Protonged expression of a tysosomat enzyme in mouse tiver after Sleeping Beauty transposon-mediated gene detivery:: imptications for non-virat gene therapy of mucopotysaccharidoses
Protonged expression of a tysosomat enzyme in mouse tiver after Sleeping Beauty transposon-mediated gene detivery:: imptications for non-virat gene therapy of mucopotysaccharidoses
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DOI：
10.1002/jgm.1028
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发表时间：
2007-05-01
影响因子：
3.5
通讯作者：
Hackett, Perry B.
中科院分区：
文献类型：
--
作者：
Aronovich, Elena L.;Bell, Jason B.;Hackett, Perry B.
背景睡美人转座子系统是一种非病毒载体系统，可以将精确的序列整合到染色体上。我们评估了SB转座子系统作为粘多糖样变性（MPS）I型和VII型基因治疗的工具。方法我们构建了SB转座子质粒，用于高水平表达人β-葡萄糖醛酸酶（hGUSB）或α-L-艾杜糖醛酸酶（hIDUA）。通过快速、高容量尾静脉注射将具有和不具有SB转座酶的质粒递送至小鼠肝脏。我们研究了持续时间的表达治疗酶活性，转基因存在的PCR，溶酶体病理甲苯胺蓝染色和细胞介导的免疫反应组织学和免疫组化staining.Results转基因频率，分布的转基因和酶的表达在肝脏和水平的转基因酶所需的改善溶酶体病理NIPS I和VII小鼠估计。在没有免疫调节的情况下，血浆中的初始GUSB和IDUA活性达到野生型（WT）水平的>100倍，但在注射后4周内下降至背景。在免疫调节转座子处理的MPS I小鼠中，血浆IDUA在三分之一的MPS I小鼠中以高达100倍的WT活性持续3个月以上，这足以逆转肝脏和部分远端器官中的溶酶体病变。在IDUA转座子处理的WT小鼠中，肝切片的组织学和免疫组织化学检查显示注射后10天的炎症主要由单核细胞组成，其中一些是CD 4或CD 8阳性的。结论我们的研究结果证明了用单剂量治疗性SB转座子实现肝脏中溶酶体酶的延长表达和逆转成年小鼠MPS疾病的可行性。版权所有（C）2007约翰威利父子有限公司
Background The Sleeping Beauty (SB) transposon system is a non-viral vector system that can integrate precise sequences into chromosomes. We evaluated the SB transposon system as a tool for gene therapy of mucopolysaccharidosis (MPS) types I and VII.Methods We constructed SB transposon plasmids for high-level expression of human beta-glucuronidase (hGUSB) or alpha-L-iduronidase (hIDUA). Plasmids were delivered with and without SB transposase to mouse liver by rapid, high-volume tail-vein injection. We studied the duration of expressed therapeutic enzyme activity, transgene presence by PCR, lysosomal pathology by toluidine blue staining and cell-mediated immune response histologically and by immunohistochemical staining.Results Transgene frequency, distribution of transgene and enzyme expression in liver and the level of transgenic enzyme required for amelioration of lysosomal pathology were estimated in NIPS I and VII mice. Without immunomodulation, initial GUSB and IDUA activities in plasma reached >100-fold of wild-type (WT) levels but fell to background within 4 weeks post-injection. In immunomodulated transposon-treated MPS I mice plasma IDUA persisted for over 3 months at up to 100-fold WT activity in one-third of MPS I mice, which was sufficient to reverse lysosomal pathology in the liver and, partially, in distant organs. Histological and immunohistochemical examination of liver sections in IDUA transposon-treated WT mice revealed inflammation 10 days post-injection consisting predominantly of mononuclear cells some of which were CD4- or CD8- positive. Conclusions Our results demonstrate the feasibility of achieving prolonged expression of lysosomal enzymes in the liver and reversing MPS disease in adult mice with a single dose of therapeutic SB transposons. Copyright (C) 2007 John Wiley & Sons, Ltd.