GENE DELIVERY TO THE CNS TO MODULATE EPILEPSY

描述：(改编自申请者摘要)常规医疗疗法 对于中枢神经系统疾病，无论是外科的还是药理学的，通常都有广泛的 对与正在治疗的疾病无关的大脑区域产生不希望看到的影响。在……里面 对此，研究人员正在开发一种“分子神经外科”，一种 使用转基因病毒治疗神经系统疾病的新方法 调节神经系统细胞的载体。这个项目的目标是 提供编码谷氨酸的工程转录单位(转基因) 脱羧酶(GAD)通过缺陷单纯疱疹病毒(HSV)载体进入细胞 中枢神经系统，以调节神经元活动。GAD直接转换为 大脑中主要的兴奋性氨基酸谷氨酸转化为γ-氨基丁酸 酸(GABA)是主要的抑制性氨基酸。有缺陷的HSV载体是一种 在体外和体内向中枢神经系统细胞传递转基因的有效手段 活着。含有大鼠GAD转基因的载体(GAD65和67亚型)和 将产生具有不同启动子的植物来靶向GAD表达和 规范GAD活动。GAD转基因表达对GABA的影响 新陈代谢将在原代中枢神经系统细胞培养中进行检测。小说的能力 将确定GABA的合成和释放以抑制癫痫样活动 在体外“癫痫”模型中，使用皮质培养物和大鼠脑片。在……里面 GAD转基因在大脑中的活体传递和表达将使我们能够 开始评估这种定点基因疗法的功能影响。 通过直接调节局部神经递质水平的能力 在大脑中原位引入相关基因可以应用于 多种神经系统疾病。GABA水平的变化在 癫痫、亨廷顿病、帕金森氏症、疼痛和精神分裂症。 超过200万美国人患有癫痫，其中约有25 %的人对药物治疗无效，可以接受手术治疗 心理治疗。目前的神经外科手术是高度侵入性的，通常涉及 切除正常大脑的大片区域。这项建议结合了这些技能 用于高度局部载体应用的神经外科，分子生物学用于 有缺陷的载体构建，用于GAD表征的细胞生物学，以及 用于新的GABA合成功能分析的生理学。

DESCRIPTION: (adapted from applicant's abstract) Conventional medical therapies for CNS disorders, both surgical and pharmacological, often have widespread unwanted effects on brain areas not involved in the disorder being treated. In response to this the investigators are developing "molecular neurosurgery", a novel approach to nervous system disorders that uses genetically modified viral vectors to modulate cells in the nervous system. The goal of this project is to deliver an engineered transcription unit (transgene) encoding glutamic acid decarboxylase (GAD) via defective herpes simplex virus (HSV) vectors to cells of the CNS, in order to modulate neuronal activity. GAD directly converts glutamate, the major excitatory amino acid in the brain, to gamma aminobutyric acid (GABA), the major inhibitory amino acid. Defective HSV vectors are an efficient means to deliver transgenes to cells of the CNS, both in vitro and in vivo. Vectors containing GAD transgenes from rat (GAD65 and 67 isoforms) and plant, with different promoters will be generated to target GAD expression and regulate GAD activity. The effect of GAD transgene expression on GABA metabolism will be examined in primary CNS cell cultures. The ability of novel GABA synthesis and release to inhibit epileptiform activity will be determined in in vitro 'seizure' models, using cortical cultures and rat brain slices. In vivo delivery and expression of the GAD transgene in the brain will allow us to begin to evaluate the functional impact of this site-directed gene therapy. The ability to modulate localized levels of neurotransmitters by directly introducing the relevant genes in situ in the brain has applications to numerous nervous system disorders. Alterations in GABA levels play a role in epilepsy, Huntington's disease, Parkinson's disease, pain and schizophrenia. Over 2 million Americans suffer from epilepsy, of which approximately 25 percent are refractory to drug treatment and are candidates for surgical therapy. Current neurosurgical procedures are highly invasive and often involve the removal of large areas of normal brain. This proposal combines the skills of neurosurgery for highly localized vector application, molecular biology for defective vector construction, cell biology for GAD characterization, and physiology for functional analysis of novel GABA synthesis.