1. Genetic testing for the epilepsy specialist- focal or generalised? East Midlands Epilepsy Interest Group 11 February 2014 Abhijit Dixit

2. GENOME EXOME Protein-coding ‘exons’ of all genes Just 1% of the genome 3.2 Gb

3. Examining genes, chromosomes, exomes and genomes ArrayCGH 1000X resolution Karyotype Sanger sequencing Next-gen sequencing

4. Outline Why? Types (new) of inheritance in epilepsy New genes Emerging landscape of epilepsy genetics Role of next generation sequencing – Multi-gene panels – Exome and genome sequencing Changing role of the genetics service (and neurology)

5. Why make a diagnosis? The George Mallory argument Alter treatment Clarify prognosis Recurrence risk; prenatal diagnosis; PGD Contribute to basic understanding of human biology

7. 4 yr old boy

8. EAST syndrome Homozygous for c.194G>C (p.Arg65Pro) mutation in KCNJ10 gene

9. No diagnosis obvious ……(most cases) Hildebrand MS, et al. J Med Genet 2013 Same model applicable to intellectual disability and autism

10. Inherited Epilepsy Autosomal Dominant X-linked (recessive or dominant) Autosomal Recessive ADNFLE Glut1DS MECP2 CASK NEMO

11. Inherited Epilepsy Autosomal Dominant X-linked (recessive or dominant) Autosomal Recessive Mitochondrial ADNFLEGlut1DS POLG MECP2 CASK NEMO MERRF

12. Inherited Epilepsy Autosomal Dominant X-linked (recessive or dominant) Autosomal Recessive Mitochondrial ADNFLEGlut1DS POLG MECP2 CASK NEMO MERRF ?

13. Inherited Epilepsy Autosomal Dominant X-linked (recessive or dominant) Autosomal Recessive Mitochondrial ADNFLEGlut1DS POLG MECP2 CASK NEMO MERRF De novo Mosaic

15. Karyotype ArrayCGH MLPA Sequencing Standard Next gen Exome Genome

16. ArrayCGH vs Next Gen Sequencing

17. Benign familial neonatal seizures KCNQ2; KCNQ3 Early myoclonic encephalopathy ERBB4 Ohtahara syndrome GNAO1, STXBP1, ARX, CASK, KCNQ2

18. West syndrome multiple Migrating partial seizures of infancy KCNT1 Benign familial infantile seizures PRRT2 Dravet syndrome SCN1A

19. Early onset benign childhood occipital epilepsy (Panayiotopoulos type) Complex Febrile seizures plus SCN1A EE with continuous spike-and-wave during sleep (CSWS) Landau-Kleffner syndrome (LKS) GRIN2A Lennox- Gastaut syndrome Multiple Autosomal dominant nocturnal frontal lobe epilepsy CHRNA4; CHRNB2; CHRNA2 Benign epilepsy with centro-temporal spikes GRIN2A Childhood absence epilepsy Complex

20. Progressive myoclonic epilepsies Unverricht-Lundborg disease CSTB, PRIKLE1, SCARB2 Lafora disease EPM2A; EPM2B Others- NCL Familial partial epilepsy with variable foci DEPDC5 Autosomal dominant partial epilepsy with auditory features (ADPEAF) LGI1 Juvenile absence epilepsy Juvenile myoclonic epilepsy Complex

21. Heterogeneity in etiology of epilepsy IGE show complex inheritance – IGE+LD has ~10% yield on arrayCGH Few EE have single gene for majority of cases – Dravet/SCN1A (~80%) and MPSI/KCNT1 (~50%) – Lesser extent CSWS-LKS/GRIN2A (~20%) Most EE (West/LGS) very heterogeneous – Multiple genes each accounting for ~1% Same gene can appear in EE and ‘benign’ lists

22. Whole Genome Sequencing

24. Sequencing is easy………… Human genome for $5000 in 15 minutes on desktop size machine Belly button-ome

25. Courtesy: The Channelopathist @ EuroEPINOMICS

29. Nature. 2013 Sep 12;501(7466):217-21

30. Neuron 80, October 2, 2013

31. Epilepsy Gene Panels No of genes No of patients Pick-upReference 1 6550010% Carvill et al Nat Genetics 2013 22653348% Lemke et al Epilepsia 2012 No of genes CostPick-upLaboratory 145£1200~15% Great Ormond St, London 231~£1000? Cardiff

32. ArrayCGH Nottingham Cytogenetics Lab ~1000 tests in last 5 years – 335 CNVs identified

33. ArrayCGH Pick-up depends on resolution of arrayCGH – Pathogenic CNV 1q21.1, 15q11.2, 15q13.3, 16p11.2, 16p13.11, 17q12, 22q11.2 Other ‘large’ deletions/duplications esp if de novo – Possibly pathogenic – Variant of unknown significance – Benign 69/335 Nottingham arrayCGH are above common CNVs

35. 15q11.2 deletion

36. 15q13.3 deletion

37. 16p13.11 deletion

38. Genetic testing in epilepsy All patients with GGE plus learning difficulties – ArrayCGH – Consider testing on suitable NGS panel All patients with ‘epileptic encephalopathy’ – NGS panel – Single gene targeted test in Dravet, MPSI or epilepsy-aphasia syndromes….may only be available as part of panel!

39. Deciphering Developmental Disorders Health Innovation Challenge Fund and Sanger Institute ddd_help@sanger.ac.uk

40. DDD Study- ~1100 results

42. Finding genes for genetic disease

43. The ‘power’ of technology…..

44. Bycatch Variants of uncertain significance, variants in more than one gene and incidental but very significant changes in other (eg cancer) genes

45. The analytical bottleneck Exome – 12000 variants Genome – 5000000 variants

47. Referral to clinical genetics ‘Syndromic’ presentations Complex result on NGS panel or arrayCGH Recruitment to DDD study Testing unaffected parents or siblings