Background Early infantile epileptic encephalopathies are severe disorders consisting of early-onset refractory seizures accompanied often by significant developmental delay. The increasing availability of next-generation sequencing has facilitated the recognition of single gene mutations as an underlying aetiology of some forms of early infantile epileptic encephalopathies.
Objectives This study was designed to identify candidate genes as a potential cause of early infantile epileptic encephalopathy, and then to provide genetic and functional evidence supporting patient variants as causative.
Methods We used whole exome sequencing to identify candidate genes. To model the disease and assess the functional effects of patient variants on candidate protein function, we used in vivo CRISPR/Cas9-mediated genome editing and protein overexpression in frog tadpoles.
Results We identified novel de novo variants in neuronal differentiation factor 2 (NEUROD2) in two unrelated children with early infantile epileptic encephalopathy. Depleting neurod2 with CRISPR/Cas9-mediated genome editing induced spontaneous seizures in tadpoles, mimicking the patients’ condition. Overexpression of wild-type NEUROD2 induced ectopic neurons in tadpoles; however, patient variants were markedly less effective, suggesting that both variants are dysfunctional and likely pathogenic.
Conclusion This study provides clinical and functional support for NEUROD2 variants as a cause of early infantile epileptic encephalopathy, the first evidence of human disease caused by NEUROD2 variants.
- neuronal differentiation factor
- epileptic encephalopathy
- epilepsy and seizures
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KL and SM-A contributed equally.
MKK and SAL contributed equally.
Contributors AGS collected Xenopus functional data, and drafted and critically reviewed the manuscript. EKM collected Xenopus functional data, and drafted and critically reviewed the manuscript. KL and SM-A collected clinical study information, conducted assessments, and critically reviewed the manuscript. WJ conducted sequencing and bioinformatic analyses, and drafted and critically reviewed the manuscript. MTC and JJ conducted sequencing and bioinformatic analyses and critically reviewed the manuscript. MK and LJ collected clinical study information and critically reviewed the manuscript. MKK contributed to Xenopus functional data and critically reviewed the manuscript. SAL contributed to Xenopus functional data, and drafted and critically reviewed the manuscript.
Funding MKK is supported by NIH/NICHD (R01HD081379). MKK is a Mallinckrodt Scholar. EKM is supported by a grant from the Hartwell Foundation and is a Hartwell Fellow.
Competing interests MTC and JJ are both paid employees of GeneDx, a wholly owned subsidiary of OPKO Health. KL reports speaking fees unrelated to this work from BioMarin, Genzyme and Alexion. SAL is part owner of Qiyas Higher Health, a start-up company unrelated to this work.
Patient consent Parental/guardian consent obtained.
Ethics approval The study was approved by the Institutional Review Board (IRB) at Yale University.
Provenance and peer review Not commissioned; externally peer reviewed.
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