Abstracts

Rationale: Patients with Early Infantile Epileptic Encephalopathy (EIEE) 52 (OMIM 617350) have inherited, homozygous variants in the gene SCN1B, encoding the voltage-gated sodium channel β1 and β1B non-pore-forming subunits. Inherited, heterozygous SCN1B variants have been linked to generalized epilepsy with febrile seizures plus, temporal lobe epilepsy, and cardiac arrhythmias. Homozygous SCN1B variants have been reported in seven epilepsy patients to date, with clinical descriptions suggestive of Dravet syndrome (DS). Scn1b null mice have a phenotype that is similar to DS. Thus, this small patient cohort was presumed to represent a subset of DS. Here, we describe a 4-year electroclinical follow-up of an eighth-reported SCN1B patient with a previously unreported variant, c.253C>T (β1-p.R85C), located in the extracellular Ig loop domain. Methods: To investigate the subcellular localization of β1-p.R85C, we performed cell surface biotinylation experiments, comparing β1-p.R85C to wildtype (WT) β1 protein. Experiments were performed in Chinese Hamster Lung fibroblasts that were stably transfected with eGFP, WT-β1, or β1-p.R85C. The WT-β1 and β1-p.R85C cDNA constructs each contained a C-terminal, in-frame, V5-epitope tag followed by a 2A endoproteolytic sequence and eGFP on the 3’ end of the construct for detection of equimolar expression of a fluorescence marker protein. Total protein and neutravidin-selected cell surface proteins were analyzed by western blot with anti-V5 antibody. Anti-HSP90 was used as an internal control to detect non-cell-surface proteins. To investigate the ability of β1-p.R85C to modulate INa, human Nav1.1 channels (hNav1.1), which were chosen due to their association with EIEE and DS, HEK-hNav1.1 cells were transiently transfected with WT-β1, β1-p.R85C or eGFP for voltage clamp analysis. Results: The female proband showed hypotonia from birth, multifocal myoclonias at 2.5 months, then focal seizures and myoclonic status epilepticus at 3 months, triggered by fever. Auditory Brainstem Response showed bilateral hearing loss. Epilepsy was refractory and the patient had virtually no development. Administration of fenfluramine resulted in a significant reduction in seizure frequency and resolution of SE episodes that persisted after a two-year follow-up. The electroclinical profile of the proband suggests a diagnosis of EIEE that is more severe than DS. Surface biotinylation and confocal immunofluorescence microscopy results showed that, similar to WT-β1, β1-p.R85C localizes to the cell surface. Electrophysiological analysis showed that WT-β1 subunits promote the cell surface expression of hNav1.1, but that the β1-p.R85C variant does not, in spite of normal cell surface expression of the protein. Conclusions: These data, showing loss-of-function of SCN1B-p.R85C, are similar to previous work with the SCN1B-p.C121W variant, also located in the Ig loop domain, showing normal cell surface expression, but loss-of-function in terms of INa modulation and for which a homozygous mouse model shows a phenotype that is similar to DS. Moreover, our results suggest reconsideration of the linkage of SCN1B variants to DS rather than to EIEE. Funding: NIH R37NS076752 to LLI