Background and Aims

A simple, safe, targeted, and efficient in vivo DNA delivery system is necessary for clinical-grade liver-targeted gene therapy in humans. Intravascular hydrodynamic gene delivery has been investigated in large animal models, but translation to humans has been hampered by its technical challenges, invasiveness, and potential for significant cardiovascular adverse events. We posited that intrabiliary delivery of DNA plasmids via ERCP-guided hydrodynamic injection could overcome these obstacles.

Methods

Twelve pigs (40-50 kg) were divided into 3 groups (4 per group) and survived 21, 30, or 60 days. ERCP was performed by inflating a balloon catheter in the common hepatic duct and creating a closed space between it and the liver parenchyma. Last, a solution composed of plasmid/sleeping beauty (SB) mix was injected under pressure through the catheter into the closed space. Swine were killed at the 3 different time points and liver tissue harvested. Plasmid DNA expression and functional translated protein expression were assessed.

Results

ERCP-guided hydrodynamic delivery of naked plasmid DNA facilitated by pCytomegalovirus-Sleep Beauty (pCMV-SB) transposons was technically feasible and devoid of cardiovascular and local adverse events in all 12 pigs. Furthermore, plasmid DNA (both single and combination) was successfully transferred into swine hepatocytes in all 12 pigs. Additionally, stable integration of the DNA constructs in hepatocyte genomic DNA was reliably noted at all 3 time points. In the 4 swine that were kept alive to 60 days, successful genomic integration and subsequent protein expression was observed in the targeted liver tissue.

Conclusions

ERCP-guided hydrodynamic delivery of gene therapy may usher in the next chapter in gene therapy with the potential to impact a variety of single-gene, complex genetic, and epigenetic liver diseases. It also raises the possibility that other nucleic acid therapeutics (microRNA, lncRNA, siRNA, shRNA) could similarly be delivered.

中文翻译：

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通过ERCP引导的流体动力注射成功地完成了肝定向基因的输送（带视频）

背景和目标

一个简单，安全，有针对性和高效的体内DNA递送系统对于人类临床级肝靶向基因治疗是必需的。已经在大型动物模型中研究了血管内流体动力学基因的传递，但是由于其技术挑战，侵袭性和潜在的重大心血管不良事件，阻碍了向人类的翻译。我们假定通过ERCP引导的流体动力注射胆汁内传递DNA质粒可以克服这些障碍。

方法

将12头猪（40-50公斤）分为3组（每组4只），存活21、30或60天。ERCP通过在肝总管中向球囊导管充气并在其与肝实质之间形成一个封闭空间来进行。最后，在压力下通过导管将由质粒/睡美人（SB）混合物组成的溶液注入封闭空间。在3个不同的时间点杀死猪并收获肝组织。评估质粒DNA表​​达和功能翻译蛋白表达。

结果

ERCP指导的由pCytomegalovirus-Sleep Beauty（pCMV-SB）转座子促进的裸质粒DNA的流体动力学递送在技术上是可行的，并且在所有12头猪中都没有心血管和局部不良事件。此外，质粒DNA（单个和组合）已成功转移到所有12头猪的猪肝细胞中。另外，在所有3个时间点可靠地注意到DNA构建体在肝细胞基因组DNA中的稳定整合。在存活60天的4头猪中，在目标肝组织中观察到成功的基因组整合和随后的蛋白表达。

结论

ERCP指导的基因治疗的水动力传递可能会开启基因治疗的新篇章，并有可能影响多种单基因，复杂遗传和表观遗传性肝病。这也增加了其他核酸治疗剂（microRNA，lncRNA，siRNA，shRNA）可以类似地递送的可能性。