Pathogenic variants of KCNQ2 lead to a spectrum of disorders including self-limited familial neonatal-infantile epilepsy (SeL(F)NIE), developmental and epileptic encephalopathies (DEEs), and neurodevelopmental disorders (NDDs) with intellectual disability (ID). This study aimed to delineate the clinical progression and underlying pathogenesis of these disorders. Particularly, we unraveled the role of gain-of-function (GoF) variants in neurodevelopmental impairment. We conducted a longitudinal study of a 90-patient Chinese cohort with KCNQ2-related disorders, classified into SeL(F)NIE, DEEs, and NDDs subgroups. Clinical phenotyping was integrated with functional analyses (electrophysiology, biochemistry) of five missense variants in homomeric and heteromeric (with Kv7.3/Kv7.5) channel assemblies. Despite comparable seizure control to SeL(F)NIE (96% vs 100%), the NDDs group exhibited significant cognitive impairment. All deceased patients (8/90) were in the DEEs group. Functional profiling revealed a spectrum from loss-of-function (LoF) to GoF. LoF variants (e.g., S247L in DEEs) were linked to severe epilepsy. Crucially, we identified strong GoF variants (P335A/L in NDDs) in the S6-HelixA domain that confer insensitivity to phosphatidylinositol 4,5-bisphosphate (PIP2) regulation and are associated with a primary neurodevelopmental phenotype, distinct from the severe epileptic phenotype of established voltage-sensing domain (VSD) GoF variants. Our integrated clinical and functional analysis showed that the clinical outcome relies on the functional consequence of a KCNQ2 variant (LoF vs GoF) and its behavior in heteromeric complexes, rather than its mere location. We defined a novel class of strong GoF KCNQ2 variants that are mechanistically and phenotypically distinct, highlighting aberrantly enhanced channel function as a key driver of cognitive dysfunction and presenting new targets for precision medicine.

Variants in KCNQ2 encoding the voltage-gated potassium channel KV7.2 are associated with developmental and epileptic encephalopathy (DEE) of varying severity. This study examined the relationship of KCNQ2 variant dysfunction with the neurodevelopmental phenotype of individuals with KCNQ2-DEE. A parent-reported survey gathered clinical and genetic data for individuals with KCNQ2-DEE. Several clinical features were analyzed separately and as a composite non-seizure phenotype severity score (PSS) for six features (mobility, communication, hand use, eating, scoliosis, cerebral visual impairment). The effect of variants on KV7.2 channel function was determined by voltage-clamp recording in heterologous cells co-expressing KV7.3. Functional effects were classified as severe loss of function (SLOF), loss of function (LOF), wild-type-like (WTL), and gain of function (GOF). The study included 48 individuals each heterozygous for one of 38 unique variants. Median seizure-onset age was 1 day. Complete or significant seizure reduction was reported in 7/13 with carbamazepine, 13/17 with oxcarbazepine, 10/13 with phenytoin, and 3/4 with retigabine. The median PSS was 1 (interquartile range 1-3). On the participant level, 29 had SLOF variants, 13 had LOF variants, and the remaining participants had variants with GOF (3) or exhibited WTL (2) function. There were no significant associations of variant function with individual phenotypes in the PSS; however, the PSS itself was higher in those with SLOF versus LOF variants (p = 0.02). Among individuals with SLOF or LOF variants, there was an intriguing lower prevalence of epileptic spasms among individuals with dominant-negative variants. Multiple and severe neurodevelopmental impairments are common in KCNQ2-DEE. There was a modest correlation between KV7.2 channel dysfunction and overall non-seizure phenotype severity in this cohort. These findings suggest that factors other than differences in channel dysfunction contribute to variable clinical severity in KCNQ2-DEE. We examined how changes in the KCNQ2 gene, which affect the function of a brain potassium channel, relate to developmental and seizure features in children with KCNQ2-related epilepsy. Using parent surveys and lab studies of gene variants, we found that variants causing the channel to lose most of its function were linked to slightly worse overall development. Our results suggest that while channel dysfunction plays a role, other biological or environmental factors likely influence how severely children are affected.

Pathogenic variants in the KCNQ2 gene cause a spectrum of neonatal onset epilepsy, from self-limited familial neonatal epilepsy (SeLNE; mild end) to developmental and epileptic encephalopathy (DEE; severe end). The associations and differences in the molecular mechanisms between the developmental outcomes of different KCNQ2 variants (SeLNE vs. DEE) remain unclear. Using brain slice patch-clamp and single-nucleus RNA sequencing, we revealed developmental dysregulation in two different phenotypic Kcnq2 mice (DEE vs. SeLNE) during postnatal days 14-28 (P14-P28). Compared to wild-type mice, both Kcnq2-SeLNE and Kcnq2-DEE mice exhibited neuronal hyperexcitability characterized by high-frequency firing of action potentials. Notably, whereas SeLNE mice showed timely recovery of excitability, DEE mice displayed delayed restoration of abnormal excitability in CA1 excitatory neurons. During P14-P28, particularly at P21, DEE mice demonstrated significant downregulation of synaptic plasticity- and cognitive development-related pathways in CA1 excitatory neuron subclusters (CA1.2/CA1.4 neurons). Conversely, SeLNE mice exhibited pronounced activation of neurodevelopmental signaling pathways. Transcriptomic analysis of differentially expressed genes between SeLNE and DEE mouse models revealed recurrent gene signatures, with persistent neuronal upregulation of Apoe in Kcnq2-DEE mice. This study identifies that the age-related spontaneous remission of seizures is due to time-limited changes in neuronal excitability, and treatment interventions for KCNQ2-DEE patients need to consider critical developmental time windows. In the future, better therapeutic outcomes may be achieved through spatiotemporal transcriptional coordination with neurodevelopmental gene networks.

Epilepsy is a chronic neurological disorder characterized by recurrent seizures, approximately one-third of whom are resistant to current anti-seizure drugs (ASDs). Retigabine (RTG) is a potential treatment for treating drug-resistant epilepsy and KCNQ2-related developmental and epileptic encephalopathy (KCNQ2-DEE). However, its use is limited by side effects from high doses and long-term use. This study aims to evaluate the anticonvulsant efficacy of RTG in combination with (+)-borneol in mouse models of maximal electroshock seizure (MES) and 6-Hz (44-mA) seizure. The individual anti-seizure efficacy of RTG and (+)-borneol was evaluated in the MES and 6-Hz seizure models, then isobolographic analysis was conducted to assess their interactions. The plasma and brain concentrations of RTG were measured with and without (+)-borneol. Electrophysiological experiments using the patch-clamp technique investigated the interactions of (+)-borneol and RTG at the α1β3γ2L-GABAAR and KCNQ2 channels. Both RTG and (+)-borneol exhibited anticonvulsant activity in MES and 6-Hz seizure models. In the isobolographic analysis, the co-administration of RTG and (+)-borneol proved to be significantly more effective than predicted based on additive effects. The ED50mix was reduced by approximately 20 to 100-fold and 2 to 6-fold compared to the ED50add in the MES and 6-Hz models, respectively. The plasma and brain levels of RTG increased following co-administration with higher doses of (+)-borneol. Patch-clamp studies indicated that both RTG and (+)-borneol positively modulated α1β3γ2L-GABAAR currents and showed additive effects. However, (+)-borneol inhibited the KCNQ2 current at 100 µM and did not enhance RTG activation on KCNQ2 channels at this concentration. These results demonstrate that (+)-borneol enhances the antiseizure effects of RTG by both pharmacokinetic and pharmacodynamic interaction and this approach may be clinically effective for patients with intractable seizures or KCNQ2-DEE.

Voltage-gated Kv7 potassium channels, particularly Kv7.2 and Kv.7.3 channels, play a critical role in modulating susceptibility to seizures, and mutations in genes that encode these channels cause heterogeneous epilepsy phenotypes. On the basis of this evidence, activation of Kv7.2 and Kv.7.3 channels has long been considered an attractive target in the search for novel antiseizure medications. Ezogabine (retigabine), the first Kv7.2/3 activator introduced in 2011 for the treatment of focal seizures, was withdrawn from the market in 2017 due to declining use after discovery of its association with pigmentation changes in the retina, skin, and mucosae. A novel formulation of ezogabine for pediatric use (XEN496) has been recently investigated in children with KCNQ2-related developmental and epileptic encephalopathy, but the trial was terminated prematurely for reasons unrelated to safety. Among novel Kv7.2/3 openers in clinical development, azetukalner has shown dose-dependent efficacy against drug-resistant focal seizures with a good tolerability profile and no evidence of pigmentation-related adverse effects in early clinical studies, and it is now under investigation in phase III trials for the treatment of focal seizures, generalized tonic-clonic seizures, and major depressive disorder. Another Kv7.2/3 activator, BHV-7000, has completed phase I studies in healthy subjects, with excellent tolerability at plasma drug concentrations that exceed the median effective concentrations in a preclinical model of anticonvulsant activity, but no efficacy data in patients with epilepsy are available to date. Among other Kv7.2/3 activators in clinical development as potential antiseizure medications, pynegabine and CB-003 have completed phase I safety and pharmacokinetic studies, but results have not been yet reported. Overall, interest in targeting Kv7 channels for the treatment of epilepsy and for other indications remains strong. Future breakthroughs in this area could come from exploitation of mechanistic differences in the action of Kv7 activators, and from the development of molecules that combine Kv7 activation with other mechanisms of action.