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2452 A Single Administration of AAV5-hFIX in Newborn, Juvenile and Adult Mice Leads to Stable hFIX Expression up to 18 Months after Dosing

Program: Oral and Poster Abstracts
Session: 801. Gene Editing, Therapy and Transfer: Poster II
Hematology Disease Topics & Pathways:
Hemophilia, Diseases, Bleeding and Clotting, Study Population
Sunday, December 6, 2020, 7:00 AM-3:30 PM

Lisa Spronk1*, Richard Logtenstein1*, Martin de Haan1*, Jaap Twisk2*, Valerie Ferreira1*, Sander van Deventer1* and Liesbeth Heijink1*

1uniQure Biopharma BV, Amsterdam, Netherlands
2uniQure biopharma B.V, Amsterdam, Netherlands

Recombinant adeno-associated viruses (rAAV) are replication deficient, non-integrating viruses commonly used as vectors for gene therapies. Currently an AAV serotype 5 vector with an hFIX transgene expression cassette designed for liver-directed expression of human Factor IX is being studied in clinical trials for hemophilia B. A major concern when using a single treatment with AAV is the possibility of losing transgene over time due to growth and/or natural turnover of cells. Here, we investigated the duration of hFIX expression in plasma after treating wildtype mice at different stages of development, from neonates to adults, with AAV5-hFIX. Since hemophilia B is predominantly found in male patients, male C57Bl/6 mice received a single intravenous infusion of 2x1014gc/kg AAV5-hFIX or vehicle. Neonates were dosed at 2 days old, weanlings at 3 weeks old, juveniles at 6 weeks of age and adult mice were dosed at 11 weeks or 6 months of age. All mice were sampled for plasma starting 4 weeks after dosing and then every three months. One cohort of mice (n=10 per group) was necropsied 4 weeks after dosing, while the other cohort (n=20 per group) was followed until 18 months of age, which is nearly the complete lifespan of a mouse. At necropsy the livers were weighed and collected for vector DNA analysis by QPCR. The plasma samples were used to determine the hFIX protein levels at each timepoint. All AAV5-hFIX injected animals showed hFIX protein expression from the first sampling timepoint at 4 weeks post dosing until sacrifice. The absolute levels of hFIX protein were lower in mice treated at a younger age, reflecting the lower total amount of vector genome copies injected in those animals due to the low body weight at dosing. The highest hFIX expression was seen 4 weeks after dosing and decreased to a stable level of hFIX protein within 3-6 months after dosing, after which the expression remained stable for the duration of the study. Remarkably, none of the animals in any group showed loss of hFIX expression after stabilization, even after more than a year of follow up and significant growth of the liver. A correlation was found between the amount of genome copies injected, the genome copies detected in the liver after necropsy and the hFIX protein expression in plasma at necropsy. This study shows that even in animals treated with AAV5-hFIX at a very young age, followed by significant growth, hFIX expression was sustained up to 18 months after dosing. This contradicts the expectation that vector DNA is lost during replication of the cells due to growth or natural turnover. Further studies to unravel the underlying mechanisms are ongoing.

Disclosures: Spronk: uniQure: Current Employment. Logtenstein: uniQure: Current Employment. de Haan: uniQure: Current Employment. Twisk: uniQure: Current Employment. Ferreira: uniQure: Current Employment. van Deventer: uniQure: Current Employment. Heijink: uniQure: Current Employment.

*signifies non-member of ASH