Optimizing lingual directed adeno-associated virus (AAV) therapy in a<i>Gaa^-/-</i>rat model of Pompe disease

Pompe disease is a recessively inherited glycogen storage disease caused by a deficiency in the lysosomal enzyme acid α-glucosidase (GAA). Excessive glycogen accumulation in skeletal muscle and motoneurons leads to motor unit dysfunction and weakness. Humans with Pompe disease commonly develop macroglossia and severe lingual motor problems. Dysphagia with secondary aspiration, pneumonias, and obstructive sleep apnea are common. The only approved treatment for Pompe disease is human recombinant GAA enzyme replacement therapy (ERT) which has a short half-life and does not cross the blood brain barrier. The ability of AAV to drive gene expression throughout the body makes it an attractive alternative to ERT. We have previously shown that AAV serotype 9 (AAV9) encoding GAA can effectively drive GAA expression in tongue myofibers and hypoglossal (XII) motoneurons following a single lingual injection in the Gaa -/- mouse model of Pompe disease. In preliminary experiments we extended this observation to a larger murine model, the Gaa -/- rat. The anterior and posterior portions of the tongue in 4-month-old female Gaa -/- rats (n=3) were injected with 100µL rAAV9-Des-hGAA (1.55x10 12 vector genomes (vg)) split into 50μL delivered to both the anterior and posterior portions of the tongue. Tissues were harvested 2-months later and GAA expression was immunohistochemically verified in XII motoneurons. However, relatively few XII motoneurons were found to express GAA. In ongoing studies, we are testing strategies to optimize the AAV delivery approach to increase XII motoneuron GAA expression. 6-mo old female Gaa -/- rats have received a lingual injection with an increased dose of AAV9-Des-hGAA (100μL, 1x10 13 vg) delivered identically with 50 µL delivered to both the anterior and poster portions of the tongue. An additional group of rats will receive an immunosuppression regimen using an anti-CD20 antibody delivered twice in two week increments prior to lingual AAV treatment and then again 2-weeks post injection. We hypothesize that the higher AAV dose coupled with immunomodulation will enable a greater portion of the XII motor nucleus to express the transgene. Based on our initial results, we conclude that a single lingual injection of AAV9-Des-hGAA can drive GAA expression in XII motoneurons in the Gaa -/- rat model of Pompe disease but optimization of this method is needed to improve efficacy to enable clinical translation. R01HD052682 (DDF and BJB), T32HG008958 (ESB), K99NS133388-01A1 (SR) This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.