Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
J Gene Med. 2020 Jan;22(1):e3138. doi: 10.1002/jgm.3138. Epub 2020 Jan 2.
Durable transgene expression from plasmid DNAs is the key to gene therapy with non-viral vectors. A comparison of the durability of transgene expression from plasmid DNAs with the CpG-free and -containing backbones is important.
We constructed plasmid DNAs with the CpG-containing backbone, various transcription regulatory sequences with and without CpG, and the gene encoding Gaussia princeps luciferase, which is apparently non-immunogenic. The tail vein hydrodynamics-based method was used for plasmid injection into mice, and the luciferase activity in serum was tracked for 28 days.
The plasmid DNAs containing the albumin promoter [with or without the cytomegalovirus (CMV) enhancer] and the elongation factor (EF)1α promoter plus the CMV enhancer exhibited long-term luciferase expression. The expression from the plasmid DNA containing the albumin promoter without the CMV enhancer was maintained for at least 24 weeks and was similar to that from the corresponding CpG-free plasmid DNA.
The results obtained in the present study suggest that special sequences/systems are unnecessary for durable transgene expression from plasmid DNAs when the proper transcription regulatory sequences are used.
从质粒 DNA 中获得持久的转基因表达是使用非病毒载体进行基因治疗的关键。比较无 CpG 和含 CpG 骨架的质粒 DNA 中转基因表达的持久性非常重要。
我们构建了含有 CpG 骨架的质粒 DNA，以及带有和不带有 CpG 的各种转录调控序列，和编码 Gaussia princeps 荧光素酶的基因，该基因显然是非免疫原性的。我们使用基于尾静脉水动力的方法将质粒 DNA 注射到小鼠体内，并跟踪血清中的荧光素酶活性 28 天。
含有白蛋白启动子（带有或不带有巨细胞病毒（CMV）增强子）和延伸因子（EF）1α 启动子加 CMV 增强子的质粒 DNA 表现出长期的荧光素酶表达。含有白蛋白启动子而不含 CMV 增强子的质粒 DNA 的表达至少维持了 24 周，与相应的不含 CpG 的质粒 DNA 相似。
本研究的结果表明，当使用适当的转录调控序列时，从质粒 DNA 中获得持久的转基因表达并不需要特殊的序列/系统。
