Loss of transgene expression limits liver gene therapy in primates

Efforts to improve liver gene therapy have focused on next-generation adeno-associated virus (AAV) vector capsids, transgene delivery, and immunomodulating drugs, such as corticosteroids, to avoid destructive T-cell responses. We conducted a detailed characterization of AAV transduction in nonhuman primate liver across multiple capsids and transgenes to better define interactions that may limit stable and efficient transgene expression. We show that the initial transduction of hepatocytes is high, but the transduction rapidly diminishes to a lower stable baseline of <1% of cells, even though ~10% of the cells retain vector DNA that is localized within the nucleus. Further characterization showed genomic vector integration at frequencies of 1/100 to 1/1,000 genomes, suggesting that one mechanism for stable expression may occur via genome integration. These studies suggest that strategies to enhance durable transgene expression by activating retained nuclear episomes or by increasing the frequency of vector integrations may improve liver directed AAV gene therapy. ### Competing Interest Statement JMW is a paid advisor to and holds equity in iECURE, Scout Bio, Passage Bio, and the Center for Breakthrough Medicines (CBM). He also holds equity in the G2 Bio-associated asset companies. He has sponsored research agreements with Amicus Therapeutics, Biogen, CBM, Elaaj Bio, FA212, G2 Bio, G2 Bio-associated asset companies, iECURE, Janssen, Passage Bio, and Scout Bio, which are licensees of University of Pennsylvania technology. JAG, CB, LW, and JMW are inventors on patents that have been licensed to various biopharmaceutical companies and for which they may receive payments.