Abstracts

This abstract is a recipient of the Young Investigator Award
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Rationale: Succinic semialdehyde dehydrogenase deficiency (SSADHD) is a rare epilepsy disorder caused by aldehyde dehydrogenase 5 family member A1 (aldh5a1) mutations. Aldh5a1 encodes SSADH which is essential for degrading GABA. In SSADHD, pathologic accumulation of GABA and its neuroactive degradation by-product g-hydroxybutyrate (GHB) in the developing brain results in global encephalopathy characterized by intellectual disability, speech delay, ataxia, intractable seizures and a significant risk of sudden unexpected death in epilepsy (SUDEP). Gene restoration therapy is a potential cure for SSADHD but is currently unavailable. To investigate the optimal parameters for safe and therapeutically effective aldh5a1-tagteted gene therapy, we developed an inducible SSADH mouse model, aldh5a1^lox-STOP, allowing on-demand, conditional Cre-mediated aldh5a1 reactivation.

Methods: We used a blood-brain barrier (BBB)-penetrating adeno-associated virus encompassing a constitutive promoter driving Cre expressions (AAV-PHP.B-CAG-Cre) for global aldh5a1 restoration in aldh5a1^lox-STOP mice. AAV was administered into homozygous (HOM) mice via intraperitoneal injection (5x10¹¹ genome copies/100ml saline) at a symptomatic stage (postnatal day 15-20) for brain-wide Cre-dependent aldh5a1 rescue. In addition, we crossed aldh5a1^lox-STOP mice with mice expressing Cre driven by the parvalbumin promoter (PV^Cre+). This allows PV cell-specific aldh5a1 rescue from an early postnatal stage. For both rescue strategies, phenotypes ranging from survivability, body weight gain, motor activity and SSADH protein expression were investigated between rescued mice and wild-type (WT) controls.

Results: AAV-Cre-mediated rescue led to nearly 50% survival at P60 (n=30), compared to 62% in PV^Cre+-rescued mice (n=24) and 0% survival in non-rescued mice (n=21). Surviving mice expressed 68.6±3.7% and 6.3±1.7% of WT SSADH protein level upon AAV-mediated (n=11) and PV^Cre+-mediated rescue (n=8), respectively. Notably, the amount of SSADH expression in PV^Cre+-mediated rescued mice correlated with their age of death. AAV-rescued mice, but not PV^Cre+-rescued mice, have their body weight returned to WT level at P60 (WT: 21.7±0.8g, n=12; HOM+AAV-Cre: 22.5±1g, n=11; HOM/PV^Cre+: 15.2±1.5g, n=7). Motor activity and gait behavior captured by a high-definition ventral camera revealed hyperactivity in HOM mice rescuable by AAV-Cre but not PV^Cre+ (distance traveled in 20 min: WT: 495±46m, n=6; HOM: 848±16m, n=5; HOM+AAV-Cre: 463±60m, n=6; HOM/PV^Cre+: 1364±17m, n=6).

Conclusions: Obligatory, pre-mature death characterized by non-recoverable seizures in aldh5a1^lox-STOP mice is amendable via brain-wide or PV cell-specific aldh5a1 restoration. AAV-mediated aldh5a1 restoration approach leads to appreciable symptom reversal. Cell type-specific gene delivery might avoid the potential oncogenic risk of aberrant aldh5a1 expression, but expressions in multiple relevant cell types are likely necessary for appreciable therapeutic effects.

Funding: SSADH Association Research Program Grant, NIH R21