Abstracts

Rationale: SCN1B encodes the ß1 subunit of the voltage gated sodium channel (VGSC), which is necessary for firing of action potentials. ß1 modulates VGSC α subunit gating kinetics, among other functions. Inherited homozygous loss-of-function variants in SCN1B result in a developmental and epileptic encephalopathy (DEE) that shares features with Dravet Syndrome. Scn1b-/- mouse models have demonstrated spontaneous seizures and increased peak amplitudes of evoked action potentials in CA3 hippocampal slice recordings. We aim to determine the effect of pathogenic SCN1B variants on excitability in human neurons to determine pathogenic mechanisms and develop precision therapy for these patients. Methods: A subject with SCN1B-related DEE was identified with a homozygous SCN1B p.R89C variant. Grown from a skin biopsy, patient fibroblasts were reprogrammed into induced pluripotent stem cells (iPSCs). Cortical-like excitatory neurons were derived from control and patient iPSCs using two methods: a dual-SMAD pathway inhibition protocol and induction of the transcription factors Neurogenin1/2 (NGN). Single cell electrophysiology was performed on the iPSC-derived neurons to determine sodium current density (I-Na) in voltage-clamp mode, and spontaneous and evoked action potentials were measured in current-clamp mode. Results: Several patient iPSC lines were generated, and two were chosen for use. Neurons derived using dual-SMAD inhibition expressed the deep-layer cortical marker CTIP2, whereas those derived from NGN induction expressed the upper-layer cortical marker Brn2. SCN1B R89C dual-SMAD neurons demonstrated a doubling of peak I-Na (n=9 control, n=13 R89C, p<0.05). SCN1B R89C NGN neurons also demonstrated increased peak I-Na (n=16 control, n=18 R89C, p<0.003). There was no significant change in the persistent I-Na. SCN1B R89C NGN neurons also had increased spontaneous firing vs. control (R89C 18.5% vs. control 9%, p<0.01), and an increase in the number of evoked action potentials (p<0.05). Conclusions: Human iPSC-derived neurons with homozygous SCN1B R89C variants display various features of hyperexcitability including increased peak I-Na, and increased spontaneous and evoked firing. This hyperexcitability likely results from dysfunctional modulation of VGSC α subunit gating kinetics and suggests that increased sodium current in excitatory neurons underlies the pathogenesis of SCN1B-related DEE. This implies that sodium channel blockers may be efficacious to treat seizures in at least a subset of SCN1B-related DEE. This is similar to SCN2A- and SCN8A-related epilepsies but differs from SCN1A-related Dravet Syndrome. Funding: Grant support is provided by NIH (NINDS) NS076752 (LLI and JMP).