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Corresponding author at: King Abdulaziz Medical City, King Saud bin Abdulaziz University for Health Sciences, Jeddah, P.O. Box: 12723, 21483, Saudi Arabia.
Affiliations
King Abdulaziz Medical City, King Saud bin Abdulaziz University for Health Sciences, Jeddah, Saudi Arabia
Center of Excellence in Genomic Medicine Research, Department of Medical Laboratory Technology, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia
This report presented a single PRICKLE1 variant segregating with PME in one family.
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The identified novel variant has never been previously reported as disease-causing.
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Heterozygous mutations in the PRICKLE1 gene may have incomplete penetrance.
Abstract
Purpose
Progressive myoclonus epilepsy (PME) comprises a group of heterogeneous disorders defined by the combination of action myoclonus, epileptic seizures, and progressive neurologic deterioration. Neurologic deterioration may include progressive cognitive decline, ataxia, neuropathy, and myopathy. A number of genes have been identified to cause either isolated PME or diseases that manifest PME. We report a Saudi family with a very rare form of autosomal dominant PME.
Methods
We included two patients from Saudi Arabia with a presumptive clinical diagnosis of PME. The patients were from a family with an affected mother I-2 and two affected siblings proband II-3 and II-4 (a girl and a boy).
Results
Genetic analysis revealed a single variant in the PRICKLE1 gene NM_153026.2: c.251 G > A (p.Arg84Gln). Segregation study was performed using DNA from the parents and two sisters. The same variant was identified in one affected parent (the mother I-2) and the two unaffected sisters II-1 and II-2 while it was absent from the unaffected father I-1.
Conclusion
This gene was linked to both autosomal dominant and autosomal recessive PME. To our best knowledge, this is the first report that demonstrates a single PRICKLE1 pathogenic variant segregating with PME in one family. The novel variant identified in this family has never been previously reported as a disease-causing variant. The presence of the same variant in the unaffected individuals may suggest that heterozygous mutations in the PRICKLE1 gene have incomplete penetrance. Further research is needed to elucidate the penetrance of heterozygous mutations in the PRICKLE1 gene.
Progressive myoclonus epilepsy (PME) comprises a group of rare heterogeneous genetic disorders that are generally manifested as a combination of myoclonic and tonic-clonic seizures with cognitive impairment, ataxia and other cerebellar signs, and other neurologic deficits. They are often encompassed under the broader term “catastrophic epilepsies”, which are invariably associated with significant neurological morbidity and often early mortality. This group also include epileptic encephalopathies [
]. Typically, the myoclonus shows a focal or segmental distribution, and is characterized by an arrhythmic, asynchronous, and asymmetric occurrence, which present at rest but activates with posture, action, or stimuli such as noise, light, or touch. The onset of symptoms is usually in late childhood or adolescence. However, they may affect all ages [
Several diseases attributed to different etiologies contribute to PME, with Unverricht–Lundborg disease as the most common PME. Less common PME disorders include Lafora disease, neuronal ceroid lipofuscinoses, Gaucher disease, action myoclonus renal failure syndrome, sialidoses, and myoclonic epilepsy with ragged-red fibers syndrome. Many of these have similar clinical presentations yet are genetically heterogeneous, making accurate diagnosis difficult, especially in the early stages of the illness. Other challenges include further problems of management including drug treatment (Table 1) [
The majority of genes implicated in PME cause autosomal recessive conditions. However, rare cases show autosomal dominant or mitochondrial inheritance. Interestingly, the PRICKLE1 gene has been linked to both autosomal recessive and autosomal dominant PME [
Pagon R.A. Adam M.P. Bird T.D. Dolan C.R. Fong C.T. Smith R.J.H. PRICKLE1-related progressive myoclonus epilepsy with ataxia. University of Washington,
GeneReviews. Seattle, WA1993
]. In this article, we report a Saudi family with autosomal dominant PME caused by a novel pathogenic variant in the PRICKLE1 gene. To the best of our knowledge, this is the first report that describes autosomal dominant PRICKLE1-related PME affecting more than one member in the same family, and it is the second report that describes this very rare form of PME.
2. Methods
We included two patients from Saudi Arabia with a presumptive clinical diagnosis of PME. The patients were from a family with an affected mother I-2 and two affected siblings proband II-3 and II-4 (a girl and a boy). Unfortunately, the proband III-4 died from aspiration pneumonia and status epilepticus at the age of 24 prior to genetic testing (Fig. 1). All family members were examined physically, neurologically, psychiatrically, and questioned about their daily activities. In addition, affected patients had basic blood work, magnetic resonance imaging (MRI) of the brain, and several electroencephalograms (EEGs) with standardized intermittent photic stimulation and hyperventilation. The patients were followed-up by a neurologist and treated with different anti-epileptic drug protocol.
Fig. 1Pedigree of the members after detailed family history. The variant carriers are marked with an asterisk sign.
The entire family was consented for genetic testing after explanation of pros and cons of such investigations. Genomic DNA was prepared from leukocytes using the QIAamp DNA Blood Mini QIAcube Kit (Qiagen, Valencia, CA) according to the manufacturer’s instructions. Whole exome sequencing (WES) for the proband was performed using the SureSelect Human All Exon kit (Agilent, Santa Clara, CA) for enrichment, and a HiSeq4000 (Illumina) instrument for the actual sequencing; variants were annotated using Annovar (http://annovar.openbioinformatics.org) and in-house ad hoc bioinformatics tools. Specific attention was paid towards epilepsy-associated genes (comprehensive epilepsy panel). The panel consisted of 194 genes, 2830 exons, 502547 bases, median coverage was 221 and percent >15X was 99.8. The panel was targeting all protein coding exons and exon-intron boundaries of all target genes. It also covered a number of mutations located outside the coding regions of these genes. This diagnostic tool covers the majority of epilepsy mutations known to date and it is used to detect mutations such as single nucleotide substitutions and small insertions and deletions (INDELs).
Sanger sequencing to confirm variants was carried out on ABI Prism 3500 Genetic Analyzer (Life Technologies, Carlsbad, CA, USA). Identified variants were classified as per the standards and guidelines recommended by the American College of Medical Genetics and Genomics and the Association for Molecular Pathology [
Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology.
A 50-year-old female I-2 that presented initially as a case of seizures at the age of 12 years with frequent myoclonic jerks and generalized tonic-clonic seizures. Her clinical course was progressive, and her myoclonus was both focal and generalized. She also had a gradually progressive cognitive impairment that started at the age of 35 with ataxia and tremor. At the age of 40, she became wheelchair bound with severe cerebellar deficit including intentional tremor, dysmetria, and dysarthria. She had an MRI of the brain, which was unremarkable. EEG demonstrated frequent bilaterally synchronous polyspike-slow wave epileptiform discharges throughout the record. She was treated with several anti-epileptic drugs and currently she is on valproic acid 500 mg twice daily, levetiracetam 500 mg twice daily, clonazepam 2 mg once daily, and acetazolamide 250 mg once daily. The patient's seizures were controlled with no seizures in the past 8 years.
2.2 Case report 2
A 25-year-old female II-3 that presented initially as a case of seizures at the age of 10 years with frequent myoclonic jerks and generalized tonic-clonic seizures. Her clinical course was progressive, and her myoclonus was both focal and generalized. She had ataxia and tremor in both hands which occur both at rest and action. She had an MRI of the brain, which was unremarkable. EEG demonstrated slow background activity of a moderate degree with frequent episodes of spikes and slow wave discharges (Fig. 2). She was treated with several anti-epileptic drugs and currently she is on valproic acid 400 mg three times daily and levetiracetam 1000 mg twice daily. She is seizure-free for the past 5 years. She also suffers from migraine attacks, which were treated with a nonsteroidal anti-inflammatory drug and beta-blockers.
Fig. 2EEG showing slow background activity of a moderate degree with frequent episodes of spikes and slow wave discharges.
The genetic analysis identified a heterozygous variant in the PRICKLE1 gene; NM_153026.2: c.251 G > A (p.Arg84Gln) (Fig. 3). This variant affects a highly conserved amino acid (Fig. 4). Prickle contains an N-terminal PET domain and three C-terminal LIM domains. Prickle has been implicated in regulation of cell movement in the planar cell polarity (PCP) pathway which requires the conserved Frizzled/Dishevelled (Dsh); Prickle interacts with Dishevelled, thereby modulating the activity of Frizzled/Dishevelled and the PCP signaling. The mutation lies in the PET domain of Prickle1 protein. It is towards N terminal. The PET domain is a protein-protein interaction domain, usually found in conjunction with the LIM domain, which is also involved in protein-protein interactions. The PET containing proteins serve as adaptors or scaffolds to support the assembly of multimeric protein complexes. Furthermore, this mutation has been predicted to be deleterious by computational in-silico analysis tools such as scale-invariant feature transform (SIFT) algorithm [
]. In addition, it has been previously identified in only one of 246,094 South Asian chromosomes by the Exome Aggregation Consortium (ExAC, http://exac.broadinstitute.org; dbSNP rs766439768) but was absent from other databases including 1000 Genomes Project (http://www.1000genomes.org), the NHLBI GO Exome Sequencing Project (ESP; http://evs.gs.washington.edu/EVS), ClinVar database of genotype-phenotype associations (http://www.ncbi.nlm.nih.gov/clinvar), the Leiden Open Variation Database (http://www.lovd.nl), and the Human Gene Mutation Database (http://www.hgmd.cf.ac.uk). Deletion/duplication assay using multiplex ligation-dependent probe amplification (MLPA) did not identify neither deletions nor duplications in PRICKLE1 in the proband. We found a mutation only in PRICKLE1 gene, and no mutations were found related to any other known epilepsy genes during the data analysis. We did not find any other pathogenic or homozygous variants in genes that may cause the disease. Segregation study was performed using DNA from the parents and two sisters. The same variant was identified in one affected parent (the mother I-2) and the two unaffected sisters II-1 and II-2 while it was absent from the unaffected father I-1 (Fig. 5). One more sibling, a deceased brother, was diagnosed with PME, however, no DNA was available to perform genotyping. Based on the overall data and genotype-phenotype correlation, the identified variant was considered the disease-causing in this family. The presence of the mutation and absence of the disease in the two unaffected sisters suggests that this mutation may have incomplete penetrance.
Fig. 3Representative chromograph of PRICKLE1 Sanger sequencing read. The red rectangle indicates the affected nucleotide. Genetic analysis identified this variant in both affected females, the proband and her mother. Genotyping was not performed for the deceased affected sibling due to DNA unavailability (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Fig. 4Multiple species protein alignment view for PRICKLE1 from the University of California, Santa Cruz (UCSC) Genome Browser. The highly conserved amino acid altered in this family (p.Arg84Gln) is indicated by a red rectangle. The high conservation of this amino acid suggests that a change at this position is probably not tolerated and would probably affect protein structure and function (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Fig. 5Representative chromograph of PRICKLE1 Sanger sequencing read of all family members. Genetic analysis identified this variant in both affected females, the proband II-3 and her mother I-2. In addition, the variant was identified in the two unaffected sisters II-1 and II-2.
The PMEs are a group of inherited disorders characterized by the presence of typically refractory myoclonic and tonic-clonic seizures with progressive cognitive and neurologic deterioration. Most of these epilepsies are caused by a pathogenic mutation inherited as an autosomal recessive trait. However, few are inherited as an autosomal dominant trait or through mitochondrial inheritance [
]. Most of the known causative diseases in this group are caused by genes that encode lysosomal proteins with few exceptions (ion channels). Despite advances in knowledge of the etiology of PME disorder, the pathogenic mechanism leading to epilepsy and neurological deterioration remain largely unknown. Finding the cause of rare genetic diseases has been tremendously improved and accelerated by next generation-based clinical genetic tests such as WES [
PME-ataxia syndrome usually presents at the age of 4–5 years with ataxia and later on develop PME phenotype with mild or absent cognitive decline. Although cognitive decline is severe and generally occur early in many forms of PME, in this disorder, intellect is generally preserved or mildly affected [
]. Cerebellar signs usually occur early in the disease with ataxia, tremor, dysarthria, and difficulty walking. Action myoclonus may affect the limbs, face, or bulbar muscles with worsening on action or exposure to the sun. Seizures can be myoclonic or generalized tonic clonic and generally occur during sleep. MRI of the brain is usually normal. EEG usually reveal generalized epileptiform discharges with spike-wave or polyspike-wave activity [
Pagon R.A. Adam M.P. Bird T.D. Dolan C.R. Fong C.T. Smith R.J.H. PRICKLE1-related progressive myoclonus epilepsy with ataxia. University of Washington,
GeneReviews. Seattle, WA1993
Genetic analysis revealed a single variant in PRICKLE1 in this patient. Segregation study identified the same variant in one affected parent (the mother) and the two unaffected sisters while it was absent from the unaffected father. The PRICKLE1 gene (MIM#608,500) is located on chromosome 12q12 and encodes the nuclear receptor prickle planar cell polarity protein 1. The Prickle1 protein has multiple protein-protein interaction domains, such as PET (Prickle, Espinas and Testin), LIM (Lin11, Isl-1 and Mec3) and C-terminal PKH (prickle homologous) [
Identification and characterization of human PRICKLE1 and PRICKLE2 genes as well as mouse Prickle1 and Prickle2 genes homologous to Drosophila tissue polarity gene prickle.
Identification and characterization of human PRICKLE1 and PRICKLE2 genes as well as mouse Prickle1 and Prickle2 genes homologous to Drosophila tissue polarity gene prickle.
]. In addition, PRICKLE1 is a well-established member of the planar cell polarity proteins and is involved in establishing cell polarity during embryonic development [
]. In addition, animal model studies showed that throughout mouse embryonic development Prickle1 is expressed in different brain regions including regions implicated in epilepsy, seizures, and ataxia [
Identification and characterization of human PRICKLE1 and PRICKLE2 genes as well as mouse Prickle1 and Prickle2 genes homologous to Drosophila tissue polarity gene prickle.
The PRICKLE1 gene was linked to different conditions including autosomal recessive and autosomal dominant PME. The mechanism by which mutations in the PRICKLE1 gene contribute to different phenotypes is not completely understood. However, the findings of PRICKLE1 functional studies and its different domains-proteins interactions, might suggest that this phenotypic heterogeneity is attributed to the involvement of the PRICKLE1 domains in multiple cellular networks. In addition, some mutations in Prickle1 were suggested to result in a dominant-negative protein that acts dominant-negatively to inhibit the function of normal Prickle1 protein in neurons [
]. This might explain why the presence of a single mutation in PRICKLE1 is enough to cause an autosomal dominant disease. However, future functional analysis of the effect of the identified variant in this study on PRICKLE1 functions would further elucidate the molecular mechanism underlying PRICKLE1-associated autosomal dominant PME.
Biallelic pathogenic variants in the PRICKLE1 gene were reported to cause autosomal recessive PME [
]. More recently, a novel de novo missense mutation in the PRICKLE1 has been reported to be associated with epilepsy, autism spectrum disorder, and global developmental delay [
] (Table 2). In addition, in-vivo studies showed that null and missense Prickle1 mutations could cause a wide spectrum of structural birth defects described to phenocopy human Robinow syndrome and velocardiofacial syndromes. [
PRICKLE1-related PME is very rare, and to date, three families of Middle Eastern descent and two other unrelated individuals were reported with this rare form of PME [
Pagon R.A. Adam M.P. Bird T.D. Dolan C.R. Fong C.T. Smith R.J.H. PRICKLE1-related progressive myoclonus epilepsy with ataxia. University of Washington,
GeneReviews. Seattle, WA1993
]. Among all the reported PRICKLE1 disease-causing pathogenic variants, only three were previously reported to cause PME (Table 2). Complete penetrance was observed in the original families studied that had PRICKLE1-related PME inherited in an autosomal recessive manner. Tao et al. [
] reported two heterozygous mutations in the PRICKLE1 gene in two unrelated individuals with PME. However, no information on the other family members of these patients was provided. In our study, the siblings of the index case were tested and were positive for the same mutation in the PRICKLE1 gene but were clinically normal although they were older than the index case (29 and 27 years old, respectively), which may indicate that heterozygous mutations in the PRICKLE1 gene have incomplete penetrance.
5. Conclusion
PME-ataxia syndrome is a rare genetic disorder characterized by ataxia, myoclonic and tonic-clonic seizure, and varying degree of neurological disability. PRICKLE1-related PME is an extremely rare condition, and to date, only three families of Middle Eastern descent and two other unrelated individuals were reported in the literature. To our best knowledge, this is the first report that demonstrates a single PRICKLE1 pathogenic variant segregating with PME in one family. The novel variant identified in this family has never been previously reported as a disease-causing variant. The presence of the same variant in the unaffected individuals may suggest that heterozygous mutations in the PRICKLE1 gene have incomplete penetrance. Further research is needed to elucidate the penetrance of heterozygous mutations in the PRICKLE1 gene.
Conflict of interest
The authors declare that they have no conflicts of interest.
Acknowledgments
We thank Miss Noha Aman and Miss Azzah Al Muqati for technical help.
Pagon R.A. Adam M.P. Bird T.D. Dolan C.R. Fong C.T. Smith R.J.H. PRICKLE1-related progressive myoclonus epilepsy with ataxia. University of Washington,
GeneReviews. Seattle, WA1993
Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology.
Identification and characterization of human PRICKLE1 and PRICKLE2 genes as well as mouse Prickle1 and Prickle2 genes homologous to Drosophila tissue polarity gene prickle.
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