Effect of pharmacological and genetic therapies on epilepsy in a TSC2 mouse model
Abstract number :
2.218
Submission category :
3. Neurophysiology / 3F. Animal Studies
Year :
2025
Submission ID :
193
Source :
www.aesnet.org
Presentation date :
12/7/2025 12:00:00 AM
Published date :
Authors :
Presenting Author: Luis Martinez, PhD – Baylor College of Medicine
Wai Lee, PhD – Baylor College of Medicine
Anne Anderson, MD – Baylor College of Medicine/Texas Children's Hospital
Rationale: Tuberous Sclerosis Complex (TSC) is a neurological disorder caused by mutations in TSC1 or TSC2 genes and characterized by brain malformations and drug resistant epilepsy. mTOR inhibitors can reduce seizures, but are not disease modifying. Gene therapy may be effective in TSC-associated epilepsy. To evaluate this, we utilized the NEX-Tsc2 knockout (KO of the Tsc2 gene in cortical excitatory neurons) mice exhibiting robust epilepsy, which were treated with an adeno-associated viral (AAV) vector carrying a Tsc2 gene construct (cTsc2).
Methods: To determine the expression of the AAV9-cTsc2 gene early neonatally, WT mice were treated by intracerebroventricular (ICV) injection with AAV9-cTsc2 (1.1x10^12) or empty vector at P0-P3 followed by collection of cortical tissue at various timepoints and assessed using western blotting. NEX-Tsc2 KO pups were then treated with AAV-cTsc2 or empty vector at P0-P3 or P21, early or late treatment groups, respectively. NEX-Tsc2 KO pups exhibit early mortality and we have previously shown that mTOR treatment extends the lifespan transiently. Therefore, a cohort of mice also were treated with the mTOR inhibitor RAD001 (6mg/kg every other day, intraperitoneally) to extend longevity during the AAV expression window. For late AAV treatment, RAD001 was discontinued at P35. EEG electrodes were implanted at P10 or P25 in the early vs. late groups, respectively. Analysis of epileptiform and seizure activity was recorded with video EEG.
Results: Cortical tissue from WT mice treated with AAV-cTsc2 exhibited a significant increase in cTsc2 protein compared to controls given empty vector as early as two days following injection (p< 0.001). At 10 days post-injection, levels of cTsc2 protein were significantly increased (p< 0.001) and remained elevated at 50 days post-treatment (p< 0.05) compared to 2 days post-treatment. Cortical tissue from AAV-cTsc2 treated KO mice exhibited significantly increased levels of the cTsc2 protein 20 days after AAV treatment; studies to evaluate phospho-S6 levels are pending. Survival was not improved in KO mice treated early with AAV-cTsc2 alone (p >0.05) but was significantly extended with AAV-cTsc2 plus RAD001 compared to vehicle KO mice (p< 0.05). Survival of KO mice treated late with AAV-cTsc2 was significantly prolonged compared to mice treated only with RAD001 from P50 to P100 (p< 0.05). Treatment with RAD001 significantly delayed the development of seizures from P12 to P50, whereas seizures in KO mice treated late with AAV-cTsc2 had delayed seizure onset to P100.
Conclusions: Our results in NEX-Tsc2 KO mice indicate that AAV-cTsc2 exhibits high expression levels of cTsc2 protein but in our pilot studies no significant decrease in mTOR activity (pS6) in the cortex. Both AAV-cTsc2 and RAD001 suppressed epileptiform activity and delayed seizure onset but did not render sustained disease modification. Additional studies are necessary to elucidate mechanisms of treatment resistance of the epilepsy phenotype in this model with potential translational relevance.
Funding: TSC Alliance Research grant
Department of Defense TSC Research Program grant
Neurophysiology