Targeting the Apoa1 locus for liver-directed gene therapy

Abstract Clinical application of somatic genome editing requires therapeutics that are generalizable to a broad range of patients. Targeted insertion of promoterless transgenes can ensure that edits are permanent and broadly applicable while minimizing risks of off-target integration. In the liver, the Albumin locus is currently the only well characterized site for promoterless transgene insertion. Here, we target the Apoa1 locus with Adeno-Associated Viral (AAV) delivery of CRISPR/Cas9, and achieve rates of 6 to 16% of targeted hepatocytes, with no evidence of toxicity. We further show that the endogenous Apoa1 promoter can drive robust and sustained expression of therapeutic proteins, such as apolipoprotein E (APOE), dramatically reducing plasma lipids in a model of hypercholesterolemia. Finally, we demonstrate that Apoa1-targeted fumarylacetoacetate hydrolase (FAH) can correct and rescue the severe metabolic liver disease hereditary tyrosinemia type I. In summary, we identify and validate Apoa1 as novel integration site that supports durable transgene expression in the liver for gene therapy applications.