Abstracts

Rationale: DEPDC5 is now recognized as one of the genes most often implicated in familial/inherited focal epilepsy and brain malformations. Individuals with pathogenic variants in DEPDC5 are at risk for epilepsy, associated neuropsychiatric comorbidities, and Sudden Unexplained Death in Epilepsy (SUDEP). Depdc5flox/flox-Syn1Cre (Depdc5cc+) neuronal-specific Depdc5 knockout mice exhibit terminal seizures and neuronal mTORC1 hyperactivation. It is not known if these mice exhibit behavioral deficits or whether mTOR inhibitors could rescue the phenotypes associated with DEPDC5-related conditions. Methods: Male and female Depdc5cc+ and littermate control mice were treated with the mTORC1 inhibitor rapamycin (6 mg/kg) or vehicle every other starting at one month of age (n > 6 animal per group). Animals were subjected to an open field paradigm and elevated plus-maze as measures of hyperactivity and anxiety between 75-90 days of age. Animals were followed until 50% of the vehicle-treated Depdc5cc+ died; the litters with remaining vehicle-treated Depdc5cc+ mice were sacrificed for pathology and biochemical experiments. Cortical lysates were analyzed by western blot and anatomically matched cortical sections were analyzed by immunofluorescence across each genotype and treatment group. Results: Rapamycin did not significantly affect the weight of rapamycin treated Depdc5cc+ mice. Adult Depdc5cc+ mice (n = 8) exhibited increased locomotor activity throughout the testing period compared to littermate controls (n = 13) (p < 0.0001). Open-field activity was reduced by 40% in rapamycin-treated Depdc5cc+ mice compared to vehicle-treated Depdc5cc+ mice (p=0.004; two-way ANOVA with Tukey’s post hoc test). Activity of rapamycin-treated Depdc5cc+ mice was reduced but not to the level of vehicle-treated controls (p=0.023; two-way ANOVA with Tukey’s post hoc test). Half of the vehicle-treated Depdc5cc+ mice spontaneously died with a median age of death of 126 days (range 93 – 128). In contrast, only one (11.1%) rapamycin-treated Depdc5cc+ mouse died spontaneously at 163 days of age without obvious signs of morbidity prior to death. Vehicle treated Depdc5cc+ cortical lysates and brain sections had evidence of hyperactive mTORC1 signaling as measured by the increased phosphorylation of downstream ribosomal S6 (p-S6 on Ser240/244). Importantly, rapamycin treatment of Depdc5cc+ mice reduced p-S6 levels in cortical lysates back to vehicle-treated control levels. Conclusions: Our data provide the first evidence of behavioral alterations in mice with Depdc5 loss. Neuronal-specific Depdc5 knockout mice die early due to terminal seizures. Rapamycin decreased mTOR hyperactivity, reduces behavioral hyperactivity, and prolongs the life of neuronal-specific Depdc5 knockout mice. Taken together, our data provide support for mTOR inhibitors as rational therapeutics for DEPDC5-related epilepsy in humans. Funding: NIH 2R25NS070682-07, NINDS K12NS098482, and Boston Children's Hospital Basic/Translational Research Executive Committee Career Development Fellowship.