Increased persistence of lung gene expression using plasmids containing the ubiquitin C or elongation factor 1α promoter
Increased persistence of lung gene expression using plasmids containing the ubiquitin C or elongation factor 1α promoter
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DOI：
10.1038/sj.gt.3301561
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发表时间：
2001-10-01
期刊：
影响因子：
5.1
通讯作者：
Hyde, SC
中科院分区：
文献类型：
--
作者：
Gill, DR;Smyth, SE;Hyde, SC
对于慢性疾病的有效基因治疗，需要治疗水平的持续转基因表达。囊性纤维化（CF）患者气道基因转移的临床研究导致了短暂的转基因表达。我们使用裸质粒DNA鼻内给药到小鼠肺作为模型，用于研究从一系列报告基因表达质粒的气道基因转移的持续时间。当由病毒启动子CMV、RSV和SV 40介导时，转基因表达是瞬时的，在2周后下降到低于峰值表达的10%，尽管腺病毒E4 ORF3基因的顺式存在导致来自CMV启动子的报道活性的持续时间延长。通过定量TaqMan PCR分析确定，这些启动子的瞬时表达不是由于载体的丢失。然而，使用来自人多聚泛素C（UbC）和延伸因子1 α（EF1 α）基因的启动子导致小鼠肺中持续的基因表达。UbC启动子指导高水平的报告活性，其维持长达8周，并且在单次给药后6个月仍可检测到。这种来自非病毒载体的持续气道转基因表达而不伴随潜在抗原的表达以前尚未报道。因此，尽管载体DNA在体内持续存在，但启动子功能的减弱可能导致转基因表达的沉默，并且建议谨慎选择启动子序列用于体内基因转移。
For effective gene therapy of chronic disease, persistent transgene expression at therapeutic levels is required. Clinical studies of airway gene transfer in patients with cystic fibrosis (CF) have resulted in short-lived transgene expression. We used intra-nasal dosing of naked plasmid DNA to the murine lung as a model for investigating the duration of airway gene transfer from a series of reporter expression plasmids. Transgene expression was transient when mediated by the viral promoters CMV, RSV and SV40, falling to less than 10% of peak expression after 2 weeks, although the presence of the adenoviral E4ORF3 gene in cis, resulted in extended duration of reporter activity from the CMV promoter. Transient expression from these promoters was not due to loss of the vector as determined by quantitative TaqMan PCR analysis. However, use of the promoters from the human polybiquitin C (UbC) and the elongation factor 1 alpha (EF1 alpha) genes resulted in persistent gene expression in the mouse lung. The UbC promoter directed high-level reporter activity which was maintained for up to 8 weeks and was still detectable 6 months after a single administration. Such persistent airway transgene expression from a nonviral vector without the concomitant expression of a potential antigen has not been reported previously. Thus, despite the persistence of vector DNA in vivo, attenuation of promoter function may lead to silencing of transgene expression and careful selection of promoter sequences is recommended for in vivo gene transfer.