So-called cryptogenic partial seizures resulting from a subtle cortical dysgenesis due to a doublecortin gene mutation

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1 Seizure 2002; 11: doi: /seiz , available online at on So-called cryptogenic partial seizures resulting from a subtle cortical dysgenesis due to a doublecortin gene mutation V. DES PORTES, L. ABAOUB, A. JOANNARD, I. SOUVILLE, F. FRANCIS, J. M. PINARD, J. CHELLY, C. BELDJORD & P. S. JOUK Service de neuropédiatrie, Hôpital Saint Vincent-de-Paul, Paris, France; INSERM U129-ICGM, Faculté Cochin, Paris, France; Service de neuropédiatrie, Hôpital R Poincaré, Garches, France; Service de génétique; Service de neurologie; Service de pediatrie, CHU de Grenoble, France Correspondence to: Dr Vincent des Portes, Service de neuropédiatrie, Hôpital Saint Vincent-de-Paul, 82 Avenue Denfert Rochereau, 75014, Paris, France. desportes@cochin.inserm.fr We report the case of a female suffering from resistant partial seizures, which were related to cryptogenic epilepsy, as the cerebral cortex was considered normal on the initial MRI images. As her son is mentally retarded and has a pachygyria, the doublecortin gene, usually involved in band heterotopia or lissencephaly, was screened for mutations. A missense mutation was identified, shared by both the son and his mother, and a subtle discontinuous subcortical heterotopia was subsequently detected on the mother s MRI. The pathophysiology of epilepsy in this woman is discussed in the light of the role of doublecortin, not only in neuronal migration, but also in axonal growth and dendritic connectivity. Key words: epilepsy; doublecortin; cortical dysgenesis. c 2002 BEA Trading Ltd. Published by Elsevier Science Ltd. All rights reserved INTRODUCTION Within the last few years, advances in genetics have reached the field of epilepsy and have led to new insights in previous clinical classifications of epileptic disorders 1. In particular, molecular mechanisms underlying idiopathic epilepsy (defined by age-related onset and specific clinical and EEG patterns) are progressively being unraveled 2. At the same time, progress in brain imaging, especially MRI, has enabled direct visualisation of many brain injuries and cortical malformations responsible for symptomatic epilepsy. Moreover, the underlying molecular mechanisms of some cortical malformations, such as lissencephaly type I 3, subcortical laminar heterotopia (SCLH) 4, or periventricular heterotopia 5, have been identified. Nevertheless, many recurrent seizures, socalled cryptogenic epilepsies, do not meet idiopathic electroclinical criteria, and remain of unknown aetiology as far as no obvious lesion can be detected on routine MRI. Here, we report an adult female suffering from recurrent partial seizures considered as cryptogenic because no obvious cortical abnormality was initially detected on routine MRI, evaluated by three independant radiologists. Only after diagnosis of a diffuse pachygyria in her son with epilepsy and mental retardation, a mutation in the doublecortin gene was identified and a thin discontinuous band of gray matter heterotopia was detected on coronal MRI. CASE REPORT A 21-year-old female (patient 1) was admitted to hospital in March 1987, following a first seizure. After a long tiring journey, the patient experienced left hand paresthesia, spreading to the arm, followed by a generalized tonic clonic seizure. Physical and neurological examinations were normal, including blood pressure. EEG showed alternating left and right high amplitude and slow discharges. A cranial /02/$22.00/0 c 2002 BEA Trading Ltd. Published by Elsevier Science Ltd. All rights reserved

2 274 V. des Portes et al. Fig. 1: Interictal EEG recording in patient 1 (closed eyes, awake). Normal background activity surrounded with delta waves and rare spikes in temporal regions. CT scan and routine brain MRI including sagittal T1-weighted images, axial T2-weighted and coronal T1-weighted images, were considered normal by three independant neuroradiologists. Carbamazepine monotherapy was introduced. Then, two types of seizures were observed: rare tonic clonic seizures and frequent partial seizures, described as left hand paresthesia or speaking and writing impairments during a few seconds, which occur in clusters of two to ten seizures a day, two to three times a month. These partial seizures have not responded, to different mono or polytherapies including carbamazepine, valproic acid, phenylhydantoin and vigabatrin. Many EEG recordings show a normal background activity surrounded with delta waves and rare spikes, mainly in temporal and parietal regions (Fig. 1). Psychometric tests were performed at the age of 34 years; no intellectual impairment was observed: verbal IQ was 113 (Binois Pichot test) and non verbal IQ was 111 (raven Progressive matrix 38). She passed her high school examinations, worked as an accountant s assistant, but lost her job several years ago and could not find a new position due to concern about her seizures. The son of patient 1 (patient 2) was born at full term without any foetal distress. During pregnancy, his mother received carbamazepine and had no tonic clonic seizures; a stable ventricle dilatation was detected during the last three months of the pregnancy. He walked at 24 months of age with unstable gait. At 3 years of age, speech delay was obvious, as he was unable to pronounce any significant words; comprehension seemed more preserved. Precise IQ evaluation was prevented by the child s opposition, but several McCarthy subtests predicted moderate mental retardation. Clinical assessment, including hearing, neurological, genital and skin examinations, were normal. On EEG records, background activity is replaced by diffuse high amplitude alpha like runs surrounded by bilateral discharges of spikes and slow waves, a pattern suggesting diffuse cortical dysplasia. Indeed, brain MRI exhibited diffuse pachygyria, more obvious over the fronto-parietal regions (Fig. 2(a)). Surprisingly, no seizure was observed until a first short episode of febrile convulsion at the age of 4 and half years. The patient remained seizure free 6 months after treatment with valproic acid. Molecular analyses of the doublecortin gene in this patient and his mother were performed as described previously 4. A point mutation was found, leading to a predicted R(196)S non-conservative substitution (Fig. 3). Taking into account the mutation in doublecortin, previous MRI sequences were checked again and new coronal T1-weighted images (inversion/recovery) were performed in patient 1, specifically to look for a subtle cortical dysgenesis. A very thin, bilateral discontinuous grey matter band heterotopia could indeed be seen, mainly beneath the right parietal cortex (Fig. 2(b) and (c)).

3 Subtle cortical dysgenesis and doublecortin mutation 275 Fig. 2: (a) Axial T1-weighted image in patient 2 (2 years old) shows a symmetric anterior pachygyria. (b) Brain MRI of patient 1, performed by a 1 tesla gyroscan machine. The pulse sequences included sagittal T1-weighted and axial T2-weighted images (not shown), and coronal T1-weighted images (inversion/recovery). Slice thickness was 4 mm with 1 mm intervals. (c) After molecular diagnosis, focus on new T1-weighted coronal sequences (inversion/recovery) did show a subtle bilateral grey matter band heterotopia beneath the parietal cortex. DISCUSSION In patient 1 suffering from resistant partial seizures, diagnosis of so-called cryptogenic epilepsy was initially considered because: (1) cognitive level and social integration were normal; (2) EEG records did not show characteristic patterns of presently identified idiopathic epilepsy syndromes and did not show any persistant focal abnormality suggestive of brain lesion; (3) routine brain MRI were considered normal. Indeed, subtle subcortical heterotopia were not seen until her son was found to have pachygyria and a mutation in the doublecortin gene subsequently detected.

4 276 V. des Portes et al. Fig. 3: Sequence gel electrophoregram showing a C A point mutation in heterozygous patient 1. Genomic DNA from subject was screened using DGGE (data not shown). A PCR fragment with an aberrant migration pattern was identified in exon 3 and sequenced using direct automated sequencing. Proportion of normal and mutated alleles are in the same range. The region of exon 3 sequence containing the mutation is shown on forward and reverse strands: the genomic DNA PCR product contains both the normal and mutated alleles of the affected patient. The C A mutation in nucleotide 1001, changes codon 196 from an arginine, a basic amino acid to serine, a neutral residue; this is predicted to change the Doublecortin protein significantly. Moreover, this mutation was neither found in healthy parents of patient 1 nor in a control set of 90 unrelated X chromosomes, which makes it unlikely to be a polymorphism. We and others have previously identified the doublecortin gene 6, 7 expressed in migrating neurons 8, 9, and responsible for lissencephaly in males and SCLH in females. The clinical severity of SCLH varies strikingly from asymptomatic clinical presentation with heterotopic bands assessed by MRI, to severe mental impairment with intractable epilepsy. In most cases, the relative thickness of the heterotopic band seems to correlate with the phenotype, as patients with thicker bands have a more severe mental retardation and seizures 10. As far as we know, no mutation in doublecortin has been previously described in a symptomatic female with resistant partial seizures, associated with a very subtle neuronal heterotopia on MRI. The discrepancy between the severity of the epilepsy and the small area of heterotopic neurons on MRI is striking and might be explained by two ways: first, it is likely that a very small number of misplaced neurons would be enough to result in a severe epilepsy. An alternative explanation should be considered in the light of recent data describing doublecortin expression in developing brain: Francis et al. 8 studied temporal and spatial expression of doublecortin in mouse and rat developing cerebral cortex. A clear expression of doublecortin was shown in the cells of the cortical plate, neurons that have apparently reached their final destination. Moreover, doublecortin expression in cultured maturating neurons is concentrated at the tips of some growing neurites. These results might suggest that mutations in doublecortin not only disrupt neuronal migration but also axonal pathfinding in differentiating postmigratory neurons 8. If this is the case, it is reasonable to imagine that the R(196)S substitution, could allow neuronal migration to proceed relatively normally but disturb the growth of axons and dendrites, affecting connectivity. This hypothesis might explain the severe epilepsy, due to abnormal axonal and dendritic networks which may act as trigger zones for seizures and contribute directly to hyperexcitability in the cerebral cortex. In either pathophysiological situation, this case report highlights the necessity of performing good brain MRI with thin sections (3 mm or less) in multiple planes allowing carefull analysis of the cortical mantle in case of so-called cryptogenic epilepsy, especially in adult females, to detect subtle neuronal migration defects. Then screening for mutations in doublecortin or other genes involved in corticogenesis should be performed in order to propose accurate genetic counseling.

5 Subtle cortical dysgenesis and doublecortin mutation 277 ACKNOWLEDGEMENTS The authors are grateful to Dr Catherine Adamsbaum, Dr Francis Brunelle, Dr Perrine Plouin and Oliver Dulac for fruitful discussions. This work was supported in part by a grant from The Assistance Publique- Hôpitaux de Paris, the Fondation Jérôme Lejeune and the Foundation pour la recherche médicale. 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: Prasad, A., Prasad, C. and Stafstrom, C. Recent advances in the genetics of epilepsy: insights from human and animal studies. Epilepsia 1999; 40: Pilz, D., Matsumoto, N., Minnerath, S., Mills, P., Gleeson, J., Allen, K. et al. LISI and XLIS (DCX) mutations cause most classical lissencephaly, but different patterns of malformation. Human Molecular Genetics 1998; 7: des Portes, V., Francis, F., Pinard, J. M., Desguerre, I., Moutard, M. L., Snoeck, I. et al. Doublecortin is the major gene causing X-linked subcortical laminar heterotopia (SCLH). Human Molecular Genetics 1998; 7: Fox, J., Lamperti, E., Eksioglu, Y., Hong, S., Feng, Y., Graham, D. et al. Mutations in filamin 1 prevent migration of cerebral cortical neurons in human periventricular heterotopia. Neuron 1998; 21: des Portes, V., Pinard, J. M., Billuart, P., Vinet, M. C., Koulakoff, A., Carrié, A. et al. Identification of a novel CNS gene required for neuronal migration and involved in X-linked subcortical laminar heterotopia and lissencephaly syndrome. Cell 1998; 92: Gleeson, J. G., Allen, K. M., Fox, J. W., Lamperti, E. D., Berkovic, S., Scheffer, I et al. Doublecortin, a brain-specific gene mutated in human X-linked lissencephaly and double cortex syndrome, encodes a putative signaling. Cell 1998; 92: Francis, F., Koulakoff, A., Boucher, D., Chafey, P., Sehaar, B., Vinet, M. C. et al. Doublecortin is a developmentally regulated, microtubule-associated protein expressed in migrating and differentiating neurons. Neuron 1999; 23: Gleeson, J., Lin, P., Flanagan, A. and Walsh, C. Doublecortin is a microtubule-associated protein and is expressed widely by migrating neurons. Neuron 1999; 23: Barkovich, A., Guerrini, R., Battaglia, G. et al. Band heterotopia: correlation of outcome with magnetic resonance imaging parameters. Annals of Neurology 1994; 36: