Abstracts

Rationale: Genetic variants in KCNT1 are associated with a spectrum of epilepsy, including epilepsy of infancy with migrating focal seizures (EIMFS) and sleep-related hypermotor epilepsy (SHE). To date, over 30 distinct pathogenic missense variants in KCNT1 have been reported in the literature. The majority of variants are hypothesized to exert a pathologic gain-of-function mechanism, increasing KCNT1 (Slack, KNa1.1) channel activity by multiple mechanisms.

Methods: In order to identify novel, potentially pathogenic variants, we analyzed genetic data from pediatric epilepsy patients screened via Invitae’s Behind the Seizure® Epilepsy Panel (https://www.invitae.com/en/behindtheseizure/). KCNT1 variants identified via the Invitae panel were subsequently expressed in xenopus oocytes; KCNT1 current-voltage relationships were then quantified with two-electrode voltage clamp recordings using the automated Robocyte system.

Results: We identified 88 unique KCNT1 variants via the Invitae panel, including 12 patients with six previously reported pathogenic mutations, three patients harboring a novel, likely pathogenic, unreported variant, and 93 patients carrying 81 novel variants of unknown significance (VUS). We additionally identified 17 reported epilepsy-associated KCNT1 variants for which in-vitro validation of a gain-of-function mechanism was not available. To validate a screening pipeline for uncharacterized, potentially pathogenic mutations, we compared the biophysical properties of novel KCNT1 variants with those of known, previously characterized pathogenic mutations. Relative to wild type KCNT1, we found increased currents at +70 mV for previously characterized EIMFS variants, while a common KCNT1 missense single nucleotide polymorphism minimally impacted current amplitude. Among previously uncharacterized KCNT1 mutations, we identified 9 variants with gain-of-function increases in current amplitude.

Conclusions: Our dataset is the largest systematic characterization of KCNT1 variants to date. These ongoing efforts have identified at least 9 previously uncharacterized KCNT1 variants as gain-of-function, further supporting the therapeutic rationale for KCNT1 down-regulation in EIMFS and SHE. Further studies systematically examining pathogenic and non-pathogenic variants will enable a deeper understanding of the structural basis of KCNT1 gain of function.

Funding: Please list any funding that was received in support of this abstract.: This work was supported by Biogen.