Testing OKN-007 as a potential intervention for ALS

肌萎缩侧索硬化症(ALS)是一种使人衰弱的疾病，目前尚无有效的治疗方法。的目标是 这项提议是为了确定一种有前途的新化合物的潜在有效性和作用机制， OKN-007，我们最近发现它在运动神经元衰老和死亡过程中具有神经保护作用。 ALS突变小鼠模型的初步研究。我们最近对衰老和骨质疏松症的研究引导我们 重点关注旨在保护运动神经元健康的干预措施，以减少神经和NMJ的损失 因此可以改善衰老过程中的肌肉效果。为此，我们有令人振奋的初步数据 使用OKN-007，减少衰老小鼠运动神经元损失的小分子。OKN-007是一种硝酮 PbN(α-苯基-N-叔丁基硝酮)的衍生物，一种先前已被证明有益的自旋陷阱 对癌症和其他病理的影响。硝酮化合物具有已知的抗氧化剂和抗炎作用。 OKN-007对创伤性脑损伤、局灶性脑缺血/卒中和脑损伤具有神经保护作用。 听力损失。我们在衰老小鼠中令人兴奋的初步数据显示，OKN-007可以减少运动神经元的损失， 减少后肢肌肉的失神经，减少老年小鼠腓肠肌质量的损失。这些 研究结果使我们推测，OKN-007也可能减少运动神经元的死亡，延缓疾病 肌萎缩侧索硬化症进展。事实上，我们的初步数据显示，马达得到了保护，这让我们感到鼓舞 G93A ALS小鼠模型脊髓内神经元数量与疾病进展的延缓 用OKN-007治疗。这个试点项目的目标是确认这些初步发现，并测试我们的 假设OKN-007可以保护ALS中运动神经元的丢失并延缓疾病的进展。在……里面 目的1，我们将确定OKN-007治疗对大鼠运动神经元和骨骼肌功能的影响。 肌萎缩侧索硬化症G93A突变小鼠模型的疾病进展和存活。对照组和G93A小鼠的队列 将在30日龄、发病时(95天)或3天开始在饮用水中使用OKN-007进行治疗 发病后几周(115天)以确定治疗的预防和治疗效果。老鼠会 从60日龄开始称重和评估疾病评分和进展。使用脊髓从 对照组和G93A小鼠，我们将测量运动神经元数量，炎症标志物，氧化损伤和 激活细胞死亡，这是ALS疾病进展中的已知变化。小鼠将在145天龄时被处死 年龄，疾病末期之前。为了评估已知的肌萎缩侧索硬化介导的肌肉表型，我们将测量握力 从60日龄开始的力量和NMJ的形态和神经支配状态将在145日龄时测量，如 也是骨骼肌失神经和肌肉萎缩的标志。将设立平行的小鼠队列，以 一旦我们确定了最有效的剂量，就可以根据药物治疗的反应来衡量存活率。在目标2中，我们将 通过测量确定OKN-007在运动神经元维持和存活中的潜在作用机制 应用RNA序列分析技术研究OKN-007对小鼠脊髓运动神经元基因表达的影响 目的1.这些研究将为OKN-007的作用机制提供重要信息，并有助于 确定OKN-007是否可能是维持ALS运动神经元健康的良好候选药物 病人。

Amyotrophic lateral sclerosis (ALS) is a debilitating disease with currently no effective treatments. The goal of this proposal is to define the potential effectiveness of and mechanism of action of a promising new compound, OKN-007, that we have recently found to have neuroprotective effects in motor neurons in aging and in preliminary studies in an ALS mutant mouse model. Our recent studies in aging and sarcopenia have led us to focus on interventions targeted to preserving motor neuron health to reduce loss of innervation and NMJ disruption and therefore improve muscle outcomes in aging. To this end, we have exciting preliminary data using OKN-007, small molecule that reduces loss of motor neurons in aging mice. OKN-007 is a nitrone derivative of PBN (α-phenyl-N-tert-butyl-nitrone), a spin trap that has previously been shown to have beneficial effects in cancer and other pathologies. Nitrone compounds have known antioxidant and anti-inflammatory properties, and OKN-007 has neuroprotective effects in traumatic brain injury, in focal ischemia/stroke and in hearing loss. Our exciting preliminary data in aging mice show that OKN-007 can reduce motor neuron loss, reduce denervation in hindlimb muscle and reduce loss of gastrocnemius muscle mass in older mice. These findings led us to hypothesize that OKN-007 may also reduce motor neuron death and delay disease progression in ALS. Indeed, we are encouraged by our preliminary data that show a preservation of motor neuron number in the spinal cord and a delay in disease progression in the G93A ALS mouse model treated with OKN-007. The goal of this pilot project is to confirm these preliminary findings and test our hypothesis that OKN-007 can preserve motor neuron loss in ALS and delay disease progression. In Aim 1, we will determine the effect of OKN-007 treatment on motor neuron and skeletal muscle outcomes in disease progression and survival in the G93A mutant mouse model of ALS. Cohorts of control and G93A mice will be treated with OKN-007 in drinking water beginning at 30 days of age, at disease onset (95 days), or at 3 weeks post-onset (115 days) to determine preventative and therapeutic effects of the treatment. Mice will weighed and evaluated for disease score and progression beginning at 60 days of age. Using spinal cord from control and G93A mice, we will measure motor neuron number, inflammatory markers, oxidative damage and activation of cell death, known alterations in disease progression in ALS. Mice will be sacrificed at 145 days of age, prior to disease end stage. To assess known ALS mediated muscle phenotypes, we will measure grip strength starting 60 days of age and NMJ morphology and innervation status will be measured at 145 days, as well as markers of denervation in skeletal muscle and muscle atrophy. Parallel cohorts of mice will be set up to measure survival in response to drug treatment once we determine the most effective dose. In Aim 2, we will identify potential mechanisms of action for OKN-007 in motor neuron maintenance and survival by measuring the effect of OKN-007 on gene expression using RNA seq analysis in spinal cord motor neurons from mice in Aim 1. These studies will provide important information on the mechanism of action of OKN-007 and help to establish whether OKN-007 may be a good candidate for treatment to maintain motor neuron health in ALS patients.