267. Image-Guided, Liver-Targeted Hydrodynamic Gene Delivery in Dogs – Preclinical Assessment of Effectiveness and Safety of the Procedure

Abstract

Image-guided, liver-targeted hydrodynamic gene delivery was evaluated in dogs to assess the safety and effectiveness of the procedure for human clinical trial. The procedure involves image-guided catheter insertion to hepatic veins in the dogs followed by computer-controlled hydrodynamic gene delivery using newly developed injector for clinical use. Optimum parameters for the procedure, including the location of the balloon catheter in the hepatic vein, intravascular pressure upon injection, injection volume, and sequential injections to multiple lobes of the injected liver, were employed as previously we reported. Hydrodynamic injection of human factor IX expressing plasmid (pBS-hFIX) was performed in 7 dogs (female, 20 kg) and short- and long-term effects of the procedure were assessed. We demonstrate that plasma level of human factor IX and coagulation activity in dogs increased from the background level at ~20 ng/ml and ~8% to 21768.1 ng/ml and 36.8%, respectively in 1 week after hydrodynamic gene delivery and sustained for 4 weeks. Transgene expression in the hepatocytes was also confirmed by immunohistochemical stainings. For safety assessment, we demonstrated that the impacts of image-guided liver-targeted hydrodynamic gene delivery are localized in the site of injection in the liver. Histological analysis showed significant expansion of sinusoids at the injected site to 190% of their original size. Hepatic microcirculation analysis of liver-targeted hydrodynamic injection using reflectance spectrophotometry showed significant decrease of the oxygen saturation of the Hb in the blood of the injected lobe from 30 to 0 unit upon the injection and recovered smoothly after the injection. Serum analysis showed a transient, 10- to 20-fold increase in hepatobiliary enzymes, including aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase after hydrodynamic injections, which recovered within 4 days. We also explored for the first time, the levels of the cytokines following the hydrodynamic injections to the large animals. There was no increase in the systemic inflammatory cytokines of IFN-γ, IL-8, IL-18, and IL-4, although an increase in serum levels of TNF-α, IL-10, MCP-1, canine KC, and IL-6, which were related to vascular stretching representing the sinusoidal expansion was observed. No impacts on their respiratory, cardiovascular conditions, or long-term body weight changes were observed throughout the study. These results of preclinical assessments support the clinical applications of image-guided, liver-targeted hydrodynamic gene delivery.This work was supported in part by NIH grants RO1EB002946 and RO1HL075542, and by Japanese Society for the Promotion of Science Grants 22890064, 23790595, and 26860354.