Abstracts

Rationale: Developmental and epileptic encephalopathies (DEE) are a large heterogeneous group of severe, early onset epilepsies characterized by refractory seizures, developmental delay and/or regression, and often a poor prognosis. Next generation sequencing approaches have revealed de novo variants in coding regions of genes as the most common cause of DEE, with clinical diagnostic labs reporting a molecular diagnosis in 24-40% of cases. Despite this significant progress, the majority of DEE cases remain unexplained, even after whole exome sequencing.We sought to investigate potential disease-causing variants in noncoding regions of the genome. Methods: We are performing whole genome sequencing (WGS) of 20 exome-negative proband-parent trios. However, a major hurdle for genome-based diagnostics is our ability to interpret the functional and clinical impact of variants in non-coding sequence. In order to determine whether noncoding variants of interest impact gene expression, we will use RNA-seq to detect changes in transcript abundance, evidence of allele specific expression, and the presence of aberrant splicing. Recent large-scale gene expression studies have highlighted the importance of using disease-relevant tissue in transcriptome sequencing; analysis of available data confirms that the majority of established DEE genes are poorly expressed in skin and whole blood, limiting the use of these easily accessible tissues. To overcome this limitation, we are approximating brain tissue by reprogramming primary skin fibroblasts into neurons. We will use genome-wide expression data from the reprogrammed neurons to interpret the clinical relevance of noncoding de novo mutations identified by WGS in patients with exome-negative DEE. Results: To date, we have performed whole genome sequencing on 5 of 20 exome-negative cases and identified 37-68 de novo variants per trio. We have collected fibroblasts from three cases and established a protocol to directly reprogram primary skin fibroblasts to neuronal cells that will be used to aid in the functional interpretation of these sequence variants. Conclusions: This work establishes a framework for the investigation of non-coding variants in DEE and will facilitate improved diagnostic testing for DEE. Funding: AES Postdoctoral Research FellowshipDSF’s Postdoctoral Research FellowshipNIH R01 NS069605