Abstracts

Rationale: While some severe epilepsy syndromes, such as Dravet syndrome, have been known to be genetic in origin, many other epileptic encephalopathies, have until recently not been explained. Now, de novo mutations in epilepsy genes have been associated with infantile spasms, Lennox-Gastaut syndrome, and other rare early onset epilepsy syndromes.Methods: We used an Illumina Truseq custom amplicon assay to design a targeted panel to capture all exon and exon-intron boundaries of 34 genes known to cause epilepsy. Pooled amplicons were used to target exons from 250ng of DNA from each proband. PCR was performed using universal primers, with the introduction of unique 8-base barcodes on both the 3 and 5 end. Pooled libraries were subjected to massively parallel sequencing using a 251-paired end protocol on the MiSeq. Libraries were prepared and sequenced in batches. Raw read data and mapping was performed with a modified Burrows-Wheeler Aligner Smith Waterman (BWA-SW). SNV and indel calling and filtering were performed using the Genome Analysis Toolkit (GATK). We defined likely pathogenic variants as those that were rare, arose de novo, and were predicted to be pathogenic (SIFT, Polyphen).Results: We applied this assay to 20 individuals with severe epilepsy syndromes including Ohtahara syndrome and infantile spasms in whom clinical evaluation for etiology (including MRI and metabolic screening) was not revealing. We discovered 7 (35%) apparently pathogenic variants in 7 cases, including in the epilepsy-associated genes SCN1A, SCN2A, SCN2B, SCN8A, KCNQ2, EFHC1, and CHRNA7. We will report Sanger validation of this cohort as well as analysis of an additional 22 cases. Conclusions: Targeted resequencing of epilepsy genes allows for rapid diagnosis in about one third of patients with severe early onset epilepsy and has a high diagnostic yield. While most of the genes we identified have been associated with severe epilepsy syndromes, EFHC1 and CHRNA7 are usually associated with milder epilepsy syndromes and these findings suggest they play a role in the epileptic encephalopathies.