No abstract

SCN2Aencodes Nav1.2, an excitatory neuron voltage-gated sodium channel and major monogenic cause of neurodevelopmental disorders, including developmental and epileptic encephalopathies (DEE) and autism. Clinical presentation and pharmocosensitivity vary with nature ofSCN2Avariant dysfunction with gain-of-function (GoF) cases presenting with pre- or peri-natal seizures and loss-of-function (LoF) patients typically having infantile spasms after 6 months of age. Here, we established and assessed patient induced pluripotent stem cell (iPSC) - derived neuronal models for two recurrentSCN2ADEE variants with GoF R1882Q and LoF R853Q associated with early- and late-onset DEE, respectively. Patient-derived iPSC lines were differentiated using aNeurogenin-2overexpression yielding populations of cortical-like glutamatergic neurons. Electrophysiological and transcriptomic profiles were assessed after 2-4 weeks in culture. Increased neuronal activity at both cellular and network level was observed for R1882Q iPSC-derived neurons at three weeks of differentiation. In contrast, R853Q neurons showed only subtle changes in excitability after four weeks in vitro. In alignment with the reported efficacy in some GoFSCN2Apatients, phenytoin (sodium channel blocker) reduced excitability of neurons to the control levels in R1882Q neuronal cultures. Transcriptomic alterations in neurons were detected for each variant and convergent pathways pointed at the shared mechanisms underlyingSCN2ADEE.

Objective: De novo missense variants in KCNQ5, encoding the voltage–gated K+ channel KV7.5, have been described as a cause of developmental and epileptic encephalopathy (DEE) or intellectual disability (ID). We set out to identify disease–related KCNQ5 variants in genetic generalized epilepsy (GGE) and their underlying mechanisms. Methods: 1292 families with GGE were studied by next-generation sequencing. Whole–cell patch–clamp recordings, biotinylation and phospholipid overlay assays were performed in mammalian cells combined with docking and homology modeling. Results: We identified three deleterious heterozygous missense variants, one truncation and one splice site alteration in five independent families with GGE with predominant absence seizures, two variants were also associated with mild to moderate ID. All three missense variants displayed a strongly decreased current density indicating a loss–of–function (LOF). When mutant channels were co–expressed with wild–type (WT) KV7.5 or KV7.5 and KV7.3 channels, three variants also revealed a significant dominant–negative effect on WT channels. Other gating parameters were unchanged. Biotinylation assays indicated a normal surface expression of the variants. The p.Arg359Cys variant altered PI(4,5)P2–interaction, presumably in the non–conducting preopen–closed state. Interpretation: Our study indicates that specific deleterious KCNQ5 variants are associated with GGE, partially combined with mild to moderate ID. The disease mechanism is a LOF partially with dominant–negative effects through functional, rather than trafficking deficits. LOF of KV7.5 channels will reduce the M–current, likely resulting in increased excitability of KV7.5–expressing neurons. Further studies on a network level are necessary to understand which circuits are affected and how the variants induce generalized seizures.