Transactivation of Fetal Hemoglobin Genes for Treatment*

描述(由申请人提供)： 目前镰状细胞病的基因治疗策略主要集中在转导方面 含抗镰状珠蛋白的病毒载体对造血干细胞的影响 基因如y-、0/5-或修饰的P-珠蛋白基因。尽管这是一种可行的 方法：治疗水平珠蛋白mRNA的长期表达和 从转导基因中获得蛋白质是很难实现的。另一种选择 一种方法是用编码转录因子的基因转导干细胞 一种能特异性增强内源性伽马或β-珠蛋白基因的蛋白质 表情。在这种情况下，一种新的转录因子水平相对较低 可能会刺激高水平的伽马或5-珠蛋白基因表达。 Sangamo BioSciences是一家率先使用锌手指的公司 Cys2-His2类型的蛋白质(ZFP)与几乎 任何感兴趣的调控序列。设计师ZFP拥有更强大的 特征多于内生因素。ZFP旨在绑定到 几乎任何具有高特异性和更大亲和力的序列 内生因素。ZFP也可以设计为正常集成功能 分布在几个内生因素之间，否则就必须 被组合在一起以达到预期的结果。 特异性和高效激活γ-和β-珠蛋白基因的ZFP 培养细胞的表达将在镰状细胞的小鼠模型中进行测试 疾病。这些小鼠从人类胎儿血红蛋白(HBF)切换到人类镰刀 出生后的血红蛋白(HBS)以类似于人类患者的方式 疾病。虽然小鼠在出生时没有症状，但会发展成严重的疾病 随着这种转变的发生，成年动物如果不是全部的话也会发展成大多数 疾病的病理学。特定和高效地激活ZFP 慢病毒将转导伽马和β珠蛋白基因的表达 载体转化从镰刀鼠分离的纯化的造血干细胞。 在将这些转基因细胞移植到接受者体内后， 将对动物进行监测，以长期纠正疾病。相对的 低水平的这些强大的转录因子应该能够 激活伽马和β-珠蛋白基因到一个水平，将改善 在大多数器官系统中观察到的病理学。这些研究将提供一个坚实的 为后续的人类患者临床试验奠定基础 毁灭性的疾病。

DESCRIPTION (provided by applicant): Current gene therapy strategies for sickle cell disease focus on transduction of hematopoietic stem cells with viral vectors containing anti-sickling globin genes such as y-, 0/5- or modified P-globin genes. Although this is a viable approach, long term expression of therapeutic levels of globin mRNA and protein from transduced genes magamma be, difficult to achieve. An alternative approach is to transduce stem cells with a transcription factor gene encoding a protein that specifically enhances endogenous gamma- or beta-globin gene expression. In this case relatively low levels of a novel transcription factor may stimulate high levels of gamma- or 5-globin gene expression. Sangamo BioSciences is a company which has pioneered the use of zinc finger proteins (ZFPs) of the Cys2-His2 type with high specific affinities to almost any regulatory sequence of interest. Designer ZFPs have more robust characteristics than endogenous factors. ZFPs are designed to bind to virtually any sequence with high specificity and greater affinity than endogenous factors. ZFPs can also be designed to integrate functions normally distributed between several endogenous factors, which would otherwise have to be added combinatorially to effect a desired outcome. ZFPs that specifically and efficiently activate gamma- and beta-globin gene expression in cultured cells will be tested in a mouse model of sickle cell disease. These mice switch from human fetal hemoglobin (HbF) to human sickle hemoglobin (HbS) after birth in a manner similar to human patients with the disease. Although the mice are asymptomatic at birth, severe disease develops as the switch occurs, and adult animals develop most if not all of the pathology of the disease. ZFPs that specifically and efficiently activate gamma- and beta-globin gene expression will be transduced with lentiviral vectors into purified hematopoietic stem cells isolated from the sickle mice. After transplantation of these genetically modified cells into recipients, the animals will be monitored for long term correction of the disease. Relatively low levels of these potent transcription factors should be capable of activating gamma- and beta-globin genes to a level that will ameliorate the pathology observed in most organ systems. These studies will provide a solid foundation for subsequent clinical trials in human patients with this devastating disease.