New Inactivators of GABA Aminotransferase for Epilepsy and Neuropathic Pain

项目摘要 参与调节脑神经元活动的两种主要神经递质是γ-氨基丁酸 酸是分布最广泛的抑制性神经递质之一，L-谷氨酸是一种兴奋性神经递质 神经递质。GABA的浓度受两种依赖于吡哆醛的5‘-磷酸(PLP)的调节 酶，L-谷氨酸脱羧酶，催化L-谷氨酸转化为γ-氨基丁酸； γ-氨基丁酸转氨酶(GABA-AT)，将γ-氨基丁酸降解为琥珀酸半醛，将α- 酮戊二酸转化为L-谷氨酸。当GABA浓度降至阈值以下时， 导致抽搐；提高GABA水平可终止癫痫发作。当癫痫被广义地定义为任何疾病时 以反复抽搐为特征，那么超过1%的世界人口(包括300万 美国人)可以被归类为癫痫。提高GABA水平的一种方法是用一种分子 跨越血脑屏障(BBB)，抑制/灭活GABA-AT。这有效地抑制了过度 在不影响基础神经元放电的情况下进行神经活动。神经病理性疼痛，包括化疗引起的疼痛 周围神经病变(60%以上接受化疗的癌症患者的问题)，影响3- 占世界人口的17%。目前对疼痛的治疗不足会因不良副作用而加剧， 如滥用倾向、镇静和精神状态改变，这些都限制了治疗的效用。的两个功能 已经确定的神经病理性疼痛是GABA水平降低和脊髓GABA能抑制功能。 这项建议的目标是设计和评估我们的新的基于机制的灭活剂类似物 先前成功的GABA-AT失活剂用于增强效力并阐明失活机制 计算机建模和结晶学(刘达利博士研究我们的晶体结构)作为设计的驱动力。 新的灭活剂正在被设计用于选择性的GABA-AT灭活。我们的新分子的有效性 将由我的合作者Andrea Hohmann博士测试它们对各种神经性疼痛的效果，包括 化疗引起的周围神经病变。 授权R01 NS123057中的特殊目标1-3描述了许多被提议的分子 合成用于试验的GABA-AT失活剂。我们有20年历史的高效液相色谱/质谱仪已经修理好了。 很多次，现在已经超过8个月没有工作了。这大大降低了我们的工作效率， 合成新分子，使分析和鉴定中间体和产品的时间非常长。 因此，提出了以下目标： 具体目标1.购买和安装新的质谱计和高性能液体 层析法。将在供应商的协助下制定常规培训/维护政策。

Project Summary The two principal neurotransmitters involved in the regulation of brain neuronal activity are γ-aminobutyric acid (GABA), one of the most widely distributed inhibitory neurotransmitters, and L-glutamic acid, an excitatory neurotransmitter. The concentration of GABA is regulated by two pyridoxal 5'-phosphate (PLP)-dependent enzymes, L-glutamic acid decarboxylase (GAD), which catalyzes the conversion of L-glutamate to GABA, and GABA aminotransferase (GABA-AT), which degrades GABA to succinic semialdehyde and converts α- ketoglutarate to L-glutamic acid. When the concentration of GABA diminishes below a threshold level, convulsions result; raising GABA levels terminates the seizure. When epilepsy is defined broadly as any disease characterized by recurring convulsive seizures, then over 1% of the entire world population (including >3 million Americans) can be classified as having epilepsy. One approach to raise GABA levels is with a molecule that crosses the blood-brain barrier (BBB) and inhibits/inactivates GABA-AT. This effectively dampens excessive neural activity without affecting basal neuronal firing. Neuropathic pain, including chemotherapy-induced peripheral neuropathy (a problem for more than 60% of cancer patients treated with chemotherapy), affects 3- 17% of the world population. Inadequate current treatments of pain are exacerbated by adverse side effects, such as abuse liability, sedation, and altered mental status, which limit treatment utility. Two features of neuropathic pain that have been identified are reduced GABA levels and spinal GABAergic inhibitory function. The objective of this proposal is to design and evaluate new mechanism-based inactivator analogs of our previously successful GABA-AT inactivators to enhance potency and elucidate inactivation mechanisms using computer modeling and crystallography (Dr. Dali Liu does our crystal structures) as the driving force for design. New inactivators are being designed for selective GABA-AT inactivation. The effectiveness of our new molecules will be tested by my collaborator, Dr. Andrea Hohmann, for their effect on various neuropathic pains, including chemotherapy-induced peripheral neuropathy. Specific Aims 1-3 in grant R01 NS123057 describes a multitude of molecules that were proposed to synthesize for testing as inactivators of GABA-AT. Our 20-year old HPLC/mass spectrometer has been repaired multiple times and now has not functioned in over 8 months. This dramatically diminishes our productivity to synthesize new molecules, making the time to analyze and identify intermediates and products very long. Consequently, the following aim is proposed: Specific Aim 1. Purchase and installation of a new mass spectrometer coupled with high-performance liquid chromatography. A routine training/maintenance policy will be established, assisted by the vendor.