Single-Stranded γPNAs for In Vivo Site-Specific Genome Editing via Watson-Crick Recognition
Single-Stranded γPNAs for In Vivo Site-Specific Genome Editing via Watson-Crick Recognition
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DOI：
10.2174/1566523214666140825154158
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发表时间：
2014-01-01
影响因子：
3.6
通讯作者：
Glazer, Peter M.
中科院分区：
文献类型：
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作者：
Bahal, Raman;Quijano, Elias;Glazer, Peter M.
三链形成肽核酸(PNA)通过特定部位形成改变的螺旋结构来刺激供体DNA在基因组DNA中的重组，从而促进基因编辑，从而进一步刺激DNA修复。然而，为形成三链而设计的PNA的序列仅限于同型嘌呤位点。在这里，我们描述了一种新的策略，在这种策略中，包含在Gamma位置上的mini PEG替换的下一代单链Gamma PNA(Gamma PNA)可以在混合序列位置靶向小鼠骨髓中的基因组DNA，从而诱导靶向基因编辑。除了增强结合，伽马PNA还增加了溶解度，并改进了聚乳酸-乙醇酸(PLGA)纳米颗粒的配方，以实现高效的细胞内递送。单链伽马PNA在体外治疗的小鼠骨髓细胞和静脉注射的体内治疗的小鼠骨髓细胞中以0.8%的频率诱导靶向基因编辑，而没有检测到毒性。这些结果表明，Gamma PNA可能为基于Watson-Crick识别的无序列限制的诱导基因编辑提供一种新的工具。
Triplex-forming peptide nucleic acids (PNAs) facilitate gene editing by stimulating recombination of donor DNAs within genomic DNA via site-specific formation of altered helical structures that further stimulate DNA repair. However, PNAs designed for triplex formation are sequence restricted to homopurine sites. Herein we describe a novel strategy where next generation single-stranded gamma PNAs (gamma PNAs) containing miniPEG substitutions at the gamma position can target genomic DNA in mouse bone marrow at mixed-sequence sites to induce targeted gene editing. In addition to enhanced binding, gamma PNAs confer increased solubility and improved formulation into poly(lactic-co-glycolic acid) (PLGA) nanoparticles for efficient intracellular delivery. Single-stranded gamma PNAs induce targeted gene editing at frequencies of 0.8% in mouse bone marrow cells treated ex vivo and 0.1% in vivo via IV injection, without detectable toxicity. These results suggest that gamma PNAs may provide a new tool for induced gene editing based on Watson-Crick recognition without sequence restriction.