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Pediatric Neurology 50 (2014) 522e524

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Pediatric Neurology

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Clinical Observations

Genetic Testing in Infantile Spasms Identifies a Chromosome 13qDeletion and Retinoblastoma

Kevin Jones MD FRCP(C) *, Berge A. Minassian MD CM FRCP(C)Division of Neurology, Department of Pediatrics, The Hospital for Sick Children, Toronto, Ontario, Canada

Article HistReceived S2013* Commu

ment of NM5G 1X8,

E-mail a

0887-8994/$http://dx.do

abstract

BACKGROUND: Infantile spasms is an epileptic encephal

opathy and the common final manifestation of numerousdisparate insults to the developing brain during infancy. The varied etiologies may be structural, metabolic, ge-netic, or unknown. Etiological diagnosis is important as it may lead to specific therapy, which may affect devel-opmental outcome. PATIENT: We report a case of infantile spasms of unknown etiology with dysmorphic features,in which genetic copy number variation microarray testing was included in the investigation of the cause of thedisease. RESULTS: A large deletion of chromosome 13 was identified in the region 13q13 to 13q21.3 encompassingthe retinoblastoma gene (13q14.2). Urgent ophthalmological evaluation revealed an asymptomatic retinoblastomaof the left eye, leading to early treatment. CONCLUSION: This is the first case report of infantile spasms specificallyassociated with a chromosome 13q deletion. Chromosomal region 13q13 to 13q21.3 may contain one or moregenes whose hemizygous loss leads to infantile spasms. Copy number variation testing for cryptogenic infantilespasms led to the discovery of a mutation responsible for retinoblastoma, enabling early diagnosis and treatmentof a potentially life-threatening cancer. High-sensitivity molecular diagnosis improves health care and substan-tially reduces expenses. This shift in diagnostic evaluation is broadly relevant to health care.

Keywords: infantile spasms, genetic testing, chromosome 13q deletion, retinoblastoma

Pediatr Neurol 2014; 50: 522-524

Crown Copyright � 2014 Published by Elsevier Inc. All rights reserved.

Introduction

Infantile spasms (IS) represent a seizure type of earlyinfancy, often associated with the electroclinical syndromenamed West syndrome, characterized by epileptic spasms,hypsarrhythmia on electroencephalograph (EEG) anddevelopmental regression. The incidence of IS is estimatedto be between 0.25 and 0.60 per 1000 live births. The pre-valence is 0.15 to 0.2 per 1000 children age 10 years oryounger.1 The etiology of IS may be related to structural,genetic, metabolic, or unknown causes. Patients with struc-tural or metabolic etiologies have a clearly defined underly-ing cause and, often, significant developmental delay before

ory:eptember 22, 2013; Accepted in final form November 23,

nications should be addressed to: Dr. Kevin Jones; Depart-eurology; Sickkids; 555 University Avenue; Toronto, OntarioCanada.ddress: kevin.jones@sickkids.ca

- Crown Copyright � 2014 Published by Elsevier Inc. All rights reserved.i.org/10.1016/j.pediatrneurol.2013.11.018

onset of spasms.2 The number of specific causes of IS hasincreased because of improved diagnostic techniques,including metabolic and genetic testing and neuroimaging.2

Case Report

A 10-month-old male infant presented with IS and global develop-mental delay. He was initially treated with high-dose vigabatrin, butfailed to respond clinically and electrographically within 2 weeks of theinitiation of treatment. He was subsequently treated with phenobarbital(4 mg/kg per day) for episodes of grimacing, head drops, and bruxismsuspected to be seizures, followed a week later by a 6-week taperingcourse of high-dose adrenocorticotropic hormone (ACTH; Synacthen) forIS. Within 2 weeks, he responded to ACTH with resolution of the IS andhypsarrhythmia. During ACTH treatment, he became irritable and hadpoor nocturnal sleep. He had significant constipation since before theonset of his spasms, which worsened with ACTH but did eventuallyimprove with stool softeners. His sleep disturbance worsened with nearcomplete day-night reversal.

Antenatal history was uncomplicated, with protective serology,normal screening, and ultrasounds. His mother was 29 years of age andthis was her first pregnancy. Hewas born at termvia cesarean section. Hewas small for gestational age with a birth weight of 2.420 kg (<third

FIGURE.The patient’s 28.61-Mb deletion in chromosomal region 13q13 to 13q21.3.The coordinates of the undeleted single nucleotide polymorphisms at theboundaries of the deleted region and the relative locations of three genesdiscussed in the text are indicated.

K. Jones, B.A. Minassian / Pediatric Neurology 50 (2014) 522e524 523

percentile), head circumference 33.5 cm (fifteenth percentile), andlength 47 cm (third percentile). He spent 5 days in the neonatal intensivecare unit with transient tachypnea of the newborn and initial difficultieswith feeding. The child had features of developmental delay withoutregression. He developed head control in the first several months of life,was able to reach for objects at 5 months, and was able to roll over at7 months. At 10 months, he was unable to track objects. At 1 year of age,he was able to pass objects to his mouth, babble without saying words,make eye contact, and smile socially. He was unable to sit unsupported.

His father is 31 years old and has protein C and S deficiency, whichlikely contributed to a myocardial infarction. His mother is healthy. Hismaternal grandmother has a bicuspid aortic valve. There was no otherfamily history of developmental delay, epilepsy, genetic disorders, ormetabolic disorders.

At 10months, his weight was 8.9 kg (third percentile), height 70.5 cm(tenth percentile), and head circumference 46 cm (fiftieth percentile). Hehad significant brachycephaly with a prominent forehead and bilateraltemporal narrowing. The eyebrows were prominent with sunken eyesand low-set ears. He had a small nose with a thin upper lip and a higharched palate. His hands had a prominent palmar crease and his feetshowed elongated third toes bilaterally. He had a sacral dimple. Therewas no evidence of a neurocutaneous manifestation.

Pupils were equal and reactive to light and 2mm in diameter. The redreflexes were present bilaterally. He was not clearly fixing and followingobjects. His face was symmetrical and he had a normal gag reflex. He hadnormal tone with symmetrical movements, symmetrical deep tendonreflexes, and bilateral extensor plantar reflexes. He withdrew to stimulisymmetrically. He had evidence of head lag and was only able to sit withsupport. In prone position, he was unable to weight-bear or roll to su-pine. Axial tone was normal. The Moro reflex and asymmetric tonic neckreflex had resolved.

Magnetic resonance imaging performed at 1 year and 3 monthsshowed generalized cerebral volume loss, greater in the anterior regionswith prominent ventricles, of uncertain etiology. EEG before and 2weeksafter vigabatrin therapy showed multiple independent spike foci,modified hypsarrhythmia with absence of sleep features, and electro-decremental events during and without clinical spasms. EEG performed2 weeks after initiation of ACTH showed resolution of hypsarrhythmia,with a low-voltage background and intermittent posterior quadrantdelta waves. EEG after completing 6 weeks of ACTH was within normallimits. Echocardiography revealed an asymptomatic bicuspid aortic valveand pulmonary valve stenosis.

Diagnostic testing for the etiology of IS in a child with dysmorphicfeatures included single nucleotide polymorphism microarray testing.This identified a 28.61-Mb deletion in the long arm of chromosome 13 inbands 13q13 and 13q21.3. It was noted that this deleted region encom-passed the retinoblastoma gene at 13q14.2. Fluorescent in situ hybridi-zation analysis of chromosome region 13q14 was consistent with thedeletion observed by microarray analysis. Fluorescent in situ hybridiza-tion analysis of parental samples indicated that the copy number changewas de novo. Based on the microarray result and the potential risk forretinoblastoma, the child was urgently referred to ophthalmology. Eyeexamination revealed the presence of retinoblastoma in the left eye,which was treated with laser therapy.

Discussion

IS may result from structural, metabolic, genetic, orunknown etiologies, and there is no current consensusregarding the optimal clinical evaluation of childrenwith IS.Etiological diagnosis is important because it may lead tospecific therapy, which may affect developmental outcome.

The diagnostic workup for IS includes neuroimaging andinvestigations for metabolic, genetic, and infectious disor-ders. Known genetic etiologies are increasing at a very rapidrate. Down syndrome and tuberous sclerosis complex havelong been associated with IS. Other IS-associated genesinclude ARX, CDKL5, FOXG1, GRIN1, GRIN2 A, MAGI2, MEF2 C,SLC25A22, SPTAN1, and STXBP1.3-5 The yield and health care

cost of genetic testing is unknown, and testing may need tobe individualized according to the clinical presentation.

In the present case, including genetic testing in theinvestigation of IS in a child with dysmorphic features, ledto the recognition of a clinically significant deletion ofchromosome 13 encompassing one copy of the retinoblas-toma gene at 13q14.2. Urgent ophthalmology evaluationdetected an asymptomatic retinoblastoma of the left eye,resulting in early treatment with laser surgery.

Retinoblastoma is the most common intraocular child-hood cancer with a worldwide incidence of 1 per 15,000 to20,000 live births. It is initiated by mutation of the tumor-suppressor gene RB1.6 Loss of one RB1 allele results in apredisposition to cancer by initiating genomic instabilitywith retinoma formation,whereas the loss of the secondRB1allele within a developing retinal cell leads to the formationof retinoblastoma.6 Molecular factors and the parents’ andclinicians’ response to the child’s presenting signs affect theage at diagnosis. The clinical signs of early retinoblastomamay be subtle and include leukocoria, strabismus, poor vi-sual tracking, glaucoma, and inflammation.6 It is imperativeto diagnose the condition in timely fashion because earlydetection and straightforward surgical treatment can curethe disorder. A delay of more than 6 months from the firstclinical sign to diagnosis is associated with 70% mortality.Mortality rates are highest in regions where the disorder ismost prevalent ranging from 40% to 70% in Asia and Africacompared with 3% to 5% in North America and Europe.6

RB1 mutation testing has a sensitivity of 95%.6 Antenataltesting in childrenwith a parental history of retinoblastomais recommended because there is a 50% risk of inheritingthe abnormal RB1 gene, and 97% of these individuals thendevelop retinoblastoma.6 Antenatal genetic screening forretinoblastoma is not performed without a family history,and de novomutations in RB1may be asymptomatic at birth,as in our patient. Had our patient presented with retino-blastoma before developing IS, sequencing of the RB1 genewould have been negative because the affected allele isdeleted and missing altogether.

A recent review of the genotypeephenotype correlationof previous patients with retinoblastoma and interstitial13q deletions reported that motor and speech delay werecommon in these patients.7 This is especially the case whenthe gene for NUFIP1 (in 13q14.12), an FMRP (fragile Xsyndrome)-interacting protein, is involved. NUFIP1 isdeleted in our patient, as is PCDH8 in 13q21.1 (Figure).PCDH8 is an integral membrane protein that may functionin central nervous systemespecific cellular adhesion.8 Thisis closely related to PCDH19, mutations of which areimportant known causes of epileptic encephalopathy.9

K. Jones, B.A. Minassian / Pediatric Neurology 50 (2014) 522e524524

Constipation, present in our patient, has been noted as ashared feature among patients with deletions spanning RB1.EDNRB, located in 13q22.3 and associated with type 2Hirschsprung disease, has been proposed as a candidate genefor this symptom in these patients.7 However, our patient’sdeletiondoes not encompass this gene, suggesting that EDNRBmay not be the gene underlying constipation in patients withdeletions spanning RB1 or that a second gene associated withthis symptom is present within 13q13 to 13q21.3.

This is the first report of IS associated with a chromo-some 13q deletion. Chromosome 13q13 to 13q21.3 maycontain one or more genes whose hemizygous loss leadsto IS. The use of microarray genetic testing for cryptogenicIS in a patient with nonspecific dysmorphic features ledto the serendipitous discovery of a mutation responsiblefor retinoblastoma, enabling early referral, diagnosis,and treatment of a presymptomatic, potentially life-threatening, condition. A general point broadly relevantto health care is highlighted by our casednamely, thatmolecular diagnosis can have major effects on health care,morbidity, and costs.

We thank the clinicians at The Hospital for Sick Children who cared for of our pa-tient. This work was supported by the endowed University of TorontoMichael BahenChair in Epilepsy Research (to B.A.M.).

Fear comes from uncertainty. When we are absolutely certaiimpervious to fear. Thus a feeling of utter unworthiness can be

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n, whether of our worth or worthlessness, we are almosta source of courage.

Eric Hoffer