PILOT STUDY--HERPES SIMPLEX VIRUS FOR GENE THERAPY FOR LESCH-NYHAN SYNDROME

许多遗传性疾病会影响神经系统。的长期目标 这项拨款提案是开发无毒的单纯疱疹病毒1型， HSV-1株作为神经系统疾病的基因治疗载体。 HSV-1感染有两个特征， 为了实现这一目标：HSV-1建立终身潜伏感染， 在潜伏感染期间，只有一个病毒启动子被激活。 活跃 该启动子指导HSV-1潜伏基因的合成， 相关转录本（LAT），其在细胞的生命周期内表达。 潜伏感染者。 为了证明HSV-1的可行性， 介导的基因治疗，人次黄嘌呤-鸟嘌呤的cDNA 磷酸核糖基转移酶（HPRT）基因将置于 HSV-1 LAT基因启动子。HPRT的完全缺乏会导致Lesch- Nyhan综合征，一种严重且无法治愈的神经系统疾病。 以来 HPRT水平非常低的个体免于神经系统疾病， 功能障碍，甚至部分替代HPRT在神经系统中可能是 有治疗作用的 HPRT阴性转基因小鼠可作为动物获得 评估基因转移有效性的Lesch-Nyhan综合征模型 技术. 有几种无毒的HSV-1毒株， 复制胜任和不胜任。这些病毒不产生任何 疾病的小鼠，即使当接种脑内（k.c.）。 发展 HSV-1基因传递到神经系统，我们将i）研究几个 用于通过神经系统传播的无毒HSV-1菌株， 细胞病理学，急性感染期间病毒RNA和蛋白质表达， 以及在治疗过程中LAT表达细胞的类型、数量和分布。 潜伏期，ii）确定高水平的polyA+/调节序列的要求， HSV-1潜伏期启动子指导的cDNA表达水平在大多数 有希望的无毒菌株，和iii）研究组织 人HPRT mRNA和蛋白质在正常小鼠和HPRT中的分布 用HSV-1/HPRT重组体感染阴性转基因小鼠。

Many genetic diseases affect the nervous system. The long term goal of this grant proposal is to develop avirulent herpes simplex virus type 1 (HSV-1) strains as gene therapy vectors for nervous system disorders. There are two characteristics of HSV-1 infections that will be exploited to achieve this goal: HSV-1 establishes life-long latent infections in neurons, and during latent infection only a single viral promoter is active. This promoter directs the synthesis of the HSV-1 latency associated transcripts (LATs), which are expressed for the lifetime of the latently infected individual. To demonstrate the feasibility of HSV-1- mediated gene therapy, a cDNA for the human hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene will be placed under the control of the HSV-1 LAT gene promoter. A complete deficiency of HPRT causes Lesch- Nyhan syndrome, a severe and untreatable neurological disease. Since individuals with very low HPRT levels are spared from neurological dysfunction, even partial replacement of HPRT in the nervous system may be therapeutic. HPRT-negative transgenic mice are available as an animal model of Lesch-Nyhan syndrome to assess the effectiveness of gene transfer techniques. There are several avirulent HSV-1 strains available, both replication competent and incompetent. These viruses do not produce any disease in mice, even when inoculated intracerebrally (k.c.). To develop HSV-1 for gene delivery to the nervous system, we will i) study several avirulent HSV-1 strains for spread through the nervous system, cytopathology, viral RNA and protein expression during acute infection, and the type, number and distribution of LAT expressing cells during latency, ii)determine the polyA+/regulatory sequence requirements for high levels of HSV-1 latency promoter directed cDNA expression in the most promising avirulent strains(s), and iii) investigate the tissue distribution of human HPRT mRNA and proteins in normal mice and in HPRT- negative transgenic mice infected with the HSV-1/HPRT recombinants.