Abstract
Missense variants in the SCN8A voltage-gated sodium channel gene are linked to early-infantile epileptic encephalopathy type 13, also known as SCN8A-related epilepsy. These patients exhibit a wide spectrum of intractable seizure types, severe developmental delay, movement disorders, and elevated risk of sudden unexpected death in epilepsy. The mechanisms by which SCN8A variants lead to epilepsy are poorly understood, although heterologous expression systems and mouse models have demonstrated altered sodium current properties. To investigate these mechanisms using a patient-specific model, we generated induced pluripotent stem cells from three patients with missense variants in SCN8A: p.R1872>L (Patient 1); p.V1592>L (Patient 2); and p.N1759>S (Patient 3). Using small molecule differentiation into excitatory neurons, induced pluripotent stem cell-derived neurons from all three patients displayed altered sodium currents. Patients 1 and 2 had elevated persistent current, while Patient 3 had increased resurgent current compared to controls. Neurons from all three patients displayed shorter axon initial segment lengths compared to controls. Further analyses focused on one of the patients with increased persistent sodium current (Patient 1) and the patient with increased resurgent current (Patient 3). Excitatory cortical neurons from both patients had prolonged action potential repolarization. Using doxycycline-inducible expression of the neuronal transcription factors neurogenin 1 and 2 to synchronize differentiation of induced excitatory cortical-like neurons, we investigated network activity and response to pharmacotherapies. Both small molecule differentiated and induced patient neurons displayed similar abnormalities in action potential repolarization. Patient induced neurons showed increased burstiness that was sensitive to phenytoin, currently a standard treatment for SCN8A-related epilepsy patients, or riluzole, an FDA-approved drug used in amyotrophic lateral sclerosis and known to block persistent and resurgent sodium currents, at pharmacologically relevant concentrations. Patch-clamp recordings showed that riluzole suppressed spontaneous firing and increased the action potential firing threshold of patient-derived neurons to more depolarized potentials. Two of the patients in this study were prescribed riluzole off-label. Patient 1 had a 50% reduction in seizure frequency. Patient 3 experienced an immediate and dramatic seizure reduction with months of seizure freedom. An additional patient with a SCN8A variant in domain IV of Nav1.6 (p.V1757>I) had a dramatic reduction in seizure frequency for several months after starting riluzole treatment, but then seizures recurred. Our results indicate that patient-specific neurons are useful for modelling SCN8A-related epilepsy and demonstrate SCN8A variant-specific mechanisms. Moreover, these findings suggest that patient-specific neuronal disease modelling offers a useful platform for discovering precision epilepsy therapies.