- Email: [email protected]
A novel inherited SCN1A mutation associated with different neuropsychological phenotypes: Is there a common core deficit?
Epilepsy & Behavior 43 (2015) 89–92
Contents lists available at ScienceDirect
Epilepsy & Behavior journal homepage: www.elsevier.com/locate/yebeh
Brief Communication
A novel inherited SCN1A mutation associated with different neuropsychological phenotypes: Is there a common core deficit? Claudia Passamonti a,⁎, Cristina Petrelli b, Davide Mei c, Nicoletta Foschi b, Renzo Guerrini c, Leandro Provinciali b, Nelia Zamponi a a b c
Regional Center for Diagnosis and Treatment of Childhood Epilepsy, Department of Neuropsychiatry, Ospedali Riuniti, Ancona, Italy Neurology, Polytechnic University of Marche, Ancona, Italy Pediatric Neurology Unit and Laboratories, Children's Hospital A. Meyer, University of Florence, Italy
a r t i c l e
i n f o
Article history: Received 3 September 2014 Revised 22 October 2014 Accepted 8 November 2014 Available online 7 January 2015 Keywords: SCN1A gene Dravet syndrome spectrum Neuropsychological phenotype
a b s t r a c t We report a three-generation, clinically heterogeneous family in which we identify a novel inherited splicing mutation of the SCN1A gene. Thirteen subjects were submitted to genetic analysis, clinical and instrumental examination, and neuropsychological assessment. In eight subjects, a heterozygous c.2946+5GNA donor splice site alteration in the SCN1A gene was found. Half of them had never had a seizure and showed normal EEG and cognitive profile, whereas the other half had a history of seizures and variable neuropsychological impairments ranging from moderate cognitive disabilities to mild visual–motor impairments. Different clinical phenotypes were identified, including generalized epilepsy with febrile seizure plus (GEFS+), Dravet syndrome, and partial epilepsy with febrile seizure plus (PEFS+). Remarkable clinical heterogeneity can be found among family members carrying the same SCN1A gene mutation. Variable involvement of visual–motor abilities might represent a neuropsychological feature which needs to be further explored in other familial cases. © 2014 Elsevier Inc. All rights reserved.
1. Introduction The SCN1A gene, which encodes the α1 subunit of the neuronal sodium channel, is the most relevant epilepsy gene with the largest number of epilepsy-related mutations. De novo mutations of the SCN1A gene are observed in about 80% of patients with Dravet syndrome (DS). Dravet syndrome is a rare and distinct epileptic encephalopathy, which begins with infantile onset of febrile hemiclonic status epilepticus and evolves into a pattern of multiple seizure types including focal, myoclonic, absence, and atonic seizures. Seizure profile is typically associated with marked slowing or stagnation of psychomotor development, often accompanied by behavioral disturbances [1–5]. Inherited SCN1A mutations, usually missense, are found in 5% of patients with DS. Families of probands with DS with inherited mutations include milder phenotypes consistent with febrile seizures (FS), febrile seizure plus (FS+), or generalized epilepsy with febrile seizure plus (GEFS+) [6,7]. Mutations of the SCN1A gene have also been found in a few patients with partial epilepsy with febrile seizure plus (PEFS +) and, more rarely, in patients with ataxia and coordination defects [6,8].
⁎ Corresponding author. E-mail address: [email protected] (C. Passamonti).
http://dx.doi.org/10.1016/j.yebeh.2014.11.009 1525-5050/© 2014 Elsevier Inc. All rights reserved.
Familial cases offer a unique opportunity to explore the clinical heterogeneity of SCN1A mutations. Previous reports have documented remarkable clinical heterogeneity in both epilepsy-related factors and cognitive level [9,10]. However, variability of neuropsychological functioning in familial cases has been incompletely investigated. More specifically, the neuropsychological profile of those family members who have not experienced seizures, despite carrying the mutation, has not been usually studied. Investigation of both affected and nonaffected family members carrying the same mutation might shed light on the interplay between genetic and epilepsy-related factors in neurocognitive development. Herein we report a three-generation, clinically heterogeneous family in which we identified a splicing mutation of the SCN1A gene and describe neuropsychological findings.
2. Materials and methods 2.1. Clinical assessment The institutional review board approved the study protocol. Written informed consent was obtained from each subject involved in the study. The proband (IIIa) was referred to the pediatric epilepsy clinic for investigation of a seizure disorder that had a familial distribution. The family
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C. Passamonti et al. / Epilepsy & Behavior 43 (2015) 89–92
Fig. 1. The diagram represents a family tree. Ordinal numbers identify generations within the same family (I: first generation, II: second generation, III: third generation). Letters identify subjects belonging to the same family nucleus, i.e., a family group consisting of a pair of adults and their children. Cardinal numbers identify birth position within the family nucleus.
originated from central Italy; a pedigree is shown in Fig. 1. Fifteen subjects belonging to the same family as proband IIIa were contacted for a clinical examination and blood sample collection for DNA analysis. Thirteen subjects gave the consent to participate. The clinical history of each family member was collected by direct interview and retrospective review of the original medical records of individuals IIa2, IIb, IIc1, IIc2, and IIIa. Standard electroencephalography (EEG) recording in sleep and awake states (minimum duration: 40 min) was performed in all subjects. In subjects with a history of seizure, a 24-hour video-EEG recording was also carried out. Electroencephalography records were assessed by an expert neurologist. Based on the clinical history, we decided to investigate the possible involvement of the SCN1A gene. A formal neuropsychological assessment was carried out at the time of the study in each subject using standardized instruments validated on an Italian sample. The mean age of subjects at testing was 37 years (range: 5–73). Assessment included an evaluation of cognitive level (IQ, intelligence quotient) by means of the Wechsler Preschool and Primary Scale of Intelligence (for subject IIIa) and the Wechsler Adult Intelligence Scale-R (for the remaining subjects), attention and executive functions (Trail-Making Test, Bells test), memory abilities (digit and visual–spatial span, short-story test), visual–motor abilities (Rey Figure test), and linguistic skills (Verbal Fluency test). A psychiatric interview was also performed [11–19].
2.2. Genetic analysis Molecular analysis was performed on genomic DNA extracted from the blood using a QIASymphony SP robot (QIAGEN, Hilden, Germany). All 26 exons of SCN1A (accession NM_001165963.1) were amplified by polymerase chain reaction (PCR) and analyzed by direct sequencing in the proband (patient IIIA) (see Supplemental material for details). The novel SCN1A substitution identified in the proband was tested in all the available family individuals to study its segregation. Since the substitution was located near a splice site, we used the Splice Site Prediction by Neural Network (http://www.fruitfly.org/seq_tools/ splice.html), Human Splicing Finder (http://www.umd.be/HSF/), and NetGene2 (http://www.cbs.dtu.dk/services/NetGene2/) splicing prediction tools to explore its role in affecting the splicing process. Unfortunately, the SCN1A gene is not expressed in easily accessible tissues (i.e., blood or fibroblasts), and therefore it has not been possible to study the impact of the substitution on mRNA splicing. This SCN1A novel splicing substitution was not found in a cohort of 190 Caucasian ethnically matched control DNAs (380 alleles)
originating from Italy and was not reported in the ESP6500 (13,006 alleles) and dbSNP137 databases.
3. Results 3.1. Genetic findings We identified a heterozygous c.2946+5GNA donor splice site alteration in the SCN1A gene in eight out of the thirteen subjects examined. Segregation analysis revealed that the mutation showed an incomplete penetrance and variable expressivity. In silico predictions indicated that the substitution likely alters the donor site thus leading to a potential loss of the coding sequence (exon skipping) or to retention of intronic sequence or may lead to a frameshift mutation. Since SCN1A is not expressed in easily accessible tissues, we could not study mRNA, and therefore we cannot establish which one of these mechanisms was the one arising from the mutation.
3.2. Subjects' characteristics and neuropsychological findings Four out of the eight individuals with SCN1A gene mutation (Ia, Ib, Ic, IIa1) have never had seizures, whereas the remaining four had manifested their initial seizures between 8 and 11 months of age. In all these cases, seizures were triggered by fever. The first seizure was complex partial (IIIa, IIb), generalized tonic–clonic (IIa2), and generalized tonic (IIc1, IIc2). Patient IIc2 experienced convulsive hemiclonic status epilepticus at the age of two years. In this subject, status epilepticus was triggered by fever N38° which lasted 4 h and required hospitalization in an intensive care unit. At the time of the study, the youngest subject (IIIa) still experienced partial seizures with secondary generalization with a frequency of twice per year. In the remaining three patients, seizures disappeared between three and twenty-four years of age. All patients with epilepsy had been treated with AEDs (IIa2, IIc1, IIb), with two or more AEDs in individuals IIIa and IIc2; three of these subjects (IIa2, IIa, IIc2) were still treated with more than one AED at the time of the study. Electroencephalography showed alterations in four subjects, all having a history of seizures. Electroencephalography abnormalities included spikes and a spike–wave complex predominantly over the right occipital lobe (IIIa), spike and sharp waves over both temporoparietooccipital lobes (IIa2), sharp waves over both temporoparietal lobes (IIc2), and sharp waves over the left temporoparietooccipital lobe (IIc1). No structural alteration was revealed by MRI in any subject.
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described two clinically heterogeneous families with a deletion encompassing the SCN1A gene [9,10]. A similar variability can be observed in the present family, in which the nucleotide alteration is thought to alter the mRNA splicing. Such variability could be explained by the different genetic backgrounds of the affected individuals [20]. In particular, other genes, such as other sodium or potassium channels, may modulate the observed phenotype as demonstrated for the SCN9A gene [6]. Variable cognitive impairments have been described in children and adults with SCN1A gene mutations [5,21,22]. In some cases, a particular involvement of the abilities concerning visual functions (visual–motor and visual–spatial deficits) has been reported [2–4,21]. Both the frequency of convulsive seizures and the early appearance of myoclonus and absences have been associated with the degree of mental impairment [23]. In the family we are presenting, the worst cognitive outcome was observed in subject IIc1, who was diagnosed with Dravet syndrome. This subject experienced prolonged status epilepticus and frequent generalized tonic seizures during childhood. By contrast, his brother (IIc2) had few febrile seizures during the first three years of life; his cognitive development was good, and only mild visual defects were detected. The youngest subject (IIIa), who has experienced only rare partial seizures with secondary generalization, showed a mild delay in visual–motor functions, while general cognitive level and linguistic abilities were preserved. The observed neuropsychological pattern in these subjects might be related to the prominent EEG activity in occipitoparietal areas. The involvement of visual and visual–motor abilities can be considered a common neuropsychological feature among most of the present family members, despite wide differences in cognitive functioning. This evidence is in line with recent results showing a specific impairment in visual abilities in children with the mild variant of Dravet syndrome [4]. It is worth noting that the genetic mutation (SCN1A) with a loss of function in Nav1.1 is expected from experimental studies to determine a decreased excitability of cerebellar Purkinje neurons [24]; this dysfunction may play a role in determining a general developmental delay and impairing visual–motor control [25,26].
As for the neurocognitive profile, a wide variability was observed. Three individuals out of the eight (Ia, Ib, IIa1) showed a normal IQ and no specific cognitive impairment or psychiatric signs. All these subjects had never had seizures. Subject Ic had a normal IQ and a mild impairment in visual–motor functions. Excepting subject Ic, all the remaining subjects had experienced at least one seizure in the first year of life. Subject IIc1 had a normal IQ, mild strabismus, and no stereopsis; subject IIc2 had moderate mental retardation associated with a severe visual–motor defect, ataxia, and psychotic symptoms, while linguistic functions were better preserved; subject IIa2 had a normal IQ, mild visual–motor impairment, and a history of bipolar disorder; subject IIIa had a normal IQ and a mild impairment in sustained attention and visual– motor abilities. A summary of clinical findings for family members affected by epilepsy is presented in Table 1.
4. Discussion The family we are reporting shows a remarkable clinical heterogeneity, comprising various phenotypes, all of which had been associated previously with SCN1A abnormalities. Heterogeneity involved both seizure severity and neuropsychological functioning. Eight subjects were found to carry the mutation. Half of them (Ia, Ib, Ic, IIa1) had never had seizures and presented a normal cognitive profile, with only one subject exhibiting mild difficulties in visual–motor abilities. Three of these subjects belonged to the first generation and one subject to the second generation. The remaining four subjects presented a history of seizures and variable neuropsychological impairments associated with psychiatric symptoms in some cases. On the basis of clinical examination, different phenotypes have been identified for these four subjects: one subject was affected by GEFS + (IIa2), one had showed rare febrile seizures (IIc2), another one was affected by Dravet syndrome (IIc1), and the last one (IIIa) has a diagnosis compatible with PEFS +: he had presented complex partial seizures in the first year of life and showed EEG alterations with a right occipital focus. Our observations confirm those by Guerrini et al. and Suls et al., who
Table 1 Clinical features of patients with SCN1A mutation. Patient ID/gender
Age
Epilepsy diagnosis
First seizure type/onset: offset
No. of seizures before the age of 1
Seizure type during follow-up
No. of SE/age
EEG abnormalities
MRI
AEDs
Cognitive level (IQ)
Neuropsychological impairments
Psychiatric illness
IIIa/M
5y
PEFS+
CPS/11 m: ongoing
1
GTCS
0
Neg
VPA; TPM; LEV
Average (95)
Sustained attention; visual–motor abilities
No
IIa2/F
42 y
GEFS+
1
GTCS
0
Neg
CNZ
Average (90)
Visual–motor abilities
Bipolar disorder
IIc2/M
29 y
1
GTS
0
Febrile and afebrile GTS; weekly
1 (4 h)/2 y
Neg
Mild strabismus, lack of stereopsis Moderate apraxia and ataxia
Dysphoria
Weekly
PB until 9y CNZ, PB
Average (92)
38 y
T-P sharp waves, bilateral T-P-O sharp waves, left
Neg
IIc1/M
Febrile seizures DS
Febrile and afebrile GTCS 8 m:2 y Febrile GTS 7 m:3 y Febrile GTS 8 m:18 y
O spikes and spike– wave complex, predominantly on the right T-P-O spike and sharp waves, bilateral
Patient ID/gender
Age
Epilepsy diagnosis
First seizure type/onset: offset
No. of seizure b12 m
Seizure type during follow-up
No. of SE/age
EEG abnormalities
MRI
AEDs
Cognitive level (IQ)
Neuropsychological impairments
Psychiatric illness
Ic/M
73 y
///
///
///
///
///
Neg
Neg
///
Average (90)
No
Ia/M Ic/M IIa1/F
67 y 65 y 47 y
/// /// ///
/// /// ///
/// /// ///
/// /// ///
/// /// ///
Neg Neg Neg
Neg Neg Neg
/// /// ///
Average (95) Average (93) Average (102)
Visual–motor abilities No No No
MMR (45)
Psychosis
No No No
Legend: CNZ, clonazepam; CPS, complex partial seizures; DS, Dravet syndrome; EEG, electroencephalography; F, female; GTS, generalized tonic seizures; GTCS, generalized tonic–clonic seizures; LEV, levetiracetam; M, male; m, months; MMR, moderate mental retardation; MRI, magnetic resonance imaging; NA, not applicable; O, occipital; P, parietal; PB, phenobarbital; T, temporal; TPM, topiramate; y, years; VPA, valproate; AEDs, antiepileptic drugs; Neg, negative.
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A careful examination of neuropsychological functioning in other familial cases should be encouraged in future investigations in order to better elucidate the specific role of SCN1A gene mutations on cognitive development. Disclosure None of the authors has any conflict of interest to disclose. Appendix A. Supplementary data Supplementary data to this article can be found online at http://dx. doi.org/10.1016/j.yebeh.2014.11.009. References [1] Commission on Classification and Terminology of the International League Against Epilepsy. Proposal for revised classification of epilepsies and epileptic syndromes. Epilepsia 1989;30:389–99. [2] Wolff M, Cass-Perrot C, Dravet C. Severe myoclonic epilepsy of infants (Dravet syndrome): natural history and neuropsychological findings. Epilepsia 2006;47: 45–8. [3] Riva D, Vago C, Pantaleoni C, Bulgheroni S, Mantegazza M, Franceschetti S. Progressive neurocognitive decline in two children with Dravet syndrome, de novo SCN1A truncations and different epileptic phenotypes. Am J Med Genet 2009;49:2339–45. [4] Chieffo D, Ricci D, Baranello G, Martinelli D, Veredice C, Lettori D, et al. Early development in Dravet syndrome; visual function impairment precedes cognitive decline. Epilepsy Res 2010;93:73–9. [5] Petrelli C, Passamonti C, Cesaroni E, Mei D, Guerrini R, Zamponi N, et al. Early clinical features in Dravet syndrome patients with and without SCN1A mutations. Epilepsy Res 2012;99:21–7. [6] Singh NA, Pappas C, Dahle EJ, Claes LR, Pruess TH, De Jonghe P, et al. A role of SCN9A in human epilepsies, as a cause of febrile seizures and as a potential modifier of Dravet syndrome. PLoS Genet 2009;5(9):e1000649. [7] Nabbout R, Gennaro E, Dalla Bernardina B, Dulac O, Madia F, Bertini E, et al. Spectrum of SCN1A mutations in severe myoclonic epilepsy of infancy. Neurology 2003;60:1961–7. [8] Escayg A, Goldin AL. Sodium channel SCN1A and epilepsy: mutations and mechanisms. Epilepsia 2010;51:1650–8.
[9] Guerrini R, Cellini E, Mei D, Metitieri T, Petrelli C, Pucatti D, et al. Variable epilepsy phenotypes associated with a familial intragenic deletion of the SCN1A gene. Epilepsia 2010;51:2474–7. [10] Suls A, Velizarova R, Yordanova I, Deprez L, Van Dyck T, Wauters J, et al. Four generations of epilepsy caused by an inherited microdeletion of the SCN1A gene. Neurology 2010;6:72–6. [11] Wechsler D. Scala Wechsler a livello prescolare e di scuola elementare. Firenze: Edizioni OS; 1973. [12] Wechsler D. WAIS-R. Scala d'Intelligenza Wechsler per Adulti-Riveduta. Firenze: Edizioni OS; 1981. [13] Giovagnoli AR, Del Pesce M, Mascheroni S, Simoncelli M, Laiacona M, Capitani E. Trail making test: normative values from 287 normal adult controls. Ital J Neurol Sci 1996;17:305–9. [14] Biancardi A, Stoppa E. Il test delle Campanelle modificato: una proposta per lo studio dell'attenzione in età evolutiva. Psichiatr Infanz Adolesc 1997;64:73–84. [15] Tressoldi PE, Vio M, Gugliotta M, Bisiacchi PS, Cendron M. Batteria di valutazione neuropsicologica per l'età evolutiva (BVN 5-11). Trento: Erickson; 2005. [16] Scarpa P, Piazzini A, Pesenti G, Brovedani P, Toraldo A, Turner K, et al. Italian neuropsychological instruments to assess memory, attention and frontal functions for developmental age. Neurol Sci 2006;27(6):381–96. [17] Spinnler H, Tognoni G. Standardizzazione e taratura italiana di test neuropsicologici. Ital J Neurol Sci 1987;8:35–50. [18] Rey A. Reattivo della Figura Complessa. Firenze: Organizzazioni Speciali; 1959. [19] Novelli G, Papagno C, Capitani E, Laiacona N, Vallar G, Cappa SF. Tre test clinici di ricerca e produzione lessicale. Taratura su soggetti normali. Arch Psicol Neurol Psichiatr 1986;47:477–506. [20] Kearney JA. Genetic modifiers of neurological disease. Curr Opin Genet Dev 2011;21: 349–353.3. [21] Ragona F, Brazzo D, De Giorgi I, Morbi M, Freri E, Teutonico F, et al. Dravet syndrome: early clinical manifestations and cognitive outcome in 37 Italian patients. Brain Dev 2010;32:71–7. [22] Zamponi N, Passamonti C, Petrelli C, Veggiotti P, Baldassari C, Verrotti A, et al. Vaccination and occurrence of seizures in SCN1A mutation-positive patients: a multicenter Italian study. Pediatr Neurol 2014;50:228–32. [23] Cass-Perrot C, Wolff M, Dravet C. Neuropsychological aspects of severe myoclonic epilepsy in infancy. In: Jambaque I, Lassonde M, Dulac O, editors. Neuropsychology of childhood epilepsy. New York: Kluwer Academic/Plenum Publishers; 2011. p. 131–40. [24] Oakley JC, Kalume F, Catterall WA. Insights into pathophysiology and therapy from a mouse model of Dravet syndrome. Epilepsia 2011;52:59–61. [25] Schmahmann JD. Disorders of the cerebellum: ataxia, dysmetria of thought, and the cerebellar cognitive affective syndrome. J Neuropsychiatry Clin Neurosci 2004;16: 367–78. [26] Cerminara NL, Apps R, Marple-Horvat DE. An internal model of a moving visual target in the lateral cerebellum. J Physiol 2009;587:429–42.
Contents lists available at ScienceDirect
Epilepsy & Behavior journal homepage: www.elsevier.com/locate/yebeh
Brief Communication
A novel inherited SCN1A mutation associated with different neuropsychological phenotypes: Is there a common core deficit? Claudia Passamonti a,⁎, Cristina Petrelli b, Davide Mei c, Nicoletta Foschi b, Renzo Guerrini c, Leandro Provinciali b, Nelia Zamponi a a b c
Regional Center for Diagnosis and Treatment of Childhood Epilepsy, Department of Neuropsychiatry, Ospedali Riuniti, Ancona, Italy Neurology, Polytechnic University of Marche, Ancona, Italy Pediatric Neurology Unit and Laboratories, Children's Hospital A. Meyer, University of Florence, Italy
a r t i c l e
i n f o
Article history: Received 3 September 2014 Revised 22 October 2014 Accepted 8 November 2014 Available online 7 January 2015 Keywords: SCN1A gene Dravet syndrome spectrum Neuropsychological phenotype
a b s t r a c t We report a three-generation, clinically heterogeneous family in which we identify a novel inherited splicing mutation of the SCN1A gene. Thirteen subjects were submitted to genetic analysis, clinical and instrumental examination, and neuropsychological assessment. In eight subjects, a heterozygous c.2946+5GNA donor splice site alteration in the SCN1A gene was found. Half of them had never had a seizure and showed normal EEG and cognitive profile, whereas the other half had a history of seizures and variable neuropsychological impairments ranging from moderate cognitive disabilities to mild visual–motor impairments. Different clinical phenotypes were identified, including generalized epilepsy with febrile seizure plus (GEFS+), Dravet syndrome, and partial epilepsy with febrile seizure plus (PEFS+). Remarkable clinical heterogeneity can be found among family members carrying the same SCN1A gene mutation. Variable involvement of visual–motor abilities might represent a neuropsychological feature which needs to be further explored in other familial cases. © 2014 Elsevier Inc. All rights reserved.
1. Introduction The SCN1A gene, which encodes the α1 subunit of the neuronal sodium channel, is the most relevant epilepsy gene with the largest number of epilepsy-related mutations. De novo mutations of the SCN1A gene are observed in about 80% of patients with Dravet syndrome (DS). Dravet syndrome is a rare and distinct epileptic encephalopathy, which begins with infantile onset of febrile hemiclonic status epilepticus and evolves into a pattern of multiple seizure types including focal, myoclonic, absence, and atonic seizures. Seizure profile is typically associated with marked slowing or stagnation of psychomotor development, often accompanied by behavioral disturbances [1–5]. Inherited SCN1A mutations, usually missense, are found in 5% of patients with DS. Families of probands with DS with inherited mutations include milder phenotypes consistent with febrile seizures (FS), febrile seizure plus (FS+), or generalized epilepsy with febrile seizure plus (GEFS+) [6,7]. Mutations of the SCN1A gene have also been found in a few patients with partial epilepsy with febrile seizure plus (PEFS +) and, more rarely, in patients with ataxia and coordination defects [6,8].
⁎ Corresponding author. E-mail address: [email protected] (C. Passamonti).
http://dx.doi.org/10.1016/j.yebeh.2014.11.009 1525-5050/© 2014 Elsevier Inc. All rights reserved.
Familial cases offer a unique opportunity to explore the clinical heterogeneity of SCN1A mutations. Previous reports have documented remarkable clinical heterogeneity in both epilepsy-related factors and cognitive level [9,10]. However, variability of neuropsychological functioning in familial cases has been incompletely investigated. More specifically, the neuropsychological profile of those family members who have not experienced seizures, despite carrying the mutation, has not been usually studied. Investigation of both affected and nonaffected family members carrying the same mutation might shed light on the interplay between genetic and epilepsy-related factors in neurocognitive development. Herein we report a three-generation, clinically heterogeneous family in which we identified a splicing mutation of the SCN1A gene and describe neuropsychological findings.
2. Materials and methods 2.1. Clinical assessment The institutional review board approved the study protocol. Written informed consent was obtained from each subject involved in the study. The proband (IIIa) was referred to the pediatric epilepsy clinic for investigation of a seizure disorder that had a familial distribution. The family
90
C. Passamonti et al. / Epilepsy & Behavior 43 (2015) 89–92
Fig. 1. The diagram represents a family tree. Ordinal numbers identify generations within the same family (I: first generation, II: second generation, III: third generation). Letters identify subjects belonging to the same family nucleus, i.e., a family group consisting of a pair of adults and their children. Cardinal numbers identify birth position within the family nucleus.
originated from central Italy; a pedigree is shown in Fig. 1. Fifteen subjects belonging to the same family as proband IIIa were contacted for a clinical examination and blood sample collection for DNA analysis. Thirteen subjects gave the consent to participate. The clinical history of each family member was collected by direct interview and retrospective review of the original medical records of individuals IIa2, IIb, IIc1, IIc2, and IIIa. Standard electroencephalography (EEG) recording in sleep and awake states (minimum duration: 40 min) was performed in all subjects. In subjects with a history of seizure, a 24-hour video-EEG recording was also carried out. Electroencephalography records were assessed by an expert neurologist. Based on the clinical history, we decided to investigate the possible involvement of the SCN1A gene. A formal neuropsychological assessment was carried out at the time of the study in each subject using standardized instruments validated on an Italian sample. The mean age of subjects at testing was 37 years (range: 5–73). Assessment included an evaluation of cognitive level (IQ, intelligence quotient) by means of the Wechsler Preschool and Primary Scale of Intelligence (for subject IIIa) and the Wechsler Adult Intelligence Scale-R (for the remaining subjects), attention and executive functions (Trail-Making Test, Bells test), memory abilities (digit and visual–spatial span, short-story test), visual–motor abilities (Rey Figure test), and linguistic skills (Verbal Fluency test). A psychiatric interview was also performed [11–19].
2.2. Genetic analysis Molecular analysis was performed on genomic DNA extracted from the blood using a QIASymphony SP robot (QIAGEN, Hilden, Germany). All 26 exons of SCN1A (accession NM_001165963.1) were amplified by polymerase chain reaction (PCR) and analyzed by direct sequencing in the proband (patient IIIA) (see Supplemental material for details). The novel SCN1A substitution identified in the proband was tested in all the available family individuals to study its segregation. Since the substitution was located near a splice site, we used the Splice Site Prediction by Neural Network (http://www.fruitfly.org/seq_tools/ splice.html), Human Splicing Finder (http://www.umd.be/HSF/), and NetGene2 (http://www.cbs.dtu.dk/services/NetGene2/) splicing prediction tools to explore its role in affecting the splicing process. Unfortunately, the SCN1A gene is not expressed in easily accessible tissues (i.e., blood or fibroblasts), and therefore it has not been possible to study the impact of the substitution on mRNA splicing. This SCN1A novel splicing substitution was not found in a cohort of 190 Caucasian ethnically matched control DNAs (380 alleles)
originating from Italy and was not reported in the ESP6500 (13,006 alleles) and dbSNP137 databases.
3. Results 3.1. Genetic findings We identified a heterozygous c.2946+5GNA donor splice site alteration in the SCN1A gene in eight out of the thirteen subjects examined. Segregation analysis revealed that the mutation showed an incomplete penetrance and variable expressivity. In silico predictions indicated that the substitution likely alters the donor site thus leading to a potential loss of the coding sequence (exon skipping) or to retention of intronic sequence or may lead to a frameshift mutation. Since SCN1A is not expressed in easily accessible tissues, we could not study mRNA, and therefore we cannot establish which one of these mechanisms was the one arising from the mutation.
3.2. Subjects' characteristics and neuropsychological findings Four out of the eight individuals with SCN1A gene mutation (Ia, Ib, Ic, IIa1) have never had seizures, whereas the remaining four had manifested their initial seizures between 8 and 11 months of age. In all these cases, seizures were triggered by fever. The first seizure was complex partial (IIIa, IIb), generalized tonic–clonic (IIa2), and generalized tonic (IIc1, IIc2). Patient IIc2 experienced convulsive hemiclonic status epilepticus at the age of two years. In this subject, status epilepticus was triggered by fever N38° which lasted 4 h and required hospitalization in an intensive care unit. At the time of the study, the youngest subject (IIIa) still experienced partial seizures with secondary generalization with a frequency of twice per year. In the remaining three patients, seizures disappeared between three and twenty-four years of age. All patients with epilepsy had been treated with AEDs (IIa2, IIc1, IIb), with two or more AEDs in individuals IIIa and IIc2; three of these subjects (IIa2, IIa, IIc2) were still treated with more than one AED at the time of the study. Electroencephalography showed alterations in four subjects, all having a history of seizures. Electroencephalography abnormalities included spikes and a spike–wave complex predominantly over the right occipital lobe (IIIa), spike and sharp waves over both temporoparietooccipital lobes (IIa2), sharp waves over both temporoparietal lobes (IIc2), and sharp waves over the left temporoparietooccipital lobe (IIc1). No structural alteration was revealed by MRI in any subject.
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described two clinically heterogeneous families with a deletion encompassing the SCN1A gene [9,10]. A similar variability can be observed in the present family, in which the nucleotide alteration is thought to alter the mRNA splicing. Such variability could be explained by the different genetic backgrounds of the affected individuals [20]. In particular, other genes, such as other sodium or potassium channels, may modulate the observed phenotype as demonstrated for the SCN9A gene [6]. Variable cognitive impairments have been described in children and adults with SCN1A gene mutations [5,21,22]. In some cases, a particular involvement of the abilities concerning visual functions (visual–motor and visual–spatial deficits) has been reported [2–4,21]. Both the frequency of convulsive seizures and the early appearance of myoclonus and absences have been associated with the degree of mental impairment [23]. In the family we are presenting, the worst cognitive outcome was observed in subject IIc1, who was diagnosed with Dravet syndrome. This subject experienced prolonged status epilepticus and frequent generalized tonic seizures during childhood. By contrast, his brother (IIc2) had few febrile seizures during the first three years of life; his cognitive development was good, and only mild visual defects were detected. The youngest subject (IIIa), who has experienced only rare partial seizures with secondary generalization, showed a mild delay in visual–motor functions, while general cognitive level and linguistic abilities were preserved. The observed neuropsychological pattern in these subjects might be related to the prominent EEG activity in occipitoparietal areas. The involvement of visual and visual–motor abilities can be considered a common neuropsychological feature among most of the present family members, despite wide differences in cognitive functioning. This evidence is in line with recent results showing a specific impairment in visual abilities in children with the mild variant of Dravet syndrome [4]. It is worth noting that the genetic mutation (SCN1A) with a loss of function in Nav1.1 is expected from experimental studies to determine a decreased excitability of cerebellar Purkinje neurons [24]; this dysfunction may play a role in determining a general developmental delay and impairing visual–motor control [25,26].
As for the neurocognitive profile, a wide variability was observed. Three individuals out of the eight (Ia, Ib, IIa1) showed a normal IQ and no specific cognitive impairment or psychiatric signs. All these subjects had never had seizures. Subject Ic had a normal IQ and a mild impairment in visual–motor functions. Excepting subject Ic, all the remaining subjects had experienced at least one seizure in the first year of life. Subject IIc1 had a normal IQ, mild strabismus, and no stereopsis; subject IIc2 had moderate mental retardation associated with a severe visual–motor defect, ataxia, and psychotic symptoms, while linguistic functions were better preserved; subject IIa2 had a normal IQ, mild visual–motor impairment, and a history of bipolar disorder; subject IIIa had a normal IQ and a mild impairment in sustained attention and visual– motor abilities. A summary of clinical findings for family members affected by epilepsy is presented in Table 1.
4. Discussion The family we are reporting shows a remarkable clinical heterogeneity, comprising various phenotypes, all of which had been associated previously with SCN1A abnormalities. Heterogeneity involved both seizure severity and neuropsychological functioning. Eight subjects were found to carry the mutation. Half of them (Ia, Ib, Ic, IIa1) had never had seizures and presented a normal cognitive profile, with only one subject exhibiting mild difficulties in visual–motor abilities. Three of these subjects belonged to the first generation and one subject to the second generation. The remaining four subjects presented a history of seizures and variable neuropsychological impairments associated with psychiatric symptoms in some cases. On the basis of clinical examination, different phenotypes have been identified for these four subjects: one subject was affected by GEFS + (IIa2), one had showed rare febrile seizures (IIc2), another one was affected by Dravet syndrome (IIc1), and the last one (IIIa) has a diagnosis compatible with PEFS +: he had presented complex partial seizures in the first year of life and showed EEG alterations with a right occipital focus. Our observations confirm those by Guerrini et al. and Suls et al., who
Table 1 Clinical features of patients with SCN1A mutation. Patient ID/gender
Age
Epilepsy diagnosis
First seizure type/onset: offset
No. of seizures before the age of 1
Seizure type during follow-up
No. of SE/age
EEG abnormalities
MRI
AEDs
Cognitive level (IQ)
Neuropsychological impairments
Psychiatric illness
IIIa/M
5y
PEFS+
CPS/11 m: ongoing
1
GTCS
0
Neg
VPA; TPM; LEV
Average (95)
Sustained attention; visual–motor abilities
No
IIa2/F
42 y
GEFS+
1
GTCS
0
Neg
CNZ
Average (90)
Visual–motor abilities
Bipolar disorder
IIc2/M
29 y
1
GTS
0
Febrile and afebrile GTS; weekly
1 (4 h)/2 y
Neg
Mild strabismus, lack of stereopsis Moderate apraxia and ataxia
Dysphoria
Weekly
PB until 9y CNZ, PB
Average (92)
38 y
T-P sharp waves, bilateral T-P-O sharp waves, left
Neg
IIc1/M
Febrile seizures DS
Febrile and afebrile GTCS 8 m:2 y Febrile GTS 7 m:3 y Febrile GTS 8 m:18 y
O spikes and spike– wave complex, predominantly on the right T-P-O spike and sharp waves, bilateral
Patient ID/gender
Age
Epilepsy diagnosis
First seizure type/onset: offset
No. of seizure b12 m
Seizure type during follow-up
No. of SE/age
EEG abnormalities
MRI
AEDs
Cognitive level (IQ)
Neuropsychological impairments
Psychiatric illness
Ic/M
73 y
///
///
///
///
///
Neg
Neg
///
Average (90)
No
Ia/M Ic/M IIa1/F
67 y 65 y 47 y
/// /// ///
/// /// ///
/// /// ///
/// /// ///
/// /// ///
Neg Neg Neg
Neg Neg Neg
/// /// ///
Average (95) Average (93) Average (102)
Visual–motor abilities No No No
MMR (45)
Psychosis
No No No
Legend: CNZ, clonazepam; CPS, complex partial seizures; DS, Dravet syndrome; EEG, electroencephalography; F, female; GTS, generalized tonic seizures; GTCS, generalized tonic–clonic seizures; LEV, levetiracetam; M, male; m, months; MMR, moderate mental retardation; MRI, magnetic resonance imaging; NA, not applicable; O, occipital; P, parietal; PB, phenobarbital; T, temporal; TPM, topiramate; y, years; VPA, valproate; AEDs, antiepileptic drugs; Neg, negative.
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