title: Targeted next generation sequencing as a diagnostic tool in epileptic disorders.
abstract: OBJECTIVE
  Epilepsies have a highly heterogeneous background with a strong genetic contribution. The variety of unspecific and overlapping syndromic and nonsyndromic phenotypes often hampers a clear clinical diagnosis and prevents straightforward genetic testing. Knowing the genetic basis of a patient's epilepsy can be valuable not only for diagnosis but also for guiding treatment and estimating recurrence risks.
  
  
  METHODS
  To overcome these diagnostic restrictions, we composed a panel of genes for Next Generation Sequencing containing the most relevant epilepsy genes and covering the most relevant epilepsy phenotypes known so far. With this method, 265 genes were analyzed per patient in a single step. We evaluated this panel on a pilot cohort of 33 index patients with concise epilepsy phenotypes or with a severe but unspecific seizure disorder covering both sporadic and familial cases.
  
  
  RESULTS
  We identified presumed disease-causing mutations in 16 of 33 patients comprising sequence alterations in frequently as well as in less commonly affected genes. The detected aberrations encompassed known and unknown point mutations (SCN1A p.R222X, p. E289V, p.379R, p.R393H; SCN2A p.V208E; STXBP1 p.R122X; KCNJ10 p.L68P, p.I129V; KCTD7 p.L108M; KCNQ3 p.P574S; ARHGEF9 p.R290H; SMS p.F58L; TPP1 p.Q278R, p.Q422H; MFSD8 p.T294K), a putative splice site mutation (SCN1A c.693A> p.T/P231P) and small deletions (SCN1A p.F1330Lfs3X [1 bp]; MFSD8 p.A138Dfs10X [7 bp]). All mutations have been confirmed by conventional Sanger sequencing and, where possible, validated by parental testing and segregation analysis. In three patients with either Dravet syndrome or myoclonic epilepsy, we detected SCN1A mutations (p.R222X, p.P231P, p.R393H), even though other laboratories had previously excluded aberrations of this gene by Sanger sequencing or high-resolution melting analysis.
  
  
  CONCLUSIONS
  We have developed a fast and cost-efficient diagnostic screening method to analyze the genetic basis of epilepsies. We were able to detect mutations in patients with clear and with unspecific epilepsy phenotypes, to uncover the genetic basis of many so far unresolved cases with epilepsy including mutation detection in cases in which previous conventional methods yielded falsely negative results. Our approach thus proved to be a powerful diagnostic tool that may contribute to collecting information on both common and unknown epileptic disorders and in delineating associated phenotypes of less frequently mutated genes.
authors.affiliation: Division of Human Genetics, University Children's Hospital Inselspital, Bern, Switzerland. johannes.lemke@insel.ch
authors.forename: Johannes R
authors.initials: JR
authors.lastname: Lemke
authors.forename: Erik
authors.initials: E
authors.lastname: Riesch
authors.forename: Tim
authors.initials: T
authors.lastname: Scheurenbrand
authors.forename: Max
authors.initials: M
authors.lastname: Schubach
authors.forename: Christian
authors.initials: C
authors.lastname: Wilhelm
authors.forename: Isabelle
authors.initials: I
authors.lastname: Steiner
authors.forename: Jörg
authors.initials: J
authors.lastname: Hansen
authors.forename: Carolina
authors.initials: C
authors.lastname: Courage
authors.forename: Sabina
authors.initials: S
authors.lastname: Gallati
authors.forename: Sarah
authors.initials: S
authors.lastname: Bürki
authors.forename: Susi
authors.initials: S
authors.lastname: Strozzi
authors.forename: Barbara Goeggel
authors.initials: BG
authors.lastname: Simonetti
authors.forename: Sebastian
authors.initials: S
authors.lastname: Grunt
authors.forename: Maja
authors.initials: M
authors.lastname: Steinlin
authors.forename: Michael
authors.initials: M
authors.lastname: Alber
authors.forename: Markus
authors.initials: M
authors.lastname: Wolff
authors.forename: Thomas
authors.initials: T
authors.lastname: Klopstock
authors.forename: Eva C
authors.initials: EC
authors.lastname: Prott
authors.forename: Rüdiger
authors.initials: R
authors.lastname: Lorenz
authors.forename: Christiane
authors.initials: C
authors.lastname: Spaich
authors.forename: Sabine
authors.initials: S
authors.lastname: Rona
authors.forename: Maya
authors.initials: M
authors.lastname: Lakshminarasimhan
authors.forename: Judith
authors.initials: J
authors.lastname: Kröll
authors.forename: Thomas
authors.initials: T
authors.lastname: Dorn
authors.forename: Günter
authors.initials: G
authors.lastname: Krämer
authors.forename: Matthis
authors.initials: M
authors.lastname: Synofzik
authors.forename: Felicitas
authors.initials: F
authors.lastname: Becker
authors.forename: Yvonne G
authors.initials: YG
authors.lastname: Weber
authors.forename: Holger
authors.initials: H
authors.lastname: Lerche
authors.forename: Detlef
authors.initials: D
authors.lastname: Böhm
authors.forename: Saskia
authors.initials: S
authors.lastname: Biskup
mesh_terms: D000293:Adolescent; D000328:Adult; D002648:Child; D002675:Child, Preschool; D004827:Epilepsy / Q000175:diagnosis / Q000235:genetics*; D005260:Female; D005796:Genes / Q000235:genetics; D020022:Genetic Predisposition to Disease; D005838:Genotype; D006801:Humans; D008297:Male; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D017422:Sequence Analysis, DNA; D000091346:Tripeptidyl-Peptidase 1; D055815:Young Adult