒信号，导致运动神经元死亡。基于这些结果，旨在中和神经胶质细胞毒性的疗法将对ALS患者非常有益。在基因筛选鉴定出新的ALS诱导基因之前，应该开发出可能延长ALS患者生命的治疗方法。神经营养因子延长生存和运动功能的机制仍然是难以捉摸的。我们实验室最近的初步工作表明，IGF-1和VEGF都可以抑制突变的胶质细胞介导的毒性。在本研究中，我们将研究IGF-1、VEGF和GDNF在抑制异常胶质活性和延缓运动神经元死亡方面的相对效率。具体来说，这些因素将利用我们实验室最近开发的宝贵的ALS体外模型进行测试。我们还将在体内AAV（腺相关病毒）基因传递范式中测试这些因素，该范式有效地靶向脊髓的所有区域。我们将分析这些营养因子介导其对星形胶质细胞影响的机制，最后，我们将利用我们在AAV载体基因传递到中枢神经系统方面的专业知识，在家族性fALS小鼠中测试这些因子的最佳组合。本提案的具体目标是：在体外fALS模型中，确定有效的神经营养素对神经保护和减少胶质细胞毒性的效率。具体目标2)在体外fALS模型中，确定IGF-1、VEGF和GDNF是否联合作用改变异常的ALS胶质活性并提供附加的神经保护。具体目标3)确定联合神经营养因子治疗是否对fALS小鼠模型有益。肌萎缩性侧索硬化症（ALS）是一种毁灭性的运动神经元疾病，可导致肌肉麻痹和最终的呼吸衰竭和死亡。肌萎缩性侧索硬化症的根本原因尚不清楚，也无法治愈。我们已经证明神经营养因子在家族性ALS啮齿动物模型中具有很高的治疗潜力，包括IGF-1、VEGF和GDNF。我们的建议侧重于评估最佳营养因子单独或组合，以延缓运动神经元退化。我们已经开发了一种基于肌萎缩侧索硬化症的体外模型，该模型利用了运动神经元和含有肌萎缩侧索硬化症星形胶质细胞的干细胞。随后，我们将在这种毁灭性疾病的啮齿动物模型中使用基因传递来测试这些因素的最佳组合，以确定这种使人衰弱的疾病的最佳治疗方法。

DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease, which results in muscle paralysis and ultimate respiratory failure and death. The underlying cause for ALS remains unknown with no cure. Numerous reports, including work from our laboratory have demonstrated the potential for neurotrophic factors to be highly therapeutic in rodent models of familial ALS (fALS). Indeed, Insulin-like growth factor-1 (IGF-1), glial derived neurotrophic factor (GDNF), and vascular endothelial growth factor (VEGF) delivered at disease onset in ALS rodent models have demonstrated profound effects in delaying disease progression. Recent studies have surprisingly demonstrated that astrocytes and microglia expressing a mutation in the enzyme superoxide dismutase can exacerbate motor neuron death, supporting earlier studies that ALS is a non-cell autonomous disease. Specifically, glial cells have been shown to develop aberrant activity, secreting toxic signals that lead to motor neuron demise. Based on these results, therapies designed to neutralize glial cell toxicity would be highly beneficial to ALS patients. Until genetic screening identifies new ALS inducing genes, therapies that could potentially prolong the lives of ALS patients should be developed. The mechanism by which neurotrophic factors prolong survival and motor function has remained elusive. Recent preliminary work by our laboratory has demonstrated that both IGF-1 and VEGF can act to suppress the mutant glial cell mediated toxicity. In this proposal, we will investigate the relative efficiency of IGF-1, VEGF and GDNF to suppress aberrant glial activity and delay motor neuron death. Specifically, these factors will be tested utilizing an invaluable in vitro model for ALS that was recently developed in our laboratory. We will also test these factors in an in vivo AAV (Adeno-Associated Virus) gene delivery paradigm that efficiently targets all regions of the spinal cord. We will analyze the mechanism by which these trophic factors mediate their effects on astrocytes and finally, we will test an optimal combination of these factors in familial fALS mice using our expertise in AAV vector gene delivery to the CNS. The specific aims of this proposal are: Specific Aim 1.) To determine the efficiency of neuroprotection and reduction of glial cell toxicity using potent neurotrophins in an in vitro based model of fALS. Specific Aim 2.) To determine whether IGF-1, VEGF, and GDNF act in combination to alter aberrant ALS glial activity and provide additive neuroprotection in an in vitro based model of fALS. Specific Aim 3.) To determine whether a combinatorial neurotrophic factor therapy is beneficial in a mouse model of fALS. PUBLIC HEALTH RELEVANCE Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease, which results in muscle paralysis and ultimate respiratory failure and death. The underlying cause for ALS remains unknown with no cure. We have shown the potential for neurotrophic factors to be highly therapeutic in rodent models of familial ALS including IGF-1, VEGF, and GDNF. Our proposal focuses on evaluating optimal trophic factors individually or in combination to delay motor neuron degeneration. We have developed an in vitro based model of ALS that utilizes stem cells directed to motor neurons and ALS containing astrocytes, which recapitulates the disease. We will subsequently test the optimal combination of these factors using gene delivery in a rodent model of this devastating disease in order to define an optimal therapy for this debilitating disorder.