Pediatric 46 (2012) 24e31

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Pediatric Neurology

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Review Article Genes of Early-Onset Epileptic : From Genotype to Phenotype

Mario Mastrangelo MD, Vincenzo Leuzzi MD *

Division of Child Neurology, Department of Pediatrics, Child Neurology, and Psychiatry, Sapienza University of Rome, Rome, Italy article information abstract

Article history: Early-onset epileptic encephalopathies are severe disorders in which cognitive, sensory, and motor Received 26 July 2011 development is impaired by recurrent clinical or prominent interictal epileptiform discharges Accepted 24 October 2011 during the neonatal or early infantile periods. They include , early myoclonic epileptic , West syndrome, , and other diseases, e.g., X-linked myoclonic seizures, spasticity and syndrome, idiopathic infantile epileptic-dyskinetic encephalopathy, and mental retardation limited to females, and severe infantile multifocal epilepsy. We summarize recent updates on the genes and related clinical syndromes involved in the pathogenesis of early-onset epileptic encephalopathies: Aristaless-related homeobox (ARX), cyclin- dependent kinase-like 5 (CDKL5), syntaxin-binding protein 1 (STXBP1), solute carrier family 25 member 22 (SLC25A22), nonerythrocytic a-spectrin-1 (SPTAN1), phospholipase Cb1(PLCb1), membrane- associated guanylate kinase inverted-2 (MAGI2), polynucleotide kinase 30-phosphatase (PNKP), sodium channel neuronal type 1a subunit (SCN1A), protocadherin 19 (PCDH19), and pyridoxamine 5-prime- phosphate oxidase (PNPO). Ó 2012 Published by Elsevier Inc.

Introduction preserved domain, octapeptide and acidic domains, four poly- alanine tracts, and three nuclear localization sequence motifs [4]. Early-onset epileptic encephalopathies are severe disorders in The Aristaless-related homeobox gene acts as both a transcrip- which cognitive, sensory, and motor development is impaired by tional repressor and activator in an incompletely characterized recurrent clinical seizures or prominent interictal epileptiform biochemical cascade, and modulates cerebral development and discharges during the neonatal or the early infantile periods [1]. patterning through the regulation of differentiation, proliferation, Several studies elucidated the pathogenic role of genetic mutations and tangential migration of neuronal precursors and cortical involved in the synaptogenesis, pruning, neuronal migration and interneurons [5,6]. differentiation, synthesis and release, structures, To date, 44 different pathogenic mutations of the Aristaless- and functions of membrane receptors and transporters [2,3]. related homeobox gene in 100 families have been associated with We review the genes more frequently associated with early- two groups of human syndromes exhibiting highly variable onset epileptic encephalopathies and their associated phenotypes phenotypes, i.e., syndromes with and without malformations [7]. (Table 1). The first group includes X-linked or hydranence- phaly, associated with abnormal genitalia (Online Mendelian Inheritance in Man number 300215), and Proud syndrome (Online Aristaless-Related Homeobox Gene (ARX, Online Mendelian Mendelian Inheritance in Man number 300004). Syndromes Inheritance in Man Number 300382) without malformations include idiopathic Ohtahara syndrome, idiopathic West syndrome, X-linked myoclonic seizures, spasticity The Aristaless-related homeobox gene maps to Xp22.13 and and intellectual disability, and idiopathic infantile epileptic- includes five coding exons. The Aristaless-related homeobox dyskinetic encephalopathy (Online Mendelian Inheritance in Man protein contains the paired/Q50 homeodomain, the Aristaless number 308350 for these four diseases), nonsyndromic X-linked mental retardation (Online Mendelian Inheritance in Man number 300419), and Partington syndrome (Online Mendelian Inheritance * Communications should be addressed to: Dr. Leuzzi; Division of Child in Man number 309510). Premature termination or null mutations Neurology; Department of Pediatrics, Child Neurology, and Psychiatry; Sapienza University of Rome; Via dei Sabelli 108; 00185 Rome, Italy. are mainly related to malformations (e.g., lissencephaly, E-mail address: [email protected] hydranencephaly, and agenesis of the corpus callosum), whereas

0887-8994/$ - see front matter Ó 2012 Published by Elsevier Inc. doi:10.1016/j.pediatrneurol.2011.11.003 M. Mastrangelo, V. Leuzzi / Pediatric Neurology 46 (2012) 24e31 25 expansion of the polyalanine tracts is linked to syndromes without More recently, Poirier et al. described a relationship between an malformations and with early epileptic encephalopathies [8,9]. expansion of the trinucleotide repeat GCG in the Aristaless-related Expansions of the polyalanine tracts represent the most homeobox gene and the presence of infantile spasms, atypical common and well-studied result of Aristaless-related homeobox , and dyskinetic movements in a 4-year-old boy gene pathologic mutations [10]. Among these, a 24-bp duplication [20]. mutation (c.429-452dup) accounts for 45% of all published cases [10-12]. Nasrallah et al. demonstrated that in these contexts, an Aristaless-related homeobox gene and Ohtahara syndrome improper folding of the Aristaless-related homeobox mutated protein produces intranuclear inclusions resulting in an increased Kato et al. [21] demonstrated a relationship between Aristaless- neuronal cell-death [10]. The pathogenic mechanism of Aristaless- related homeobox gene mutations and Ohtahara syndrome related homeobox gene-related epileptic encephalopathies prob- through the definition of a hemizygous, de novo 33-bp duplication ably involves the subsequent loss of function of cortical GABAergic in exon 2, 298-330dupGCGGCA(GCG)9, in two of three unrelated interneurons, resulting in overall increased neuronal excitability male patients. This mutation expanded the original 16 alanine [7,11,12]. residues to 27 alanine residues in the first polyalanine tract of the Aristaless-related homeobox protein. The authors suggested Aristaless-related homeobox gene, infantile spasms, and West a possible link between the longer expansion of the polyalanine syndrome (early infantile epileptic encephalopathy 1) tract in Ohtahara syndrome and its earlier onset and more severe phenotype than in West syndrome [21]. Strømme et al. [13] demonstrated the pathogenic role of the Absoud et al. [22] described a rapidly progressive neuro- Aristaless-related homeobox gene in an X-linked subtype of West degeneration resulting in death before age 1 year in a boy with syndrome. They reported four mutations in seven families: a GCG Ohtahara syndrome, a dyskinetic movement disorder, and a shorter expansion and the 24-bp duplication (c.429-452dup), resulting in expansion of seven alanine residues in the Aristaless-related an expansion of the polyalanine tracts of the Aristaless-related homeobox gene. This fatal course is not compatible with the homeobox gene, a missense mutation within the Aristaless- hypothesis of a correlation between the expansion length of poly- related homeobox gene homeodomain (P353L), and a truncation alanine tracts and clinical severity [22]. mutation [13]. Kato et al. also observed polyalanine expansions in Fullston et al. described a single Aristaless-related homeobox a patient with sporadic cryptogenic West syndrome [14]. protein truncation mutation (c.81C>G /p.Y27X) in two cousins with Subsequent observations by Strømme et al. [15] and Wohlrab different clinical onsets: one with Ohtahara syndrome, and one et al. [16] indicated that the clinical severity of epileptic (infantile with West syndrome [23]. spasms and myoclonic seizures) and nonepileptic (mental retar- More recently, Kato et al. [24] reported on two novel frameshift dation, , , or ) manifestations is greater when mutations in the terminal exon of the Aristaless-related homeobox the expansion of alanine residues is located in the first polyalanine gene (Ala524fsX534 and E536fsX672) in two members of a family tract. with six males affected by Ohtahara syndrome. The clinical Reish et al. [17] reported on a de novo familial duplication presentation appeared less severe in these patients than in patients mutation of 27 bp, c.430-456dup, in exon 2 of the Aristaless-related with a 33-bp expansion in the first polyalanine tract (minor homeobox gene (the same locus as the more frequent 24-bp abnormalities of the genitalia and of brain structure, and delayed duplication, c.429-452dup) in three affected males because of onset of seizures) [24]. a postzygotic mosaicism, and suggested that the related reduction of the second polyalanine tract of the Aristaless-related homeobox Cyclin-Dependent Kinase-Like 5 (CDKL5, Online Mendelian gene beyond its natural limit of 20 amino acids resulted in a nega- Inheritance in Man Number 300203) tive prognosis [17]. The cyclin-dependent kinase-like 5 gene maps to Xp22 and Aristaless-related homeobox gene and X-linked myoclonic seizures, contains 20 coding exons. The related protein is a large, serine- spasticity, and intellectual disability syndrome threonine kinase of 1030 amino acids, including a preserved serine-threonine kinase domain in the N-terminal, and a C-terminal Scheffer et al. defined the features of “X-linked myoclonic zone that regulates its catalytic activity and nuclear placement [25]. seizures, spasticity, and intellectual disability syndrome” through Cyclin-dependent kinase-like 5 is part of an incompletely linkage studies and mutational analyses of six boys with generalized characterized cascade. A recent study seems to rule out a previous spasticity, , and intellectual impairment. X-linked hypothesis that cyclin-dependent kinase-like 5 protein could be myoclonic seizures, spasticity, and intellectual disability syndrome involved in the same molecular pathways as the methyl-cytosine was mainly related to a missense mutation (1058C>T/P353L) with- phosphate guanine-binding protein 2 (MeCP2) gene [26,27]. in the homeodomain of the Aristaless-related homeobox gene, and To the best of our knowledge, 53 pathologic mutations of cyclin- an X-linked recessive transmission was observed. The obligate dependent kinase-like 5 (47 in females, and six in males) have been female typically presented osteotendineous hyperreflexia [18]. reported: 12 missense, four nonsense, eight splice-site, 22 deletion, and seven frameshift mutations [28-30]. No correlations between Aristaless-related homeobox gene and idiopathic infantile epileptic- clinical severity and sites of the mutations have been demonstrated dyskinetic encephalopathy [29,30]. A severe epileptic encephalopathy (early infantile epileptic Guerrini et al. [19] reported on a new clinical entity, “idiopathic encephalopathy 2; Online Mendelian Inheritance in Man number infantile epileptic-dyskinetic encephalopathy,” with infantile 300672) that is characterized by intractable seizures with infantile spasms, mental retardation, and generalized dystonia leading to spasms, mental retardation, and a Rett-like phenotype, is attrib- severe quadriplegic dyskinesia. They defined an expansion from 10 utable to cyclin-dependent kinase-like 5 mutations [31-34]. Bahi- to 17 GCG repeats (c.333-334 ins[GCG]7) in the Aristaless-related Buisson et al. [35] defined three clinical stages of epilepsy in 13 homeobox gene sequences encoding for the first polyalanine tract patients with early infantile epileptic encephalopathy 2 and cyclin- in six boys from four families [19]. dependent kinase-like 5 mutations: (1) a first stage with early- 26 M. Mastrangelo, V. Leuzzi / Pediatric Neurology 46 (2012) 24e31

Table 1. Genes and related epileptic encephalopathies that are discussed in this review

Gene Locus Protein Function Epileptic Encephalopathy Clinical Features Available Gene Test* Aristaless-related Xp22.13 Transcriptional repressor X-linked infantile spasms/ Infantile spasms Sequence analysis of the homeobox gene (ARX, and activator West syndrome or early Mental retardation entire coding region OMIM number 300382) infantile epileptic Hypsarrhythmia Targeted mutation analysis encephalopathy 1 (OMIM Deletion/duplication number 308350) analysis X-linked myoclonic Generalized spasticity Mutation scanning of select seizures, spasticity, and Myoclonic epilepsy exons intellectual disability Intellectual impairment Prenatal diagnosis syndrome (OMIM number Carrier testing 308350) Idiopathic infantile Infantile spasms epileptic-dyskinetic Mental retardation encephalopathy (OMIM Dyskinetic movements number 308350) Generalized dystonia Ohtahara syndrome (OMIM Tonic spasms and other number 308350) seizures Neurocognitive impairment Suppression burst on EEG Cyclin-dependent kinase- Xp22 Serine-threonine kinase Early infantile epileptic Infantile spasms Sequence analysis of the like 5 (CDKL5,OMIM encephalopathy 2 (OMIM Mental retardation and entire coding region number 300203) number 300672) severe motor impairment FISH-metaphase Hypotonia Deletion/duplication Poor eye contact analysis Rett-like phenotype Prenatal diagnosis (secondary deceleration of Carrier testing head growth, sleep disturbances, hand apraxia, and stereotypies) Solute carrier family 25, 11p15.5 Mithocondrial glutamate/ Ohtahara syndrome (OMIM Myoclonic seizures Sequence analysis of the þ member 22 (SLC25A22, H symporter number 308350) Hypotonia entire coding region OMIM number 609302) Early infantile epileptic Deletion/duplication encephalopathy 3 (OMIM Suppression burst pattern analysis number 609304) on EEG Prenatal diagnosis Abnormal Carrier testing electroretinogram Syntaxin binding protein 1 9q341 Modulator of synaptic Ohtahara syndrome (OMIM Tonic spasms or tonic- Sequence analysis of the (STXBP1, OMIM number vesicle release number 308350) clonic seizures entire coding region 602926) Early infantile epileptic Mental retardation Deletion/duplication encephalopathy 4 (OMIM Hypotonia analysis number 612164) Suppression-burst on EEG Prenatal diagnosis Nonerythrocytic a- 9q33-q34 Cytoskeletal protein Early infantile epileptic Infantile spasms with Sequence analysis of the spectrin-1 (SPTAN1, encephalopathy 5 (OMIM hypsarrhythmia entire coding region OMIM number 182810) number 613477) Generalized seizures Prenatal diagnosis Mental retardation Spastic quadriplegia Progressive microcephaly Hypomyelination and diffuse brain atrophy on MRI Phospholipase Cb1(PLCb1, 20p12.3 Enzyme involved in cellular Ohtahara syndrome (OMIM Tonic seizures Sequence analysis of the OMIM number 607120) signalling number 308350) Infantile spasms entire coding region (not West syndrome, or early commercially available) infantile epileptic Deletion/duplication encephalopathy 1 (OMIM analysis (not commercially number 308350) available) Membrane-associated 7q11.23-q21.1 Synaptic scaffolding protein West syndrome, or early Infantile spasms Sequence analysis of the guanylate kinase infantile epileptic Mental retardation entire coding region (not inverted-2 (MAGI2, encephalopathy 1 (OMIM commercially available) OMIM number 606382) number 308350) Deletion/duplication analysis (not commercially available) Polynucleotide kinase 30- 19q13.33 Enzyme involved in DNA Ohtahara syndrome (OMIM Polymorphic seizures Sequence analysis of the phosphatase (PNKP, repair number 308350) Microcephaly entire coding region OMIM number 605610) West syndrome, or early Developmental delay Linkage analysis infantile epileptic Behavioral disorders Prenatal diagnosis encephalopathy 1 (OMIM Carrier testing number 308350) Sodium channel neuronal 2q24.3 Subunit of a voltage-gated Dravet syndrome (OMIM Febrile or afebrile seizures Sequence analysis of the type 1a subunit (SCN1A, sodium channel number 607208) Generalized or unilateral entire coding region OMIM number 182389) clonic seizures Mutation scanning of the Myoclonic seizures entire coding region Atypical absences Linkage analysis Partial seizures FISH-metaphase Mental retardation Deletion/duplication Motor impairment analysis Prenatal diagnosis M. Mastrangelo, V. Leuzzi / Pediatric Neurology 46 (2012) 24e31 27

Table 1 (continued )

Gene Locus Protein Function Epileptic Encephalopathy Clinical Features Available Gene Test* Protocadherin 19 (PCDH19, Xq22 Adhesion protein Epilepsy and mental Febrile and afebrile seizures Sequence analysis of the OMIM number 300460) retardation limited to Rare myoclonic jerks and entire coding region females (OMIM number atypical absences Sequence analysis of select 300088) Mental retardation exons Dravet syndrome (OMIM Motor impairment Deletion/duplication number 607208) analysis Prenatal diagnosis Carrier testing Pyridoxamine 5-prime- 17q21.32 Enzyme in pyridoxine PNPO deficiency (OMIM Parental consanguinity Sequence analysis of the phosphate oxidase activation cascade number 610090) Low Apgar scores entire coding region (PNPO, OMIM number Perinatal respiratory Analyte 603287) distress Prenatal diagnosis Pyridoxine-unresponsive Carrier testing seizures Suppression burst pattern on EEG Abbreviations: EEG ¼ Electroencephalogram FISH ¼ Fluorescence in situ hybridization MRI ¼ Magnetic resonance imaging OMIM ¼ Online Mendelian Inheritance in Man * Laboratories offering the reported gene testing are listed at the website www.genereviews.org.

onset, recurrent convulsive seizures, severe hypotonia, and normal a missense mutation (p.Pro206Leu) in the SLC25A22 gene in four findings of interictal electroencephalograms; (2) a second stage consanguineous Arab infants with early myoclonic epilepsy (early involving epileptic encephalopathy with infantile spasms and infantile epileptic encephalopathy 3, Online Mendelian Inheritance hypsarrhythmia; and (3) a third stage with refractory tonic or in Man number 609304). In the same study, the authors did not myoclonic epilepsy. The authors observed that progression to the identify any mutation in the coding zone of the solute carrier family third stage was more frequent and occurred more quickly in 25, member 22 gene in 25 unrelated children with early myoclonic patients with truncation mutations involving the catalytic domain encephalopathy and progressive microcephaly. They also demon- [35]. strated the prominent expression of solute carrier family 25, The same authors expanded the description of clinical features member 22 in cerebral regions involved in the pathogenesis of of cyclin-dependent kinase-like 5-related encephalopathy in myoclonic seizures (red nuclei, substantia nigra, and olivocerebellar a larger series of 20 affected girls with early-onset seizures, severe networks). Moreover, they speculated that an epileptic phenotype hypotonia, poor eye contact, and some Rett-like signs (secondary in patients carrying solute carrier family 25, member 22 mutations deceleration of head growth, severe motor impairment, sleep could be attributable to abnormal glutamate synthesis and turn- disturbances, hand apraxia, and stereotypies). Cranial magnetic over in the cytoplasm of both astrocytes and neurons, to resonance imaging of these patients indicated diffuse cortical a secondary impairment of the mitochondrial respiratory chain, or atrophy associated with hyperintensities in the white matter of the to alterations of neuronal oxygen handling [40]. temporal lobe as a possible diagnostic marker, without prognostic The same group described a novel homozygous solute carrier significance, in this disease. The authors speculated that the vari- family 25, member 22 mutation (p.G236W) in the son of ability of clinical presentations in cyclin-dependent kinase-like consanguineous Algerian parents, and tested the total loss of 5-related encephalopathy resulted from the transcriptional or activity of the mutated protein in vitro. The proband manifested translational effects of cyclin-dependent kinase-like 5 mutations. severe myoclonic epileptic encephalopathy beginning in the first No links were evident concerning the nature and site of mutations days after birth, with hypotonia and microcephaly, a suppression- and the X-chromosome inactivation pattern of patients [36]. burst pattern on electroencephalogram, and an abnormal elec- Various nonspecific electroencephalographic abnormalities troretinogram [41]. (hypsarrhythmia, focal or multifocal activity, diffuse high-voltage sharp waves, and generalized q rhythm) were reported in chil- dren carrying cyclin-dependent kinase-like 5 mutations, whereas Syntaxin Binding Protein 1 (STXBP1, Online Mendelian Pintaudi et al. observed a presumably much more specific pseudo- Inheritance in Man Number 602926) periodic trace in two girls [37]. Melani et al. reported on three electroclinical stages in six children less than 1 year of age: tonic- The syntaxin binding protein 1 (STXBP1,orMunc18) gene maps vibratory contractions associated with an electrodecremental to 9q341, and includes 20 exons. Syntaxin binding protein 1 event, a clonic phase related to an irregular series of sharp waves modulates the release of synaptic vesicles through specific inter- and spike slow waves, and final myoclonic jerks, linked with actions with syntaxin A (Stx1a) and with the soluble N-ethyl- bilateral, rhythmic sharp waves [38]. maleimide-sensitive factor attachment protein receptor complex. An open conformation of syntaxin 1A that promotes the formation of the soluble N-ethylmaleimide-sensitive factor attachment Solute Carrier Family 25, Member 22 (SLC25A22, Online protein receptor complex and subsequent vesicular release, and Mendelian Inheritance in Man Number 609302) a closed conformation of syntaxin 1A that controls synaptic vesic- ular docking, are involved in these processes [42]. The solute carrier family 25, member 22 gene maps to 11p15.5, Saitsu et al. reported on a de novo 2.0-Mb microdeletion and contains nine encoding exons, and encodes for a mitochondrial four heterozygous missense mutations within the syntaxin binding þ glutamate/H symporter [39]. Molinari et al. [40] identified protein 1 gene in five patients (two females and three males) with 28 M. Mastrangelo, V. Leuzzi / Pediatric Neurology 46 (2012) 24e31

Ohtahara syndrome (early infantile epileptic encephalopathy 4, described a homozygous loss-of-function 0.5-Mb deletion, Online Mendelian Inheritance in Man number 612164) [43]. Ham- including the promoter and exons 1, 2, and 3 of phospholipase Cb1, dan et al. described two de novo truncating mutations in the region in a single male infant who developed tonic seizures and then of binding with syntaxin 1A in two unrelated French-Canadian infantile spasms. The same authors excluded a linkage to the patients with early-onset seizures, mental retardation, hypotonia, phospholipase C b1 gene in 12 consanguineous families of other abnormal gait, and tremor [44]. All mutations in both these reports children with infantile spasms [50]. were located in the hydrophobic center of the protein, and probably resulted in its misfolding. All the reported patients presented tonic Membrane-Associated Guanylate Kinase Inverted-2 Gene spasms or tonic-clonic seizures before age 3 months, mental (MAGI2, Online Mendelian Inheritance in Man Number retardation, hypotonia, a suppression-burst pattern on electroen- 606382) cephalograms, and hypomyelination according to cranial magnetic fi resonance imaging [43,44]. More recently, Deprez et al. identi ed Membrane-associated guanylate kinase inverted-2 maps to four new truncating mutations and two microdeletions that abol- 7q11.23-q21.1, and encodes for a scaffolding enzyme interacting ished syntaxin binding protein 1 gene function in six of 106 with different presynaptic and postsynaptic receptors (including patients. None of the patients manifested a phenotype compatible the N-methyl-D-aspartic acid receptor) [51]. with Ohtahara syndrome (one had West syndrome, whereas the Marshall et al. described a hemizygous 1.4-Mb deletion fi others did not manifest a speci c recognized epileptic syndrome), encompassing membrane-associated guanylate kinase inverted-2 whereas profound mental retardation, ataxia, and dyskinetic gene in 15 of 16 patients with infantile spasms, whereas the same fi movements were observed in ve subjects [45]. Seven novel mutation was absent in 11 of 12 controls without a history of mutations (two frameshift mutations, three nonsense mutations, seizures [51]. one splicing mutation, and one missense mutation) were discov- Röthlisberger et al. recently reported that in children with ered by Saitsu et al. [46] in six males and three females belonging to infantile spasms, membrane-associated guanylate kinase inverted- a larger cohort, including 29 patients with Ohtahara syndrome. The 2 is not necessarily involved in the hemizygous 1.4-Mb deletion, same mutations were absent in 54 patients with West syndrome. and thus further studies are required to clarify this finding [52]. The same study demonstrated that the reported mutations produced nonsense-mediated mitochondrial RNA decay in vitro, 0 PNKP associated with an abnormal splicing and subsequent degradation Polynucleotide Kinase 3 -Phosphatase ( , Online of the resultant protein [46]. Mendelian Inheritance in Man Number 605610)

In a cohort study of 86 patients with epileptic encephalopathies, 0 Otsuka et al. defined a novel missense mutation (c.1654T>C) in the The polynucleotide kinase 3 -phosphatase gene encodes for an syntaxin binding protein 1gene in a child with West syndrome and enzyme that is involved in DNA repair networks and maps to without previous Ohtahara syndrome. The same mutation was 19q13.33. Through a genome-wide linkage analysis in seven evident in a 2-year-old boy with classic Ohtahara syndrome [47]. consanguineous families, Shen et al. associated mutations in this gene with an autosomal recessive syndrome involving early-onset, Nonerythrocytic a-Spectrin-1 (SPTAN1, Online Mendelian drug-resistant seizures, microcephaly, developmental delay, and Inheritance in Man Number 182810) behavioral disorders [53].

The nonerythrocytic a-spectrin-1 gene maps to 9q33-q34, and Sodium Channel Neuronal Type 1a Subunit (SCN1A, Online encodes for a filamentous cytoskeletal protein that regulates the Mendelian Inheritance in Man Number 182389) stability of axonal structure [48]. Saitsu et al. [48] recently described de novo heterozygous Voltage-gated sodium channels present a large, multimeric a- nonerythrocytic a-spectrin-1 gene mutations in three unrelated subunit and two smaller, auxiliary b-subunits. The sodium channel patients, previously reported by Tohyama et al. [49], with drug- neuronal type 1a subunit belongs to a family of nine genes resistant seizures, hypsarrhythmia, mental retardation, spastic encoding for a subunits. The sodium channel neuronal type 1a quadriplegia, and progressive microcephaly (early infantile subunit maps to 2q24.3, and is expressed in nine isoforms (Nav1.1- epileptic encephalopathy 5, Online Mendelian Inheritance in Man Nav1.9) [54]. number 613477). Magnetic resonance imaging indicated hypo- More than 600 sodium channel neuronal type 1a subunit myelination and diffuse brain atrophy [48,49]. One patient mutations have been discovered [55]. In sodium voltage channels, demonstrated a 2.25-Mb microdeletion, encompassing both the they can promote both a gain of function, resulting in increased syntaxin binding protein 1 and nonerythrocytic a-spectrin-1 genes, neuronal susceptibility, and a loss of function, with subse- that impaired spectrin heterodimer stability. That patient man- quent decreased inhibitory activity of GABAergic interneurons [56]. ifested a milder phenotype, with controlled seizures and no Two epileptic syndromes have been linked to sodium channel significant structural brain anomalies. The authors speculated that neuronal type 1a subunit mutations: genetic epilepsy with febrile this phenotype could result from syntaxin binding protein 1 hap- seizure plus (previously known as “ with loinsufficiency, whereas the effects on myelination were attribut- plus”) and Dravet syndrome (this eponym assembles able to the haploinsufficiency of nonerythrocytic a-spectrin-1 “severe myoclonic epilepsy of infancy” and “severe myoclonic [43,46,48]. epilepsy of infancy-borderline”) [57]. Generalized epilepsy with febrile seizure plus is an autosomal Phospholipase Cb1(PLCb1, Online Mendelian Inheritance in dominant disorder characterized by febrile seizures occurring Man Number 607120) beyond their common ages and subsequent generalized or partial . Mutations of the sodium channel neuronal type 1a The phospholipase Cb1(PLCb1) gene maps to 20p12.3, and subunit account for the 10% of patients with generalized epilepsy encodes for an enzyme that is involved in cellular signaling through with febrile seizure plus. Patients who manifest generalized the production of inositol 1,4,5 triphosphate and diacylglycerol epilepsy with febrile seizure plus present with milder clinical from phosphatidylinositol 4,5-bisphosphate. Kurian et al. [50] severity and usually normal psychomotor development [58]. M. Mastrangelo, V. Leuzzi / Pediatric Neurology 46 (2012) 24e31 29

Dravet syndrome (Online Mendelian Inheritance in Man channel neuronal type 1a subunit gene-positive form because of number 607208) is an early-onset, intractable epileptic encepha- minor variability in seizures (myoclonic jerks and atypical absences lopathy with polymorphic clinical manifestations, including pro- are relatively rare) [69]. longed generalized or unilateral clonic seizures triggered by fever, More recently, Marini et al. [70] reported on various mutations photo stimulation, or hot water, myoclonic seizures, atypical in 13 probands (seven with a clinical diagnosis of Dravet syndrome, absences, and partial seizures. Developmental milestones are and six with focal epilepsy) in a cohort of 117 females with both usually normal before the onset of seizures, but are gradually febrile seizures and different epileptic phenotypes. Eleven of these impaired by recurrent epileptic episodes, resulting in mental delay, 13 patients presented with mental retardation. Mutation types spasticity, or ataxia [59]. were variable (six missense, 2two truncating, three frameshift, and About 85% of patients with Dravet syndrome manifest sodium two splicing), and a familial transmission was demonstrated in channel neuronal type 1a subunit mutations. In contrast with three cases [70]. generalized epilepsy with febrile seizure plus, mutations of the Depienne et al. also demonstrated that protocadherin 19 sodium channel neuronal type 1a subunit in Dravet syndrome are mutations can be found in epileptic families without an associated mainly de novo, and more commonly arise from the paternal history of mental retardation. In the same study, based on a cohort chromosome [60]. Neurologic impairment is generally greater with of 150 patients, a strong sensitivity to fever of the affected subjects truncation mutations than with missense/nonsense or frameshift and 15 novel point mutations and three deletions were reported mutations, but impairment can be also influenced by mutations in [71]. modulator genes other than the sodium channel neuronal type 1a subunit (“genetic modifiers”) [61]. Martin et al. [62], Ohmori et al. Pyridoxamine 5-Prime-Phosphate Oxidase (PNPO, Online [63], and Singh et al. [64] suggested the sodium channel neuronal Mendelian Inheritance in Man Number 603287) type 8a subunit (SCN8A), the calcium channel voltage-dependent b4 subunit (CACNB4), and the sodium channel neuronal type 9a Pyridoxamine 5-prime-phosphate oxidase is an enzyme that subunit (SCN9A) as possible candidates for this role [62-64]. Harkin produces pyridoxal 5-prime-phosphate in the pathway of activa- et al. [65] enlarged the spectrum of clinical phenotypes of sodium tion of pyridoxine (an important cofactor in neurotransmitter channel neuronal type 1a subunit gene mutations to cryptogenic synthesis). Its encoding gene maps to 17q21.32, and contains seven generalized and focal epilepsies other than Dravet syndrome in exons [72]. a study of 188 patients with seizures occurring before age 2 years. Mills et al. [72] defined homozygous missense, splice site, and They also defined a new “severe infantile multifocal epilepsy” that stop codon pyridoxamine 5-prime-phosphate oxidase gene muta- was distinguished from Dravet syndrome by a later onset of tions in five preterm infants from three families with parental cognitive decline, a lack of absence and myoclonic seizures, and consanguinity, low Apgar scores, perinatal respiratory distress a lack of generalized spike and wave activity on electroencepha- resulting in intubation, pyridoxine-unresponsive seizures in the lograms [65]. first hours after birth, and suppression bursts on electroencepha- lograms. All but one patient studied by Mills et al. [72] died during Protocadherin 19 (PCDH19, Online Mendelian Inheritance in the neonatal period. The only survivor, who was treated with Man Number 300460) pyridoxal 5-prime-phosphate, manifested marked acquired microcephaly, severe developmental delay, central hypotonia, The protocadherin 19 gene maps to Xq22, and encodes for painful dystonic spasms, and persistent seizures by age 2 years [72]. a transmembrane protein that controls calcium-dependent cell-cell adhesion. Protocadherin 19 may be involved in specific synaptic connections and transmissions [66]. The Role of Genetic Workups in the Diagnostic Approach to Dibbens et al. reported on protocadherin 19 mutations in seven Early-Onset Epileptic Encephalopathies families with “epilepsy and mental retardation limited to females” (Online Mendelian Inheritance in Man number 300088) [67]. Hines The diagnostic workup of early-onset epileptic encephalopa- et al. described two protocadherin 19 mutations in three cases of thies remains a challenge because of frequent difficulties in female mental retardation, whereas they excluded a pathogenic defining etiologies. Acquired structural abnormalities, such as role of protocadherin 19 in and autistic spectrum hypoxic-ischemic insults and isolated cortical malformations, disorder [68]. represent the most common causes of epileptic encephalopathy in Epilepsy and mental retardation limited to females is charac- infancy, and they should be excluded in the first diagnostic steps terized by an onset of seizures between ages 6 and 36 months, [73]. a combination of febrile and afebrile seizures, and variable Specific tests for the abovementioned genes should always be psychomotor and cognitive impairment. The typical prominent performed: expression in females, although protocadherin 19 gene is on the X chromosome, has been explained through two possible mecha- In newborns or infants with , myoclonic nisms: the existence of compensatory factors in males with seizures, and other drug-resistant seizures, or with status mutated protocadherin 19 (e.g., the protocadherin 19Y gene), and epilepticus not otherwise explicable; the formation of tissue mosaicism with protocadherin 19-positive In children with a progressively worsening clinical course, and protocadherin 19-negative cells and subsequent altered inter- including developmental delay or a loss of previously achieved actions between the two cellular populations [67,68]. milestones; Depienne et al. [69] demonstrated mutations of protocadherin In children with seizures associated with movement disorders, 19 in 12 sodium channel neuronal type 1a subunit gene-negative dysmorphism, alterations of head circumference, or abnormal patients with Dravet syndrome. A hemizygous deletion in a male genitalia; and nine point mutations (four missense and five truncating In children with a clinical history compatible with classic age- mutations) in 11 females were reported, and a cellular mosaicism related epileptic syndromes (e.g., Ohtahara syndrome or West was demonstrated in the fibroblasts of the affected male. Proto- syndrome) or other specific epileptic encephalopathies, such as cadherin 19-positive Dravet syndrome differs from the sodium Dravet syndrome; 30 M. Mastrangelo, V. Leuzzi / Pediatric Neurology 46 (2012) 24e31

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