Abstract
The biliary system is routinely accessed for clinical purposes via endoscopic retrograde cholangiopancreatography (ERCP). We previously pioneered ERCP-mediated hydrodynamic injection in large animal models as an innovative gene delivery approach for monogenic liver diseases. However, the procedure poses potential safety concerns related mainly to liver or biliary tree injury. Here, we sought to further define biliary hydrodynamic injection parameters that are well-tolerated in a human-sized animal model. ERCP was performed in pigs, and hydrodynamic injection carried out using a novel protocol to reduce duct wall stress. Each pig was subjected to multiple repeated injections to expedite testing and judge tolerability. Different injection parameters (volume, flow rate) and injection port diameters were tested. Vital signs were monitored throughout the procedure, and liver enzyme panels were collected pre- and post-procedure. Pigs tolerated repeated biliary hydrodynamic injections with only occasional, mild, isolated elevation in aspartate aminotransferase (AST), which returned to normal levels within one day post-injection. All other liver tests remained unchanged. No upper limit of volume tolerance was reached, which suggests the biliary tree can readily transmit fluid into the vascular space. Flow rates up to 10 mL/sec were also tolerated with minimal disturbance to vital signs and no anatomic rupture of bile ducts. Measured intrabiliary pressure was up to 150 mmHg, and fluid-filled vesicles were induced in liver histology at high flow rates, mimicking the changes in histology observed in mouse liver after hydrodynamic tail vein injection. Overall, our investigations in a human-sized pig liver using standard clinical equipment suggest that ERCP-guided hydrodynamic injection will be safely tolerated in patients. Future investigations will interrogate if higher flow rates and pressure mediate higher DNA delivery efficiencies.
Highlights
Hydrodynamic injection represents a promising method of non-viral gene therapy
We showed the feasibility of using endoscopic retrograde cholangio-pancreatography (ERCP) to deliver genes via hydrodynamic injection into the livers of pigs [24]
Due to the potential trauma to liver tissue and bile ducts, we further investigated the safety and limitations of endoscopic retrograde cholangiopancreatography (ERCP)-mediated hydrodynamic injection, which is crucial for translation into humans
Summary
The technique consists of briefly exposing tissues to increased pressure and/or fluid volume levels, transiently causing disruptions in cell membranes, thereby mediating delivery of DNA inside the cell [1]. Hydrodynamic injection was first pioneered for liver gene delivery, but has subsequently been shown to mediate delivery in the skin, muscle, and kidney [2,3,4]. Beyond the delivery of DNA inside the cell, hydrodynamic injection has been shown to mediate delivery of siRNA and proteins inside hepatocytes [5]. Hydrodynamic tail vein injection (HTVI) into mouse liver involves administration of 10% body fluid volume DNA solution in 4–7 seconds, yielding expression of delivered DNA throughout the liver [6, 7]. Once inside the liver cells, DNA is stable due to slowly dividing hepatocytes, it can be slowly epigenetically silenced over time [15, 16]
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