# From Genetics to Genomics of Epilepsy

Authors: Silvio Garofalo; Marisa Cornacchione; Alfonso Di Costanzo
Journal: Neurology Research International
 (2012)

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## Abstract

The introduction of DNA microarrays and DNA sequencing technologies in medical genetics and diagnostics has been a challenge that has significantly transformed medical practice and patient management. Because of the great advancements in molecular genetics and the development of simple laboratory technology to identify the mutations in the causative genes, also the diagnostic approach to epilepsy has significantly changed. However, the clinical use of molecular cytogenetics and high-throughput DNA sequencing technologies, which are able to test an entire genome for genetic variants that are associated with the disease, is preparing a further revolution in the near future. Molecular Karyotype and Next-Generation Sequencing have the potential to identify causative genes or loci also in sporadic or non-familial epilepsy cases and may well represent the transition from a genetic to a genomic approach to epilepsy.


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## Body

## 1. Introduction

In the last decades a large number of gene discoveries have changed our views of idiopathic and symptomatic epilepsy [1]. Indeed, idiopathic epilepsy has the considerable genetic advantage to be found very often in informative autosomal dominant families that have been of great relevance to map and to positional clone the causative gene, opening insight into the biology and molecular pathology of this condition [2, 3].The search of epilepsy genes has allowed the identification of several genes in idiopathic generalized epilepsy (Table1), the vast majority of which are channelopathies [4, 5] or affect the activity of excitatory or inhibitory neurotransmitters in central nervous system [6]. It is possible that the dominant nature of these genes due to the multisubunit composition of the molecules have greatly overestimated the role of their mutations in the disease.Table 1
Disease genes identified in generalized myoclonic epilepsy, febrile seizures, absences (37 genes).
Gene SymbolGene name and descriptionALDH7A1Aldehyde dehydrogenase 7 family, member A1BRD2Bromodomain containing 2CACNA1ACalcium channel, voltage-dependent, P/Q type, alpha 1A subunitCACNA1HCalcium channel, voltage-dependent, T type, alpha 1H subunitCACNB4Calcium channel, voltage-dependent, beta 4 subunitCASRCalcium-sensing receptorCHRNA2Cholinergic receptor, nicotinic, alpha 2 (neuronal)CHRNA4Cholinergic receptor, nicotinic, alpha 4CHRNB2Cholinergic receptor, nicotinic, beta 2 (neuronal)CLCN2Chloride channel 2CSTBCystatin B (stefin B)EFHC1EF-hand domain (C-terminal) containing 1EPM2AEpilepsy, progressive myoclonus type 2A, Lafora disease (laforin)GABRA1Gamma-aminobutyric acid (GABA) A receptor, alpha 1GABRB3Gamma-aminobutyric acid (GABA) A receptor, beta 3GABRDGamma-aminobutyric acid (GABA) A receptor, deltaGABRG2Gamma-aminobutyric acid (GABA) A receptor, gamma 2GPR98G protein-coupled receptor 98GRIN2AGlutamate receptor, ionotropic, N-methyl D-aspartate 2AGRIN2BGlutamate receptor, ionotropic, N-methyl D-aspartate 2BKCNMA1Potassium large conductance calcium-activated channel, subfamily M, alpha member 1KCNQ2Potassium voltage-gated channel, KQT-like subfamily, member 2KCNQ3Potassium voltage-gated channel, KQT-like subfamily, member 3KCTD7Potassium channel tetramerisation domain containing 7MBD5Methyl-CpG-binding domain protein 5ME2Malic enzyme 2, NAD(+)-dependent, mitochondrialNHLRC1NHL repeat containing 1PCDH19Protocadherin 19PRICKLE1Prickle homolog 1 (Drosophila)PRICKLE2Prickle homolog 2 (Drosophila)SCARB2Scavenger receptor class B, member 2SCN1ASodium channel, voltage-gated, type I, alpha subunitSCN1BSodium channel, voltage-gated, type I, beta subunitSCN2ASodium channel, voltage-gated, type II, alpha subunitSCN9ASodium channel, voltage-gated, type IX, alpha subunitSLC2A1Solute carrier family 2 (facilitated glucose transporter), member 1TBC1D24TBC1 domain family, member 24Other important insights came from the discoveries of causative genes of syndromic epilepsy (Table2) [7] and other disorders where epilepsy is associated with encephalopathies (Table 3) [8], mental retardation with brain malformation (Table 4) [9, 10], other neurologic conditions including neuronal migration disorders (Table 5) [11], and inborn errors of metabolism (Tables 6 and 7) [12, 13]. Without any doubt, these discoveries have been great advances in the field; however, their impact on the management of epileptic patients was limited because of the failure to collect significant genetic information from each patient to distinguish the large number of genetic defects that can lead to the disease. Therefore, genetic testing was possible only for few or selected family cases.Table 2
Disease genes identified in syndromic epilepsy (47 genes).
Gene symbolGene name and descriptionSyndromeARFGEF2ADP-ribosylation factor GEF2Periventricular heterotopiaARHGEF9Cdc42 GEF 9Hyperekplexia with epilepsyA2BP1Ataxin 2-binding protein 1 (RNA binding protein fox-1 homolog 1)Mental retardation and epilepsyASPAAspartoacylaseCanavan syndromeATP1A2ATPase, Na/K transporting, alpha 2 polypeptideFamilial hemiplegic migraineATP2A2ATPase, Ca transporting, cardiac muscle, slow twitch 2Darier-White syndromeATP6V0A2ATPase, H+ transporting, lysosomal V0 subunit a2Cutis laxa with epilepsy and mental retardationCACNA1ACalcium channel, voltage-dependent, P/Q type, alpha 1A subunitFamilial hemiplegic migraineCCDC88CCoiled-coil domain containing 88CHydrocephalus with medial diverticulumCLCNKAChloride channel KaBartter syndromeCLCNKBChloride channel KbBartter syndromeCOH1Cohen syndrome protein 1—vacuolar protein sorting 13 homolog BCohen syndromeDLGAP2Discs, large (Drosophila) homolog-associated protein 2Progressive epilepsy with mental retardationGFAPGlial fibrillary acidic proteinAlexander diseaseGLI3GLI family zinc finger 3Pallister-hall syndromeGLRA1Glycine receptor, alpha 1HyperekplexiaGLRBGlycine receptor, betaHyperekplexiaGPHNGephyrinHyperekplexiaKCNA1Potassium voltage-gated channel, shaker-relatedEpisodic ataxiaKCNJ1Potassium inwardly rectifying channel, subfamily J, member 1Bartter syndromeKCNJ10Potassium inwardly rectifying channel, subfamily J, member 10Seizures, deafness, ataxia, mental retardationKIAA1279Kinesin family member 1 binding proteinGoldberg-ShprintzenLAMA2Laminin, alpha 2Merosin deficiencyLBRLamin B receptorPelger-Huet syndromeLGI1Leucine-rich, glioma inactivated 1Autosomal dominant lateral temporal lobe epilepsyMLC1Megalencephalic leukoencephalopathy with subcortical cysts 1Megalencephalic leukoencephalopathy with cystsMLL2Myeloid/lymphoid or mixed-lineage leukemia 2Kabuki syndromeNF1Neurofibromin 1NeurofibromatosisNIPBLNipped-B homolog (Drosophila)Cornelia de Lange syndromePANK2Pantothenate kinase 2Neurodegeneration with brain iron accumulationPI12Serpin peptidase inhibitor, clade I (neuroserpin), member 1Encephalopathy with neuroserpin inclusion bodiesPIGVPhosphatidylinositol glycan anchor biosynthesis, class VHyperphosphatasia with mental retardationPLA2G6Phospholipase A2, group VI (cytosolic, calcium independent)Infantile neuroaxonal dystrophyRAI1Retinoic acid induced 1Smith Magenis syndromeSCN8ASodium channel, voltage gated, type VIII, alpha subunitCerebellar atrophy, ataxia, and mental retardationSETBP1SET binding protein 1Schinzel-Giedion midface retraction syndromeSHHSonic hedgehogHoloprosencephalySLC4A10Solute carrier family 4, sodium bicarbonate transporter, member 10Epilepsy with mental retardationSLC6A5Solute carrier family 6 (neurotransmitter transporter, glycine), member 5HyperekplexiaSMC1AStructural maintenance of chromosomes 1ACornelia de lange syndromeSMC3Structural maintenance of chromosomes 3Cornelia de lange syndromeSYNGAP1Synaptic Ras GTPase activating protein 1Epilepsy and mental retardationTBX1T-box 1Di George syndromeTSC1Tuberous sclerosis 1Tuberous sclerosisTSC2Tuberous sclerosis 2Tuberous sclerosisVPS13AVacuolar protein sorting 13 homolog ANeuroacanthocytosisZEB2Zinc finger E-box binding homeobox 2Mowat-Wilson syndromeTable 3
Disease genes identified in epileptic encephalopathies (30 genes).
Gene symbolGene Name and DescriptionDiseasesARHGEF9Cdc42 guanine nucleotide exchange factor (GEF) 9Early infantile epileptic encephalopathyARXAristaless related homeoboxEarly infantile epileptic encephalopathyCDKL5Cyclin-dependent kinase-like 5Early infantile epileptic encephalopathyCNTNAP2Contactin associated protein-like 2Pitt Hopkins syndromeFOXG1Forkhead box G1Rett syndromeGABRG2Gamma-aminobutyric acid (GABA) A receptor, gamma 2Early infantile epileptic encephalopathyGRIN2AGlutamate receptor, ionotropic, N-methyl D-aspartate 2AEarly infantile epileptic encephalopathyGRIN2BGlutamate receptor, ionotropic, N-methyl D-aspartate 2BEarly infantile epileptic encephalopathyMAPK10Mitogen-activated protein kinase 10Lennox Gastaut syndromeMECP2Methyl CpG binding protein 2Rett syndromeNRXN1Neurexin 1Pitt Hopkins SyndromePCDH19Protocadherin 19Early infantile epileptic encephalopathyPNKPPolynucleotide kinase 3’-phosphataseEarly infantile epileptic encephalopathyRNASEH2ARibonuclease H2, subunit AAicardi-Goutieres syndromeRNASEH2BRibonuclease H2, subunit BAicardi-Goutieres syndromeRNASEH2CRibonuclease H2, subunit CAicardi-Goutieres syndromeSAMHD1SAM domain and HD domain 1Aicardi-Goutieres syndromeSCN1ASodium channel, voltage-gated, type I, alpha subunitEarly infantile epileptic encephalopathySCN1BSodium channel, voltage-gated, type I, beta subunitEarly Infantile epileptic encephalopathySCN2ASodium channel, voltage-gated, type II, alpha subunitEarly infantile epileptic EncephalopathySCN9ASodium channel, voltage-gated, type IX, alpha subunitEarly infantile epileptic encephalopathySLC2A1Solute carrier family 2 (facilitated glucose transporter), member 1GLUT1 deficiency syndromeSLC25A22Solute carrier family 25 (mitochondrial carrier: glutamate), member 22Early infantile epileptic encephalopathySLC9A6Solute carrier family 9 (sodium/hydrogen exchanger), member 6Angelman syndromeSPTAN1Spectrin, alpha, non-erythrocytic 1 (alpha-fodrin)Early infantile epileptic encephalopathySTXBP1Syntaxin binding protein 1Early infantile epileptic encephalopathyTCF4Transcription factor 4Pitt Hopkins syndromeTREX1Three prime repair exonuclease 1Aicardi-Goutieres syndromeUBE3AUbiquitin protein ligase E3AAngelman syndromeZEB2Zinc finger E-box binding homeobox 2Mowat-Wilson syndromeTable 4
Epilepsy with mental retardation and brain malformations.
Gene symbolNameDisease(a) Mental retardation (25 genes)ARHGEF9Cdc42 guanine nucleotide exchange factor (GEF) 9Early infantile epileptic encephalopathyARXAristaless related homeoboxEarly infantile epileptic encephalopathyATP6AP2ATPase, H+ transporting, lysosomal accessory protein 2Epilepsy with XLMR*ATRXAlpha thalassemia/mental retardation syndrome X-linkedEpilepsy with XLMR*CASKCalcium/calmodulin-dependent serine protein kinase (MAGUK family)Mental retardation and microcephalyCDKL5Cyclin-dependent kinase-like 5Early infantile epileptic encephalopathyCUL4BCullin 4BEpilepsy with XLMR*CXORF5Oral-facial-digital syndrome 1Simpson-Golabi-Behmel syndromeDCXDoublecortinLissencephalyFGD1FYVE, RhoGEF and PH domain containing 1Aarskog-Scott syndromeGPC3Glypican 3Simpson-Golabi-Behmel syndromeGRIA3Glutamate receptor, ionotrophic, AMPA 3Epilepsy with XLMR*HSD17B10Hydroxysteroid (17-beta) dehydrogenase 10Epilepsy with XLMR*JARID1CLysine (K)-specific demethylase 5CEpilepsy with XLMR*OPHN1Oligophrenin 1Epilepsy with XLMR*PAK3P21 protein (Cdc42/Rac)-activated kinase 3Epilepsy with XLMR*PHF6PHD finger protein 6Borjeson Forssmann Lehmann syndromePLP1Proteolipid protein 1Pelizaeus-Merzbacher diseasePQBP1Polyglutamine binding protein 1Epilepsy with XLMR*RAB39BRAB39B, member RAS oncogene familyEpilepsy with XLMR*SLC9A6Solute carrier family 9 (sodium/hydrogen exchanger), member 6Angelman-Like syndromeSMC1AStructural maintenance of chromosomes 1ACornelia De Lange syndromeSMSSpermine synthaseEpilepsy with XLMR*SRPX2Sushi-repeat containing protein, X-linked 2Rolandic epilepsySYPSynaptophysinEpilepsy with XLMR**XLMR: X-linked mental retardation(b) Joubert syndrome (10 genes)AHI1Abelson helper integration site 1Joubert syndromeARL13BADP-ribosylation factor-like 13BJoubert syndromeCC2D2ACoiled-coil and C2 domain containing 2AJoubert syndromeCEP290Centrosomal protein 290 kDaJoubert syndromeCXORF5Oral-facial-digital syndrome 1Joubert syndromeINPP5EInositol polyphosphate-5-phosphatase, 72 kDaJoubert syndromeNPHP1Nephronophthisis 1 (juvenile)Joubert syndromeRPGRIP1LRetinitis pigmentosa GTPase regulator interacting protein 1 likeJoubert syndromeTMEM67Transmembrane protein 67Joubert syndromeTMEM216Transmembrane protein 216Joubert syndrome(c) Lissencephaly and polymicrogyria (18 genes)COL18A1Collagen, type XVIII, alpha 1PolymicrogyriaCPT2Carnitine palmitoyltransferase 2PolymicrogyriaDCXDoublecortinLissencephalyEOMESEomesoderminPolymicrogyriaFGFR3Fibroblast growth factor receptor 3PolymicrogyriaFLNAFilamin A, alphaPeriventricular heterotopiaGPR56G protein-coupled receptor 56PolymicrogyriaPAFAH1B1Platelet-activating factor acetylhydrolase 1b, regulatory subunit 1 (45kDa)LissencephalyPAX6Paired box 6PolymicrogyriaPEX7Peroxisomal biogenesis factor 7PolymicrogyriaRAB3GAP1RAB3 GTPase activating protein subunit 1 (catalytic)Warburg microsyndromeRELNReelinLissencephalySNAP29Synaptosomal-associated protein, 29 kDaCerebral dysgenesisSRPX2Sushi-repeat containing protein, X-linked 2Rolandic epilepsyTUBA1ATubulin, alpha 1aLissencephalyTUBA8Tubulin, alpha 8PolymicrogyriaTUBB2BTubulin, beta 2BPolymicrogyriaVDAC1Voltage-dependent anion channel 1Polymicrogyria(d) Severe microcephaly and pontocerebellar hypoplasia (22 genes)ASPMAsp (abnormal spindle) homolog, microcephaly associated (Drosophila)MicrocephalyATRAtaxia telangiectasia and Rad3 relatedMicrocephalyBUB1BBudding uninhibited by benzimidazoles 1 homolog beta (yeast)MicrocephalyCASKCalcium/calmodulin-dependent serine protein kinase (MAGUK family)MicrocephalyCDK5RAP2 [Microcephaly]CDK5 regulatory subunit associated protein 2MicrocephalyCENPJCentromere protein JMicrocephalyCEP152Centrosomal protein 152 kDaMicrocephalyLIG4Ligase IV, DNA, ATP-dependentMicrocephalyMCPH1Microcephalin 1MicrocephalyMED17Mediator complex subunit 17MicrocephalyNHEJ1Nonhomologous end-joining factor 1MicrocephalyPCNTPericentrinMicrocephalic osteodysplastic DwarfismPNKPPolynucleotide kinase 3'-phosphataseMicrocephalyPQBP1Polyglutamine binding protein 1X-linked mental retardationRARS2Arginyl-tRNA synthetase 2, mitochondrialPontocerebellar hypoplasiaSLC25A19Solute carrier family 25 (mitochondrial thiamine pyrophosphate carrier), member 19MicrocephalySTILSCL/TAL1 interrupting locusMicrocephalyTSEN2tRNA splicing endonuclease 2 homolog (S. cerevisiae)Pontocerebellar hypoplasiaTSEN34 [Pontocerebellar Hypoplasia]tRNA splicing endonuclease 34 homolog (S. cerevisiae)Pontocerebellar hypoplasiaTSEN54 [Pontocerebellar Hypoplasia]tRNA splicing endonuclease 54 homolog (S. cerevisiae)Pontocerebellar hypoplasiaVRK1Vaccinia related kinase 1Pontocerebellar hypoplasiaWDR62WD repeat domain 62Microcephaly, cortical malformationsand mental retardation(e) Walker-Warburg syndrome (WWS) or muscle, eye and brain disease (6 genes) anomalies type A2 (MDDGA2)FKRPFukutin-related proteinWalker-Warburg syndromeFKTNFukutinWalker-Warburg syndromeLARGELike-glycosyltransferaseWalker-Warburg syndromePOMGNT1Protein O-linked mannose beta1,2-N-acetylglucosaminyltransferaseWalker-Warburg syndromePOMT1Protein-O-mannosyltransferase 1Walker-Warburg syndromePOMT2Protein-O-mannosyltransferase 2Walker-Warburg Syndrome(f) Holoprosencephaly (HPE) (8 genes)FGF8Fibroblast growth factor 8 (androgen-induced)HoloprosencephalyGLI2GLI family zinc finger 2Holoprosencephaly 9GLI3GLI family zinc finger 3Greig cephalopolysyndactyly syndromePTCH1patched 1Holoprosencephaly 7SHHSonic HedgehogHoloprosencephaly 3SIX3SIX homeobox 3Holoprosencephaly 2TGIF1TGFB-induced factor homeobox 1Holoprosencephaly 4ZIC2Zic family member 2Holoprosencephaly 5Table 5
Epilepsy with other neurological problems.
Gene symbolNameDisease(a) Leukodystrophies (20 genes)ARSAArylsulfatase ALeukodystrophy metachromatic (MLD)ASPAAspartoacylaseCanavan diseaseEIF2B1Eukaryotic translation initiation factor 2B, subunit 1 alpha, 26 kDaLeukodystrophyEIF2B2Eukaryotic translation initiation factor 2B, subunit 2 beta, 39 kDaLeukodystrophyEIF2B3Eukaryotic translation initiation factor 2B, subunit 3 gamma, 58 kDaLeukodystrophyEIF2B4Eukaryotic translation initiation factor 2B, subunit 4 delta, 67 kDaLeukodystrophyEIF2B5Eukaryotic translation initiation factor 2B, subunit 5 epsilon, 82 kDaLeukodystrophyGALCGalactosylceramidaseLeukodystrophy globoid cell (GLD)GFAPGlial fibrillary acidic proteinAlexander diseaseMLC1Megalencephalic leukoencephalopathy with subcortical cysts 1Megalencephalic leukoencephalopathyNOTCH3Notch3CADASILPLP1Proteolipid protein 1Leukodystrophy hypomyelinating type 1 (HLD1)PSAPProsaposinLeukodystrophy metachromaticRNASEH2ARibonuclease H2, subunit AAicardi-Goutieres syndrome type 4 (AGS4)RNASEH2BRibonuclease H2, subunit BAicardi-Goutieres syndrome type 2 (AGS2)RNASEH2CRibonuclease H2, subunit CAicardi-Goutieres syndrome type 3 (AGS3SAMHD1SAM domain and HD domain 1Aicardi-Goutieres syndrome type 5 (AGS5)SDHASuccinate dehydrogenase complex, subunit A, flavoprotein (Fp)Leigh syndromeSUMF1Sulfatase modifying factor 1Multiple sulfatase deficiency (MSD)TREX1Three prime repair exonuclease 1Aicardi-Goutieres syndrome type 1 (AGS1)(b) Migraine (6 genes)ATP1A2ATPase, Na+/K+ transporting, alpha 2 polypeptideMigraine familial hemiplegic type 2 (FHM2)CACNA1ACalcium channel, voltage-dependent, P/Q type, alpha 1A subunitSpinocerebellar ataxia type 6 (SCA6)NOTCH3Notch 3CADASILPOLGPolymerase (DNA directed), gammaProgressive external ophthalmoplegiaSCN1ASodium channel, voltage-gated, type I, alpha subunitMigraine familial hemiplegic type 3 (FHM3)SLC2A1Solute carrier family 2 (facilitated glucose transporter), member 1GLUT1 deficiency type 1(GLUT1DS1) syndrome(c) Disorders of Ras-MAPK pathway with epilepsy (13 genes)BRAFV-raf murine sarcoma viral oncogene homolog B1Cardiofaciocutaneous (CFC ) syndromeCBLCas-Br-M (murine) ecotropic retroviral transforming sequenceNoonan syndrome-like disorder (NSL)HRASV-Ha-ras Harvey rat sarcoma viral oncogene homologFaciocutaneoskeletal (FCSS) syndromeKRASV-Ki-ras2 Kirsten rat sarcoma viral oncogene homologNoonan type 3 (NS3) syndromeMAP2K1Mitogen-activated protein kinase kinase 1cardiofaciocutaneous (CFC) syndromeMAP2K2Mitogen-activated protein kinase kinase 2cardiofaciocutaneous (CFC) syndromeNF1Neurofibromin 1Neurofibromatosis type 1NRASNeuroblastoma RAS viral (v-ras) oncogene homologNoonan type 6 (NS6) syndromePTPN11Protein tyrosine phosphatase, non-receptor type 11LEOPARD type 1 (LEOPARD1) syndromeRAF1V-raf-1 murine leukemia viral oncogene homolog 1Noonan type 5 (NS5) syndromeSHOC2Soc-2 suppressor of clear homolog (C. elegans)Noonan syndrome-like with loose anagen hairSOS1Son of sevenless homolog 1 (Drosophila)Noonan type 4 (NS4) syndromeSPRED1Sprouty-related, EVH1 domain containing 1Neurofibromatosis type 1-like syndrome(d) Hyperekplexia (5 genes)ARHGEF9Cdc42 guanine nucleotide exchange factor (GEF) 9Hyperekplexia with epilepsyGLRA1Glycine receptor, alpha 1Hyperekplexia with epilepsyGLRBGlycine receptor, betaHyperekplexia with epilepsyGPHNGephyrinHyperekplexia with epilepsySLC6A5solute carrier family 6 (neurotransmitter, transporter, glycine), member 5Hyperekplexia with epilepsy(e) Neuronal migration disorders (31 genes)ARFGEF2ADP-ribosylation factor guanine nucleotide-exchange factor 2 (brefeldin A-inhibited)MicrocephalyARXAristaless-related homeoboxEarly infantile epileptic encephalopathyCOL18A1Collagen, type XVIII, alpha 1PolymicrogyriaCOL4A1Collagen, type IV, alpha 1PorencephalyCPT2Carnitine palmitoyltransferase 2PolymicrogyriaDCXDoublecortinLissencephalyEMX2Empty spiracles homeobox 2SchizencephalyEOMESEomesoderminPolymicrogyriaFGFR3Fibroblast growth factor receptor 3PolymicrogyriaFKRPFukutin related proteinWalker-Warburg syndromeFKTNFukutinWalker-Warburg syndromeFLNAFilamin A, alphaPeriventricular heterotopiaGPR56G protein-coupled receptor 56PolymicrogyriaLAMA2Laminin, alpha2Merosin deficiencyLARGELike-glycosyltransferaseWalker-Warburg syndromePAFAH1B1Platelet-activating factor acetylhydrolase 1b, regulatory subunit 1 (45 kDa)LissencephalyPAX6Paired box 6PolymicrogyriaPEX7Peroxisomal biogenesis factor 7PolymicrogyriaPOMGNT1Protein O-linked mannose beta1,2-N-acetylglucosaminyltransferaseWalker-Warburg syndromePOMT1Protein O-mannosyltransferase 1Walker-Warburg syndromePOMT2Protein O-mannosyltransferase 2Walker-Warburg syndromePQBP1Polyglutamine binding protein 1X-linked mental retardationRAB3GAPRAB3 GTPase activating protein subunit 1 (catalytic)Warburg microsyndromeRELNReelinLissencephalySNAP29Synaptosomal-associated protein, 29 kDaCerebral dysgenesisSRPX2Sushi-repeat containing protein, X-linked 2Rolandic epilepsyTUBA1ATubulin, alpha 1aLissencephalyTUBA8Tubulin, alpha 8PolymicrogyriaTUBB2BVoltage-dependent anion channel 1PolymicrogyriaVDAC1Voltage-dependent anion channel 1PolymicrogyriaWDR62WD repeat domain 62Microcephaly, cortical malfor, mental retardatationTable 6
Inherited errors of metabolism with epilepsy (49 genes).
Gene symbolDefective enzyme nameDiseaseABCC8ATP-binding cassette, subfamily C (CFTR/MRP), member 8HypoglcemiaACY1Aminoacylase1Aminoacylase1 deficiencyADSLAdenylosuccinate lyaseAdenylosuccinase deficiencyAGAAspartylglucosaminidaseAspartylglucosaminuriaALDH4A1Aldehyde dehydrogenase 4 family, member A1HyperprolinemiaALDH5A1Aldehyde dehydrogenase 5 family, member A1Succinic Semialdehyde dehydrogenase deficiencyALDH7A1Aldehyde dehydrogenase 7 family, member A1Pyridoxine deficiencyARG1Liver arginaseArgininemiaARSAArylsulfatase AMetachromatic leukoodystrophyASPAAspartoacylaseCanavan diseaseATIC5-aminoimidazole-4-carboxamide ribonucleotide  (AICAr) formyltransferase/IMP cyclohydrolaseAICAr transformylase/IMP cyclohydrolase deficiency (ATIC Deficiency)BTDBiotinidaseBiotinidase deficiencyCPT2Carnitine palmitoyltransferase 2Carnitine palmitoyltransferase II deficiencyCTSACathepsin AGalactosialidosisDPYDDihydropyrimidine dehydrogenaseDihydropyrimidine dehydrogenase deficiencyETFAElectron-transfer-flavoprotein, alpha polypeptideGlutaraciduriaETFBElectron-transfer-flavoprotein, beta polypeptideGlutaraciduriaETFDHElectron-transferring-flavoprotein dehydrogenaseGlutaraciduriaFHFumarate hydrataseFumarase deficiencyFOLR1Folate receptor 1 (adult)Cerebral folate transport deficiencyFUCA1Fucosidase, alpha-L- 1, tissueFucosidosisGALCGalactosylceramidaseKrabbe diseaseGAMTGuanidinoacetate N-methyltransferaseGuanidinoacetate N-methyltransferase deficiencyGCDHGlutaryl-CoA dehydrogenaseGlutaraciduriaGCSHGlycine cleavage system protein H (aminomethyl carrier)Glycine encephalopathyGCSTGlycine cleavage system protein T (aminomethyltransferase)Glycine encephalopathyGLB1Galactosidase, beta 1GangliosidosisGLDCGlycine dehydrogenase (decarboxylating)Glycine encephalopathyGNEGlucosamine (UDP-N-acetyl)-2-epimerase/N-acetylmannosamine kinaseSialuriaHEXAHexosaminidase A (alpha polypeptide)GangliosidosisHEXBHexosaminidase B (beta polypeptide)GangliosidosisHPD4-Hydroxyphenylpyruvate dioxygenaseTyrosinemiaL2HGDHL-2-Hydroxyglutarate dehydrogenaseL-2-Hydroxyglutaric aciduriaLAMA2Laminin, alpha 2Muscular dystrophyMOCS1Molybdenum cofactor synthesis 1Molybdene cofactor deficiencyMOCS2Molybdenum cofactor synthesis 2Molybdene cofactor deficiencyNEU1Sialidase 1 (lysosomal sialidase)Neuraminidase deficiencyNPC1Niemann-Pick disease, type C1Niemann-Pick diseaseNPC2Niemann-Pick disease, type C2Niemann-Pick diseasePGK1Phosphoglycerate kinase 1GAMT deficiencyPRODHProline dehydrogenase (oxidase) 1HyperprolinemiaPSAPProsaposinKrabbe diseaseQDPRQuinoid dihydropteridine reductaseHyperphenylalaninemiaSLC17A5Solute carrier family 17 (anion/sugar transporter), member 5SialuriaSLC25A15Solute carrier family 25 (mitochondrial carrier; ornithine transporter) member 15Ornithine translocase deficiencySLC46A1Solute carrier family 46 (folate transporter), member 1Folate malabsorptionSMPD1Sphingomyelin phosphodiesterase 1, acid lysosomalNiemann pick diseaseSUMF1Sulfatase modifying factor 1SulfatidosisSUOXSulfite oxidaseSulfitoxidasisTable 7
Other inherited errors of metabolism with epilepsy.
Gene symbolDefective enzyme nameDisease(a) Congenital Disorder of Glycosylation (CDG) (23 genes)ALG1N-linked glycosylation 1, beta-1,4-mannosyltransferase homologCDGALG2N-linked glycosylation 2, alpha-1,3-mannosyltransferase homologCDGALG3N-linked glycosylation 3, alpha-1,3-mannosyltransferase homologCDGALG6N-linked glycosylation 6, alpha-1,3-glucosyltransferase homologCDGALG8N-linked glycosylation 8, alpha-1,3-glucosyltransferase homologCDGALG9N-linked glycosylation 9, alpha-1,3-glucosyltransferase homologCDGALG12N-linked glycosylation 12, alpha-1,3-glucosyltransferase homologCDGB4GALT1UDP-Gal: betaGlcNAc beta 1,4-galactosyltransferase, polypeptide 1CDGCOG1Component of oligomeric golgi complex 1CDGCOG7Component of oligomeric golgi complex 7CDGCOG8Component of oligomeric golgi complex 8CDGDOLKDolichol kinaseCDGDPAGT1Dolichyl-phosphate (UDP-N-acetylglucosamine) N-acetylglucosamine phosphotransferase 1 (GlcNAc-1-P transferase)CDGDPM1Dolichyl-phosphate mannosyltransferase polypeptide 1, catalytic subunitCDGDPM3Dolichyl-phosphate mannosyltransferase polypeptide 3CDGMOGSMannosyl-oligosaccharide glucosidaseCDGMGAT2Mannosyl (alpha-1,6-)-glycoprotein beta-1,2-N-acetylglucosaminyltransferaseCDGMPDU1Mannose-P-dolichol utilization defect 1CDGMPIMannose phosphate isomeraseCDGPMM2Phosphomannomutase 2CDGRFT1Requiring fifty three 1 homologCDGSLC35A1Solute carrier family 35 (CMP-sialic acid transporter), member A1CDGSLC35C1Solute carrier family 35, member C1CDG(b) Neuronal ceroid lipofuscinosis (NCL) (8 genes)CLN3Ceroid-lipofuscinosis, neuronal 3NLCCLN5Ceroid-lipofuscinosis, neuronal 5NLCCLN6Ceroid-lipofuscinosis, neuronal 6NLCCLN8Ceroid-lipofuscinosis, neuronal 8NLCCTSDCathepsin DNLCMFSD8Major facilitator superfamily domain containing 8NLCPPT1Palmitoyl-protein thioesterase 1NLCTPP1Tripeptidyl peptidase INLC(c) Defects of mitochondrial metabolism including coenzyme Q deficiency (35 genes)APTXAprataxinCoenzyme Q10 DeficiencyATPAF2ATP synthase mitochondrial F1 complex assembly factor 2ATPase deficiencyBCS1LBCS1-likeLeigh syndromeC12ORF65Chromosome 12 open reading frame 65Leigh syndromeC8ORF38Chromosome 8 open reading frame 38Leigh syndromeCABC1Chaperone activity of bc1 complex-like, mitochondriaCoenzyme Q10 deficiencyCOQ2Coenzyme Q2 homolog, prenyltransferase (yeast)Coenzyme Q10 deficiencyCOQ9Coenzyme Q9 homolog (S. cerevisiae)Coenzyme Q10 deficiencyCOX10COX10 homolog, cytochrome c oxidase assembly protein,          heme A: farnesyltransferase (yeast)Leigh syndromeCOX10COX15COX15 homolog, cytochrome c oxidase assembly protein (yeast)Leigh syndromeDLDDihydrolipoamide dehydrogenaseLeigh syndromeGCSHGlycine cleavage system protein H (aminomethyl carrier)Glycine encephalopathyGCSTAminomethyltransferase (glycine cleavage system protein T)Glycine encephalopathyGLDCGlycine dehydrogenase (decarboxylating)Glycine encephalopathyHSD17B10Hydroxysteroid (17-beta) dehydrogenase 10HSD17B10 deficiencyLRPPRCLeucine-rich PPR-motif containingLeigh syndromeNDUFA2NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 2,8 kDaLeigh syndromeNDUFS1NADH dehydrogenase (ubiquinone) Fe-S protein 1, 75 kDaLeigh syndromeNDUFS3NADH dehydrogenase (ubiquinone) Fe-S protein 3, 30 kDaLeigh syndromeNDUFS4NADH dehydrogenase (ubiquinone) Fe-S protein 4, 18 kDaLeigh syndromeNDUFS7NADH dehydrogenase (ubiquinone) Fe-S protein 7, 20 kDaLeigh syndromeNDUFS8NADH dehydrogenase (ubiquinone) Fe-S protein 8, 23 kDaLeigh syndromeNDUFV1NADH dehydrogenase (ubiquinone) flavoprotein 1, 51 kDaLeigh syndromePCPyruvate carboxylaseLeigh syndromePDHA1Pyruvate dehydrogenase (lipoamide) alpha 1Leigh syndromePDSS1Prenyl (decaprenyl) diphosphate synthase, subunit 1Coenzyme Q10 deficiencyPDSS2Prenyl (decaprenyl) diphosphate synthase, subunit 2Coenzyme Q10 deficiency]POLGPolymerase (DNA directed), gammaMitochondrial DNA depletion SyndromeRARS2Arginyl-tRNA synthetase 2, mitochondrialPontocerebellar hypoplasiaSCO2SCO cytochrome oxidase deficient homolog 2 (yeast)Leigh syndromeSDHASuccinate dehydrogenase complex, subunit A, flavoprotein (Fp)Leigh syndromeSURF1Surfeit 1Leigh syndromeTACO1Translational activator of mitochondrially encoded cytochrome c oxidase ILeigh syndromeTMEM70Transmembrane protein 70EncephalocardiomyopathyVDAC1voltage-dependent anion channel 1VDAC deficiency(d) Mucopolysaccharidosis (MPS) and mucolipidosis (MLP) (15 genes)ARSBArylsulfatase BMPS 6 (Maroteaux-Lamy syndrome)GALNSGalactosamine (N-acetyl)-6-sulfate sulfataseMPS 4A (Morquio syndrome)GLB1Galactosidase, beta 1GM1-gangliosidosisGNPTABN-acetylglucosamine-1-phosphate transferase, alpha and beta subunitsMucolipidosi 2 (I cell disease) and 3AGNPTGN-acetylglucosamine-1-phosphate transferase, gamma subunitMucolipidosi 3CGNSGlucosamine (N-acetyl)-6-sulfataseMPS 3D (Sanfilippo D syndrome)GUSBGlucuronidase, betaMPS 7 (Sly syndrome)HGSNATHeparan-alpha-glucosaminide N-acetyltransferaseMPS 3C (Sanfilippo C syndrome)HYAL1Hyaluronoglucosaminidase 1MPS 9IDSIduronate 2-sulfataseMPS 2 (Hunter syndrome)IDUAIduronidase, alpha-L-MPS 1H (Hurler syndrome)MCOLN1Nucolipin 1Mucolipidosi 4NAGLUN-acetylglucosaminidase, alphaMPS 3B (Sanfilippo B syndrome)SGSHN-sulfoglucosamine sulfohydrolaseMPS 3A (Sanfilippo A syndrome)SUMF1Sulfatase modifying factor 1Multiple sulfatase deficiency(e) Peroxisome biogenesis disorders (PBD) (9 genes): Zellweger syndrome (ZWS): neonatal adrenoleukodystrophy (NALD): infantile refsum disease (IRD): rhizomelic chondrodysplasia punctata type 1 (RCDP1)PEX1Peroxisomal biogenesis factor 1ZWS-NADL-IRDPEX2Peroxisomal biogenesis factor 2ZWS-IRDPEX3Peroxisomal biogenesis factor 3ZWSPEX5Peroxisomal biogenesis factor 5ZWS-NADLPEX6Peroxisomal biogenesis factor 6ZWSPEX7Peroxisomal biogenesis factor 7RCDP1PEX12Peroxisomal biogenesis factor 12ZWSPEX14Peroxisomal biogenesis factor 14ZWSPEX26Peroxisomal biogenesis factor 26ZWS-NADL-IRDTechnical improvements in human chromosomes recognition and better definition of chromosome regions realized by increasing the number of detectable chromosome bands have provided higher resolution of normal and pathological karyotype. It is today well established an association between epileptic seizures and chromosome abnormalities recognized by high-resolution chromosome banding [14, 15]. However, the type and the size of the chromosome defects are not always easy to detect even by the highest-resolution cytogenetic techniques available for light microscopes.The identification of the specific genetic defect in a patient with epilepsy may clarify the diagnosis (diagnostic testing), suggest the prognosis, assist with treatment and management (e.g., the use of a ketogenic diet in glucose transporter type 1 deficiency syndrome or the avoidance of lamotrigine, phenytoin, and carbamazepine in Dravet syndrome), elucidate the risk of a disease in family members and future children, and save the patient from further diagnostic evaluation and potentially invasive testing.In asymptomatic subjects with increased risk of seizures because of a family history, genetic test may predict onset of epilepsy (predictive testing) [16, 17]. Despite such potential benefits, genetic testing has also potential harms, such as its ethical, legal, and social implications, and the potential for stigma, distress, adverse labeling, and nonconfidentiality that exists in the setting of inadequate safeguards against discrimination [18]. Considering that our understanding of the epidemiology and clinical utility of genetic testing in the epilepsies is incomplete, the assessment of these potential benefits and harms is particularly complex and is closely linked to the clinical scenario.The International League Against Epilepsy (ILAE) Genetic Commission presented a tool in the approach to specific tests for epilepsy [16]. According to ILAE report, the diagnostic genetic testing is “very useful” in individual affected by early-onset spasms, X-linked infantile spasms, Dravet and related syndromes, Ohtahara syndrome, epilepsy and mental retardation limited to females, early-onset absence epilepsy, autosomal dominant nocturnal frontal lobe epilepsy, and epilepsy with paroxysmal exercise-induced dyskinesia; the predictive testing is “very useful” in unaffected relatives of individuals affected by Dravet syndrome and epilepsy and mental retardation limited to females [16]. Considering the potential harms, genetic testing should always be performed with the patient’s consent or parental consent in the case of minors. A team approach, including a genetic counselor, a psychologist, and a social worker, is recommended throughout the process of evaluation.In the last years a number of new molecular genetic technologies became available and they promise to change genetic testing for epilepsy, allowing to extend genetic analysis also to sporadic or nonfamilial cases. Two are the major new technologies that can affect the management of epileptic patients: Oligonucleotide Arrays Comparative Genomic Hybridization (Array-CGH) and Next-Generation Sequencing (NGS).


## 2. Molecular Karyotype

During the last 50 years cytogenetics has evolved from simple chromosome counting or banded chromosome morphological identification under light microscope to a molecular approach where chromosomes are analyzed through sophisticated computer system for their ability to hybridize to specific oligonucleotides spanning the entire genome [19]. Array-CGH is nowadays a basic diagnostic tool for clinical diagnosis of several types of developmental delays [20], intellectual disabilities [21, 22], and congenital abnormalities [23]. Epilepsy is also enjoying several advantages from the use of this technology that significantly improves diagnostic resolution of classic cytogenetics [24, 25].Chromosomes did not become individually identifiable before the discovery that several procedures could create reproducible, permanent, and specific banding patterns [26, 27]. This was fundamental for gene mapping and positional cloning of disease genes and also revealed a large number of rare and subtle pathological conditions that disturbed the normal band patterning of chromosomes. The improvements of high-resolution banding techniques allowed the identification of several subtle chromosomal abnormalities associated with epilepsy [14, 15]. The possibility to study these chromosomes regions with specific hybridization DNA probes through fluorescence in situ hybridization (FISH) greatly improved sensitivity to detect small chromosomal aberrations in specific regions [28].Today molecular karyotyping is rapidly replacing conventional cytogenetics and FISH. This name refers to the analysis of all chromosomes using hybridization to standard DNA sequences arranged on a “chip” rather than microscope observation. The technological development of this approach allows now clinicians to evaluate the entire genome for copy number variants (CNVs, duplications, deletions) in a single test. The high resolution of this approach is however limited by the difficulty to identify balanced chromosome translocations or inversions, even if this powerful technique recognizes in many of them microdeletions or cryptic anomalies at the chromosomal breakpoints. Detection of deletions or duplications is based on the comparison of two genomes (Figure1). Labelled patient DNA is cohybridized with control DNA to an array spotted with oligonucleotide DNA probes spanning the entire genome at critical intervals. The distance between these oligonucleotide sequences in the genome marks the resolution of the technique and can be as low as 1000 bp. The intensity of the signal from patient and control are then read and normalized by an electronic scanning device coupled with a software that generates a graphic plot of intensities for each probe.Figure 1A search of Online Mendelian Inheritance in Man (OMIM) clinical synopsis with the term “seizure” reveals that there are at least 754 mendelian disorders in which epilepsy is or can be part of the clinical condition, but not the main feature. Many of these disorders can be associated with DNA sequence mutations or subtle chromosomal anomalies that can be conveniently detected by array-CGH. Some will be private or sporadic cases and others will be familial. With the widespread use of CGH in both circumstances, many more genetic events will be reported in patients and the genetic aetiology will be recognized making possible over the time to saturate the genome with all possible loci and events that have an epileptogenic role.


## 3. Next-Generation Sequencing

From the publication of the draft of human genome sequence inNature, on 15th February 2001 issue, our view and knowledge of human genome has considerably changed [29] and the technologies to sequence DNA are today of common use in diagnostic practice and much cheaper. Chain termination or Sanger’s method [30] was largely used for the Human Genome Project and has dominated the past decades. The logic of this technology was to create by synthesis a population of DNA fragments of different size each one terminated at all possible positions by one of the four labelled dideoxynucleotides (ddNTPs) terminators. Separation of these fragments by polyacrylamide gel or capillary electrophoresis allowed the reading of the sequence through the first developed sequencing machines that could distinguish the fluorescence emitted by the blocking ddNTP [31]. Therefore, these are considered the “first generation” of DNA sequencing technologies.The need to reduce the cost of large sequencing projects has stimulated the development of a variety of cheaper sequencing technologies that are generally called “Next-Generation Sequencing” (NGS) [32, 33]. The final goal of this new field is to reduce the cost of human genome sequencing till or lower than $1,000 per genome to make it available for common medical practice and diagnostic use [34]. The development of further third-generation sequencing technologies should make possible to sequence single DNA molecules in real time with a cost that it is projected to be very close to the goal [35].The development of NGS platforms was a major progress in the technology because, differently from Sanger method, rather than producing about one thousand nucleotides for run, they are able to produce orders of magnitude more sequence data using massive parallel process, resulting in substantial increase of data at a lower cost per nucleotide [36, 37].Several commercial platforms are today available, including Roche/454 [38], Illumina/Solexa and Life Technologies/SOLiD (Table 8(a)). In very general terms these platforms follow similar process that includes: (a) template preparation by breaking large DNA macromolecule to generate short fragment libraries with platform-specific synthetic DNA adapters at the fragment ends, (b) massive and parallel clonal amplification of individual DNA fragment molecules on glass slide or microbeads by PCR [39] to generate a sufficient copy number of the labelled fragment to be detected by the machine optical system, and (c) sequencing by several cycles of extensions that are repeated and detected automatically to create short reads [40]. The data of these reads are then collected by the device, and the alignment of the short reads with specific software allows to rebuild the initial template sequence. Helicos and Pacific Biosystem platforms (Table 8(b)) are substantially different because they use a more advanced laser-based detection system that does not require massive parallel amplification with the considerable advantages to simplify preparation process, to eliminate PCR-induced bias and errors, and to make easy data collection. Ion Torrent developed an entirely new approach to sequencing based on hydrogen ion release when a nucleotide is incorporated into a DNA strand by polymerase (Table 8) [41]. An ion sensor can detect hydrogen ions and convert this ion chemical signal to digital sequence information eliminating the need of optical reading at each dNTP incorporation.Table 8
Comparison of commercially available sequencing platforms.
(a) Massive parallel clonal amplification with optical detectionRoche 454Life Technologies SOLiDIlluminaLibrary amplificationemPCR*emPCR*On glassSequencingIncorporation of unlabeled dNTPsLigase-mediated addition of fluorescent oligoNTPs (2bp)Incorporation of end-blocked fluorescent dNTPsDetectionLight emission from release of PPiFluorescence emission from ligated dye-labeled oligoNTPsFluorescence emission from incorporated labeled oligoNTPsProgressionUnlabeled dNTPS added in base-specific fashionChemical cleavage removes dye and oligoNTPChemical cleavage fluorescent dye and blocking groupErrorsInsertion/deletionEnd of readEnd of readLength400 bp75 bp150 bpOverall yield/run500 Mbp>100 Gbp200 Gbp(b) Fluorescent and semiconductor single molecule sequencingHelicosPacific biosystemIontorrentLibrary amplificationN/A-tSMS**N/A-SMRT*** sequencingOptional PCRSequencingIncorporation of fluorescent labeled dNTPsPolymerase incorporation terminal phosphate labeled dNTPsPolymerase incorporation of dNTPs releases H+DetectionLaser-induced emission from incorporated dNTPReal time detection of fluorescent dye in polymerase active siteSemiconductor ion sensor detects H+ released during dNTPs incorporationProgressionChemical cleavage of dNTP fluorescent groupN/A fluorescent dyes are removed as PPi with dNTPs incorporationH+ signal during each dNTP incorporation is converted in voltage signalErrorsInsertion/deletionInsertion/deletionInsertion/deletionLength35 bp1000 bp200–400 bpOverall yield/run21–37 Gbp>100 Gbp1 Gbp*emPCR (emulsion PCR) is an amplification method where DNA library fragments are mixed with beads and PCR reagents in an oil emulsion that allows massive amplification of bead-DNA in a single reaction.**tSMS: true Single Molecule Sequencing.***SMRT (Single Molecule Real Time)bp: base pair, Mbp: Mega base pair (106 bp), Gbp: Giga base pair (109 bp), dNTP: deoxynucleotide-tri-phosphate, PPi: pyrophosphate.Other third-generation platforms under development make use of nanophotonic visualization chamber, ion semiconductor, electron microscopy, a variety of nanotechnologies like nanopores (Oxford Nanopore Technologies), nanochannels (BioNanomatrix/nanoAnalyzer), nanoparticles (GE Global Research), nanoballs (Complete Genomics), nanowells (CrackerBio), nanoknifes (Renveo), and specially engineered sensor DNA polymerase (VisiGen Biotechnologies) [42]. They promise even larger and faster data production although they are still under development and a few years away from commercial use. In principle also DNA microarrays could allow sequencing by hybridization using ultrafast nonenzymatic methods (Genizon BioSciences) and somebody even suggests that mass spectrometry might be used to determine mass differences between DNA fragments produced by chain termination [43].The beginning of several individual genome projects has gradually decreased the cost of sequencing an individual genome, and it is likely that the $1,000 cost per person will be reached in few years. In medicine, the “personal genome” age made possible by NGS will be an important milestone for the entire genomic field and will mark a transition from single gene testing to whole genome evaluation [44].It is impossible to predict today which NGS will eventually dominate genomic research, but it is sure that cost reductions, sequencing speed, and better accuracy will make NGS an essential molecular tool in several areas of biology and medicine.Although the cost of whole genome sequencing has dropped significantly, it remains a major obstacle since it can reach $100,000 for a single individual. However, targeting sequencing of specific regions of interest can decrease the overall cost and improve efficiency of NGS making this technology ready for diagnostic use [45].Also the field of epilepsy is potentially affected by NGS. Indeed too many genes and genetic conditions can be associated with epilepsy to make impossible for the clinicians a general use of specific monogene test for the vast majority of nonsyndromic or idiopathic epileptic patients. NGS is changing this situation by targeting several genome regions where known epilepsy genes are located and using enrichment techniques to significantly reduce the cost and improve efficiency. Targeted sequencing usually tests all protein-coding exons (functional exosome) which only requires roughly 5% as much sequencing than whole genome. This strategy will reduces the cost to about $3,000 or even less per single individual. Targeted selection technologies have been marketed and successfully used in different NGS projects and are becoming the tool of choice in several conditions, including epilepsy [46].


## 4. Targeting Sequencing and Epilepsy Gene Panels

A diagnostic panel is the contemporaneous targeted sequencing of a number of known genes that have already been identified as cause of a particular disease. A diagnostic panel is very different from whole genome or exome sequencing. Only genes clearly associated with a disease are examined. The genes included in the panel can be decided by the prescribing physician or by ad hoc committees of experts that can reach a consensus on the number and type of genes to test making commercially available diagnostic panel kits for specific diseases. This strategy should make easier to detect genetic variants that after validation by Sanger sequencing can be interpreted as the cause of the disease. Of course diagnostic panels and targeted sequencing make sense only if the condition is caused by several or very large number of genes. Many genetic disorders fall in this condition and are excellent candidate for the development of diagnostic panels. Epilepsy is an excellent example of such situation since it is a relative frequent disease affecting 1% of the population in a variety of forms, at different ages, with different progression. A genetic cause of epilepsy can be reasonably supposed in sporadic cases if trauma, tumor, or infection can be ruled out. In such circumstance all genetic information about epilepsy genes identified over the years in familial cases can be used to identify the causative gene through an epilepsy diagnostic panel. Indeed in the case of epilepsy the identified genes are so many that they can be classified in subpanels of genes that underline a common clinical entity (Table9). Clinical considerations may suggest the clinician to include in a diagnostic panel other genes or genes from different sub-panels. At the present the first diagnostic panels for epilepsy that can analyze up to 400 different genes (CeGaT GmbH) are commercially available [47].Table 9
Epilepsy diagnostic panels.
Subpanels With Homogeneous Clinical EntitiesTableNumber of genesMyoclonic epilepsy, febrile seizures, absences137Encephalopathies330X-linked mental retardation (XLMR)4(a)25Joubert syndrome4(b)10Lissencephaly and polymicrogyria4(c)18Severe Microcephaly and pontocerebellar hypoplasia4(d)22Walker-Warburg syndrome4(e)6Holoprosencephaly4(f)8Leukodystrophies5(a)20Migraine5(b)6Disorders of the Ras-MAPK pathway5(c)13Hyperekplexia for defective glycine neurotransmission5(d)5Neuronal migration disorders5(e)31Inherited errors of metabolism649Congenital disorder of glycosylation (CDG)7(a)23Neuronal ceroid lipofuscinosis (NCL)7(b)8Defects of mitochondrial metabolism including coenzyme Q deficiency7(c)35Mucopolisaccaridosis and mucolipidosis7(d)15Peroxisome biogenesis disorders (PBD)7(e)9Syndromic epilepsy247


## 5. Future Perspectives and Conclusions

The genetics of epilepsy has evolved from ion channel and neurotransmitter receptor subunits to newly discovered genes highlighting the importance of different pathways in the epileptogenesis. Furthermore, it has been demonstrated that copy number variations collectively explain a larger portion of idiopathic epilepsy than any single gene. These studies have identified structural genomic variations associated with idiopathic epilepsy, representing a change from the conventional knowledge that chromosome microarray analysis is useful only for patients with intellectual disability or dysmorphism [48]. Genetic testing techniques are rapidly evolving and whole exome or whole genome sequencing, performing at increasingly cheaper costs, will allow rapid discovery of other pathogenic mutations, variants in noncoding DNA, and copy number variations encompassing several genes. This rapidly accumulating genetic information will expand our understanding of epilepsy, and will allow more rational and effective treatment. However, along with the ability to identify genetic variants potentially associated with epilepsy it is imperative to validate genetic associations and analyze their clinical significance.Is it worth? The main objective of a diagnostic panel for epilepsy is to discover the molecular defect in all possible cases to create a specific and personalized treatment of the disease than can be pharmacologically different for different types of molecular defects. Personalized therapy will be possible only within a genomic medicine. But genomic medicine at the same time will raise other questions: what to do if more than one genetic variant is identified in the same epileptic patient? Can we understand how genetic interactions will modulate the disease severity and prognosis? Can interaction of specific genetic variants and environmental factors modulate the clinical spectrum of the disorder? What to do if the diagnostic panel is inconclusive? Are the costs affordable? These are only few questions that the genomics of epilepsy will raise. The answers will require time, a lot of sequencing, and probably the development of new and cheaper sequencing technologies.



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