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The IS2 Element Improves Transcription Efficiency of Integration-Deficient Lentiviral Vector Episomes
Molecular Therapy - Nucleic Acids ( IF 6.5 ) Pub Date : 2018-08-18 , DOI: 10.1016/j.omtn.2018.08.007
Sabina Sánchez-Hernández , Alejandra Gutierrez-Guerrero , Rocío Martín-Guerra , Marina Cortijo-Gutierrez , María Tristán-Manzano , Sandra Rodriguez-Perales , Laura Sanchez , Jose Luis Garcia-Perez , Jesus Chato-Astrain , Ricardo Fernandez-Valades , Ana Belén Carrillo-Galvez , Per Anderson , Rosa Montes , Pedro J. Real , Francisco Martin , Karim Benabdellah

Integration-defective lentiviral vectors (IDLVs) have become an important alternative tool for gene therapy applications and basic research. Unfortunately, IDLVs show lower transgene expression as compared to their integrating counterparts. In this study, we aimed to improve the expression levels of IDLVs by inserting the IS2 element, which harbors SARs and HS4 sequences, into their LTRs (SE-IS2-IDLVs). Contrary to our expectations, the presence of the IS2 element did not abrogate epigenetic silencing by histone deacetylases. In addition, the IS2 element reduced episome levels in IDLV-transduced cells. Interestingly, despite these negative effects, SE-IS2-IDLVs outperformed SE-IDLVs in terms of percentage and expression levels of the transgene in several cell lines, including neurons, neuronal progenitor cells, and induced pluripotent stem cells. We estimated that the IS2 element enhances the transcriptional activity of IDLV LTR circles 6- to 7-fold. The final effect the IS2 element in IDLVs will greatly depend on the target cell and the balance between the negative versus the positive effects of the IS2 element in each cell type. The better performance of SE-IS2-IDLVs was not due to improved stability or differences in the proportions of 1-LTR versus 2-LTR circles but probably to a re-positioning of IS2-episomes into transcriptionally active regions.



中文翻译:

IS2元素提高了整合缺陷型慢病毒载体附加体的转录效率

整合缺陷型慢病毒载体(IDLV)已成为基因治疗应用和基础研究的重要替代工具。不幸的是,与它们的整合对应物相比,IDLVs显示出较低的转基因表达。在这项研究中,我们旨在通过将带有SAR和HS4序列的IS2元件插入其LTR(SE-IS2-IDLV)中来提高IDLV的表达水平。与我们的预期相反,IS2元素的存在并未消除组蛋白脱乙酰基酶的表观遗传沉默。此外,IS2元素降低了IDLV转导细胞中的附加体水平。有趣的是,尽管存在这些负面影响,但SE-IS2-IDLV在包括神经元,神经元祖细胞和诱导性多能干细胞在内的几种细胞系中,转基因的百分比和表达水平都优于SE-IDLV。我们估计,IS2元素可将IDLV LTR环的转录活性提高6至7倍。IDLV中IS2元素的最终效果将在很大程度上取决于目标细胞以及每种细胞类型中IS2元素的负效应与正效应之间的平衡。SE-IS2-IDLVs更好的性能不是由于提高了稳定性或1-LTR与2-LTR环的比例不同,而是归因于IS2-异构体重新定位到转录活性区中。

更新日期:2018-08-18
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