Using human liver tissue equivalents to optimize AAV-mediated GT and better define age-related clinical risks

The overall goal is to utilize a human liver tissue equivalent (hLTE) platform to answer critical questions regarding AAV-mediated gene therapy for hemophilia A and to determine the impact recipient age has on these variables. The project will test the hypothesis that FVIII expression can be improved, pre-existing immunity to AAV overcome, and toxicity avoided by optimizing fVIII transgene codon usage/sequence or by targeting transduction to hepatic endothelium. Specifically, the work involves defining age-dependent impacts of AAV transduction vs. hepatocyte-targeted FVIII expression on human liver biology, innate immunity, and the potential for optimization to prevent immune responses. It will also test if targeting AAV transduction to hepatic endothelium improves FVIII expression, prevents inflammation, preserves liver function, and protects against anti-capsid immunity. Finally, it will investigate if genomic integration frequency is higher at younger ages and if targeting hepatic endothelial cells decreases genotoxicity.