Leukodystrophies (LDs) are heterogeneous genetic disorders characterized by abnormal white matter in the central nervous system. Some of the LDs are progressive and often fatal. In general, LD is primarily diagnosed based on the neuroimaging; however, definitive diagnosis of the LD type is done using genetic testing such as next-generation sequencing. The aim of this study is to identify the genetic causes of LD in two independent Jordanian cases that exhibit MRI findings confirming LD with no definitive diagnosis using whole exome sequencing (WES). The most likely causative variants were identified. In one case, the homozygous pathogenic variant NM_000049.2:c.914C>A;p.Ala305Glu, which is previously reported in ClinVar, in the gene
Leukodystrophies (LDs) are a group of heterogeneous inherited metabolic disorders affecting the white matter and caused by defect in the myelin sheet [
Thirty types of leukodystrophies are described with specific clinical characteristics and genetic causes [
List of the LD diseases and the associated inheritance patterns.
LD | Inheritance |
---|---|
Pol-III related disorders (4H syndrome (hypomyelination, hypodontia and hypogonadotropic hypogonadism)) | AR |
18q minus syndrome | AD |
X linked Adrenoleukodystrophy | XLD |
Adult onset leukodystrophy with neuroaxonal spheroids and pigmented glia (including hereditary diffuse leukoencephalopathy with spheroids, HDLS, and pigmentary type of orthochromatic leukodystrophy with pigmented glia, POLD) | AD |
Aicardi–Goutières Syndrome | AD |
Alexander disease | AD |
Autosomal Dominant Leukodystrophy with Autonomic disease | AD |
Canavan disease | AR |
Cerebrotendinous Xanthomatosis | AR |
Chloride Ion Channel 2 (ClC-2) related leukoencephalopathy with intramyelinic oedema | AR |
eIF2B related disorder (Vanishing White Matter Disease or Childhood ataxia with central nervous system hypomyelination (CACH)) | AR |
Fucosidosis | AR |
Globoid cell Leukodystrophy (Krabbe) | AR |
Hypomyelination with atrophy of the basal ganglia and cerebellum (H-ABC) | AD |
Hypomyelination with brainstem and spinal cord involvement and leg spasticity (HBSL) | AR |
Hypomyelination with congenital cataract (HCC) | AR |
Leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation (LBSL) | AR |
Leukoencephalopathy with thalamus and brainstem involvement and high lactate (LTBL) | AR |
Megalencephalic Leukoencephalopathy with subcortical cysts (MLC) | AR |
Metachromatic leukodystrophy (MLD) and its biochemical variants | AR |
Oculodentodigital dysplasia | AD |
Pelizaeus Merzbacher disease (PMD) | XLR |
Pelizaeus Merzbacher like-disease (PMLD) | AR |
Peroxisomal Biogenesis disorders (including Zellweger, neonatal Adrenoleukodystrophy and Infantile Refsum) | AR |
Polyglucosan Body Disease (PGBD) | AR |
RNAse T2 deficient leukoencephalopathy | AR |
Sialic acid storage disorders (Salla disease, Infantile Sialic Acid Storage Disease and Intermediate form) | AR |
Single enzyme deficiencies of peroxisomal fatty acid beta oxidation (including only D-Bifunctional Protein Deficiency; Sterol Carrier Protein X (SCPx) deficiency; Peroxisomal acyl-CoA-Oxidase Deficiency) | AR |
Sjögren–Larsson syndrome | AR |
SOX10-associated PCWH: peripheral demyelinating neuropathy, central dysmyelinating leukodystrophy, Waardenburg syndrome, and Hirschsprung disease | AD |
Recognizing the specific type of LDs is challenging because of the limited knowledge about their etiology as all types share white matter signals on brain MRI (the term
Patients are often presented to the neurologists with concern to LD based on the abnormal neuroimaging. However, several other clinical features alert the clinicians for the possibility of LD including adrenal insufficiency, endocrine disturbances, ophthalmologic abnormalities, cortical visual impairment, hypodontia and oligodontia, dysmorphic facial features, tendinous xanthomas, skeletal impairment, hearing impairment, hepatosplenomegaly, cutaneous abnormalities, ovarian dysgenesis, and gastrointestinal symptoms [
Many genes have been identified that cause myelin defects by genetic linkage analysis or by next-generation sequencing (NGS) [
In Jordan, the epidemiological data on LD are very scarce and dozens of LD cases in Jordan are with unknown genetic causes. Therefore, this study aims to identify the genetic causes of LD in two affected girls of two unrelated consanguineous families in Jordan using whole exome sequencing (WES).
Two patients (TF106_1 and TF107_1) were enrolled in this study at the Department of Biotechnology and Genetic Engineering at Philadelphia University in Jordan. The two patients were clinically diagnosed with LD based on the brain MRI. The two patients are unrelated girls; TF106_1 belongs to first-cousin parents once removed, and TF107_1 belongs to first-cousin parents (Figure
Pedigrees of the studied cases. The dead proband TF106_1 belongs to first-cousin parent once removed. The patient TF107_1 belongs to a first-cousin parent.
Blood samples were collected from the two patients, their parents, and their healthy siblings. Total Genomic DNA was extracted according to the standard protocols of the Qiagen FlexiGene DNA kit.
Because the majority of monogenic diseases can be detected in the coding part of the genome, the DNA samples of the two patients underwent WES using Illumina NOVASEQ6000 platform (Illumina Inc., San Diego, CA, USA). Exomes were captured using SureSelectXT Library Prep Kit (Agilent Technologies, USA). The sequencing reads (150 bp pair end) were mapped to the reference genome (UCSC hg19) using the Burrows-Wheeler Aligner software [
To identify possible disease-causing mutations, all high-quality variants were identified that are located in the protein coding region (according to Ensembl database v68) and/or two base pair flanking splice sites. We maintained only the variants meeting the following quality criteria: (1) at least 10X coverage and (2) mapping quality score ≥60.
CNV deletions and duplications were checked using the whole exome sequence (WES) coverage data based on the used WES pipeline (megSAP;
Rare homozygous variants with minor allele frequency (MAF) ≤ 0.01 were maintained. Filtration was then based on predicted effects of the variants on the protein, maintaining only loss of function (LOF) variants (stop gain, frameshift, splice site acceptor, and splice site donor) and nonsynonymous variants predicted to be probably damaging or possibly damaging by Polyphen2 Humvar [
Sanger sequencing was performed to segregate and confirm the candidate variants. Primers were designed using primer3 version 4.1.0 [
Besides the LD, as reported in the MRI findings (not shown) of the two patients, patient TF106_1 exhibits atonia in her muscles (neck, arms, and legs), blindness, and seizure. The patient died by the age of 23 months. Patient TF107_1 exhibits the following phenotypes progressing slowly: muscle weakness, unsteady gait, and mental deterioration.
CNV deletions and duplications were checked first based on the WES coverage data and no significant deletions or duplications were detected in both patients.
The total number of variants that were revealed after the filtration procedure was 6. The prioritized variant is NM_000049.2:c.914C>A;p.Ala305Glu in the gene
Segregation analysis of the identified variant in the patient TF106_1.
Canavan disease is neurodegenerative disease that belongs to LDs. And debate is still for the pathophysiology between the accumulation of NAA and demyelination [
Nine variants were revealed after the filtration procedure. One variant NM_000487.5:c.256C>G;p.Arg86Gly in the gene
Segregation analysis of the identified variant in the patient TF107_1.
The phenotypes of the patient show significant overlap with the clinical synopsis of metachromatic leukodystrophy including mental deterioration, loss of speech, hypotonia, muscle weakness, gait disturbances, ataxia, and demyelination [
In summary, the studied patients were genetically diagnosed with Canavan disease and metachromatic leukodystrophy. The two diseases are caused by the lack of important enzymes and thus result in LD. The clinicians in Jordan diagnosed the two independent causes with LD without definitive diagnosis to the type of LD. Here, with the aid of next-generation sequencing (NGS) the genetic causes of the two diseases were assigned. This will further help the two families to avoid having more children with similar diseases and will alert the carrier healthy siblings for their future. Together with the few studies that were conducted in Jordan to identify the genetic causes of neurodevelopmental diseases [
The data used to support the findings of this study are included within the article.
The author declares that there are no conflicts of interest.
The author would like to thank the participating families. Thanks also go to the lab technicians: Aya Baraghiti and Tala Issa. This study was supported by Philadelphia University in Jordan.