A Role of SCN9A in Human Epilepsies, As a Cause of Febrile Seizures and As a Potential Modifier of Dravet Syndrome

Abstract

A follow-up study of a large Utah family with significant linkage to chromosome 2q24 led us to identify a new febrile seizure (FS) gene, SCN9A encoding Na_v1.7. In 21 affected members, we uncovered a potential mutation in a highly conserved amino acid, p.N641Y, in the large cytoplasmic loop between transmembrane domains I and II that was absent from 586 ethnically matched population control chromosomes. To establish a functional role for this mutation in seizure susceptibility, we introduced the orthologous mutation into the murine Scn9a ortholog using targeted homologous recombination. Compared to wild-type mice, homozygous Scn9a^N641Y/N641Y knockin mice exhibit significantly reduced thresholds to electrically induced clonic and tonic-clonic seizures, and increased corneal kindling acquisition rates. Together, these data strongly support the SCN9A p.N641Y mutation as disease-causing in this family. To confirm the role of SCN9A in FS, we analyzed a collection of 92 unrelated FS patients and identified additional highly conserved Na_v1.7 missense variants in 5% of the patients. After one of these children with FS later developed Dravet syndrome (severe myoclonic epilepsy of infancy), we sequenced the SCN1A gene, a gene known to be associated with Dravet syndrome, and identified a heterozygous frameshift mutation. Subsequent analysis of 109 Dravet syndrome patients yielded nine Na_v1.7 missense variants (8% of the patients), all in highly conserved amino acids. Six of these Dravet syndrome patients with SCN9A missense variants also harbored either missense or splice site SCN1A mutations and three had no SCN1A mutations. This study provides evidence for a role of SCN9A in human epilepsies, both as a cause of FS and as a partner with SCN1A mutations. Febrile seizures are the most common seizure disorder of early childhood, and exhibit a prevalence of 2%–5% in European and North American children. While the genetic basis of febrile seizures is well-documented, efforts to uncover these genes have yielded only a few genes in a small proportion of cases. In a genomic region on human chromosome 2 known to harbor the febrile seizure SCN1A sodium channel gene, we now report a disease-causing mutation in the adjacent gene, SCN9A (Na_v1.7), in a large family with febrile seizures. We introduced the family mutation (N641Y) into the orthologous mouse gene to create a knockin mouse model, and tested seizure susceptibility in these mice. Compared to wild-type mice, our Scn9a knockin mice have a significantly lower threshold to electrically induced seizures and experience seizures at a significantly faster rate with repeated subthreshold stimulation. We also report novel missense SCN9A mutations in unrelated febrile seizure patients. Furthermore, we show that a subset of patients with the catastrophic early-onset Dravet syndrome who commonly have mutations in SCN1A also harbor mutations in SCN9A. This finding is important as it demonstrates for the first time mutational evidence for a modifying digenic mechanism of human epilepsy. For infants with Dravet syndrome, a genetic diagnosis will be of immediate benefit to guide therapeutics away from the sodium channel blocking class of anticonvulsant drugs that exacerbate seizures but are often the first administered.