150. Liver Directed Gene Therapy Using Non-Viral Minicircle-DNA Vector for Ornithine Transcarbamylase Deficiency (OTCD)

As an alternative to (AAV-) viral vector-transduction, we are using minicircle (MC) naked-DNA vectors devoid of any viral or bacterial sequences for delivery into the ornithine transcarbamylase (OTC) deficient murine model, Spfash. The MC-vectors which do not have a defined size limitation contained expression cassettes with a short (0.3 kb) synthetic liver-specific promoter or the natural (endogenous) Otc promoter with a length of up to 2 kb, the murine Otc-cDNA (mOtc) or the codon-optimized murine Otc (mcoOtc) with or without truncated 5’-intron, and a polyA signal. MC vectors were delivered to mouse liver by hydrodynamic tail vein injection. Upon sacrificing mice 2-3 weeks after vector infusion, transgenic OTC enzyme activity was found in liver extracts to be up to 2 or 3-fold higher than in normal wild-type controls. However, similar to what we have observed in our study to correct hyperphenylalaninemia in the phenylketonuria (PKU) mouse using hydrodynamic injection of MC vectors (Viecelli et al., Hepatology 2014), gene expression was mainly found at the perivenous area also in the OTC-treated mouse liver. Since the full active urea cycle is restricted to the periportal area and thus delivery of MC vectors to treat OTC deficiency needs to target these periportal hepatocytes, we have established delivery via hydrodynamic portal vein injection for gene transfer into Spfash mice. Preliminary analysis showed transgene expression in MC-treated mice however at a much lower level than via hydrodynamic tail vein injection. Analysis of urinary orotic acid and in vivo ureagenesis is ongoing for the evaluation of the therapeutic efficacy.