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The utility of exome sequencing for genetic diagnosis in a familial microcephaly epilepsy syndrome
- Laura M McDonell†1,
- Jodi Warman Chardon†2,
- Jeremy Schwartzentruber3,
- Denise Foster4,
- Chandree L Beaulieu1,
- FORGE Canada Consortium1,
- Jacek Majewski5,
- Dennis E Bulman1, 6 and
- Kym M Boycott1, 2, 6Email author
© McDonell et al.; licensee BioMed Central Ltd. 2014
Received: 11 November 2013
Accepted: 22 January 2014
Published: 31 January 2014
Despite remarkable advances in genetic testing, many adults with syndromic epilepsy remain without a molecular diagnosis. The challenge in providing genetic testing for this patient population lies in the extensive genetic heterogeneity associated with epilepsy. Even for the subset of epilepsy patients that present with a defining feature, such as microcephaly, the number of possible genes that would require interrogation by Sanger sequencing is extensive and often prohibitively expensive.
We report a family of French Canadian descent with four adult children affected with severe intellectual disability, epilepsy and microcephaly born to consanguineous parents and evaluated by the Genetics Service to provide informed genetic counseling to unaffected family members regarding possible recurrence risks. We used whole-exome sequencing (WES) of DNA from one affected sibling as a first-line diagnostic tool and compared the prioritization of variants using two strategies: 1) focusing on genes with homozygous variants; and, 2) focusing on genes associated with microcephaly. Both approaches prioritized the same homozygous novel frameshift mutation (p.Arg608Serfs*26) in WDR62, a gene known to cause autosomal recessive primary microcephaly. Sanger sequencing confirmed the presence of the homozygous mutation in the other three affected siblings.
WES and subsequent filtering of the rare variants in a single affected family member led to the rapid and cost-effective identification of a novel homozygous frameshift mutation in WDR62, thereby explaining the severe neurodevelopmental disorder in this family and facilitating genetic counseling. Our findings support WES as an effective first-line diagnostic tool in families presenting with rare genetically heterogeneous neurological disorders.
Although up to 70% of patients with epilepsy have an underlying genetic cause , most patients with epilepsy will not receive a molecular diagnosis. Genetic testing for epilepsies is challenging since genetic heterogeneity for this group of conditions is significant. For instance, epilepsy can be caused by mutations in over 260 genes and an accurate diagnosis can be particularly difficult with nonspecific epilepsy phenotypes . Even when epilepsy is associated with additional features, such as microcephaly, the number of genes potentially responsible can still be significant and assessing each gene in turn by Sanger sequencing would be time-consuming and expensive. However, establishing a molecular diagnosis is important for informed genetic counseling, prenatal testing, insight into natural history and increasingly for therapeutic management as there are now many instances where medications are tailored based on molecular results [3–5].
Whole exome sequencing (WES) has emerged as a very successful tool for novel disease gene discovery  and its utility in the rapid and cost-effective analysis of heterogeneous disorders is being established. Both a WES approach and the application of targeted panels of channelopathy or encephalopathy genes have been successful in identifying the molecular cause for many patients with rare epilepsies [2, 7]. In addition, WES may offer a rapid and less expensive first line test to establish a molecular diagnosis for disabled patients residing in remote areas where ready access to standard diagnostic assessments can be more challenging. We report here the utility of WES as the initial diagnostic tool for a consanguineous family with four affected adult children with epilepsy, microcephaly and severe intellectual disability from a remote area of Canada.
Clinical and neuroimaging features of the four MCPH2 patients
Age at assessment (yrs)
Occipito-frontal circumference (cm)
Occipito-frontal circumference (SD)
-4 to -5 SD
-2 to -3 SD
-4 to -5 SD
Self injurious behaviour
Self injurious behaviour
Corpus callosum hypoplasia
Following our preliminary filtering steps that identified 183 rare non-synonymous variants, we examined the sequencing data for homozygous variants as the pedigree was consistent with autosomal recessive inheritance and the parents were distantly related (see Method 1, Figure 2). WES identified ten exome-wide homozygous variants including a rare variant in WDR62, encoding a protein essential to cerebral cortex development and known to cause microcephaly [9–11] (Figure 1B) and thus felt to be a strong candidate gene for the disorder in this family. The WDR62 mutation identified is a novel homozygous single-base pair insertion NM_001083961.1:c.1821dupT) producing a frameshift mutation (p.Arg608Serfs*26) in exon 14 (Figure 1C). The protein-truncating mutation is absent from the 6500 exomes in the NHLBI Exome Variant Server. We next examined the WES data using a second approach to identify all rare variants in genes known to be associated with epilepsy and microcephaly (see Method 2, Figure 2); the same mutation in WDR62 was identified as the only candidate.
The mutation in WDR62 was validated by Sanger sequencing in the other three siblings. After the diagnosis, unenhanced MRI of the brain was performed in both sisters and demonstrated microcephaly without polymicrogyria, cerebellar hypoplasia, cortical thickening or corpus callosum hypoplasia. Patient II-2 died suddenly of a cardiac arrest but was not known to have cardiac disease prior to the fatal event; the family declined an autopsy.
Primary autosomal recessive microcephaly (MCPH, OMIM 251200) is a genetically heterogeneous group of disorders characterized by congenital microcephaly, cognitive impairment and variable epilepsy . Mutations in WDR62 are associated with primary microcephaly-2 (MCPH2, OMIM 604317), which is believed to account for 10% of all cases of MCPH . Clinical features of MCPH2 include epilepsy, marked cognitive impairment, incontinence, sloping forehead and prominent ears. Radiological features are variable and may include polymicrogyria, corpus callosum hypoplasia, heterotopias, pachygyria with cortical thickening, lissencephaly and schizencephaly, although patients can present with isolated microcephaly [14–18]. To date, 34 families have been identified with mutations in WDR62, the reported affected individuals ranging in age from infancy to mid-teens [14–18].We present here the oldest patients known to have WDR62 mutations associated with MCPH, providing additional insight into the natural history and phenotypic spectrum of this disorder. It is unclear whether the sudden cardiac death for Patient II-2 is associated with WDR62 mutations or an unrelated cardiac event. Of the mutations in WDR62 reported thus far, approximately half have been frameshift mutations, as described here, distributed throughout the gene and predicted to result in premature truncation of the protein (Figure 1C).
At least ten genes have been associated with MCPH [ASPM, MCPH1, WDR62, CDK5RAP2, CASC5, CENPJ, STIL, CEP135, CEP152 and ZNF335] and are implicated in proliferation and migration of neuronal progenitors during embryonic cortical development . Clinical-radiological features have not been loci-specific and MCPH patients remain phenotypically indistinguishable . Thus, conventional molecular testing can be a long and expensive process and may not yield a diagnosis. Currently, sequential analysis of the eight clinically available MCPH genes would total more than $22,000 USD. Alternatively, eight of the ten MCPH genes could be sequenced as part of a microcephaly panel for $7,000 USD (http://www.centogene.com accessed November 6th, 2013, eight genes). Comparatively, WES would interrogate all ten microcephaly genes and is currently available in clinical laboratories at $5000 USD per patient, making it a cost-effective solution for rapid diagnosis in the clinic.
We studied four siblings from a consanguineous French Canadian family presenting with microcephaly, severe developmental delay and epilepsy. Given their relatively nonspecific phenotype which did not suggest a single likely candidate gene, sequential gene testing was not undertaken. Alternatively, WES provided a cost-effective and rapid method to identify the genetic cause of epilepsy and microcephaly in this family. We highlight, that particularly for patients in remote areas, WES via DNA blood/saliva offers a non-invasive and readily accessible method for shortening the diagnostic process for these patients.
Informed consent was obtained for participation in the Finding of Rare Disease Genes (FORGE) Canada study. The Research Ethics Board of the Children’s Hospital of Eastern Ontario approved this study in accordance with the Declaration of Helsinki. A copy of the written consent is available for review by the Editor of this journal.
The authors would first like to thank the study patients, their family and their caring guardians, without whose participation this work would not be possible. This work was funded by the Government of Canada through Genome Canada, the Canadian Institutes of Health Research (CIHR) and the Ontario Genomics Institute (OGI-049) (to KMB). Additional funding was provided by Genome Quebec and Genome British Columbia (to KMB). The authors wish to acknowledge the contribution of the high throughput sequencing platform of the McGill University and Génome Québec Innovation Centre, Montreal, Canada. This work was selected for study by the FORGE Canada Steering Committee, consisting of K. Boycott (U. Ottawa), J. Friedman (U. British Columbia), J. Michaud (U. Montreal), F. Bernier (U. Calgary), M. Brudno (U. Toronto), B. Fernandez (Memorial U.), B. Knoppers (McGill U.), M. Samuels (U. de Montreal), and S. Scherer (U. Toronto). LMM is supported by a Frederick Banting Graduate Scholarship from CIHR. KMB is supported by a Clinical Investigatorship Award from the CIHR Institute of Genetics.
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