Genetic disorders are individually rare, but when you add them up, they’re relatively common in pediatric neurology. They can underlie epilepsy, neuromuscular disease, stroke, autism, intellectual disability, inflammatory brain disease, white matter disease and movement disorders.
A molecular diagnosis can spare children from further unnecessary diagnostic testing and allows for anticipatory guidance. Test results can help guide family planning, and may allow for preimplantation genetic diagnosis if parents are pursuing in vitro fertilization (IVF). A genetic diagnosis can also directly affect therapy, as in inborn errors of metabolism and channelopathies for which treatments are already available.
When should you consider genetic testing, and where should you start?
Neurogenetic testing: the challenges
There are more than 6000 known genetic disorders, many having some neurologic manifestations. In considering testing, it’s good to keep in mind:
- Genetic disorders often have a spectrum of symptoms and severity, which can make them hard to recognize clinically. Different genetic alterations in a gene can produce different effects on the encoded protein, and an individual’s genetic milieu, environment and other factors can modify the effects of a genetic variation.
- Many neurogenetic disorders are not yet described. New disorders are literally being reported on a weekly basis, accelerated greatly by whole-exome sequencing.
- Some mutations are somatic and do not occur in every cell, making them harder to detect with standard genetic testing.
- Some patients have more than one disorder.
- Testing can sometimes uncover “variants of uncertain significance,” or genetic changes whose effects require further investigation.
- Many genetic tests cost in the thousands of dollars, making access an issue. A patient’s insurance coverage may limit what you can test for.
- Families should receive genetic counseling if tested.
Genetic testing methodologies
Approaches to genetic testing range from sequencing a single suspect gene to chromosome-level analyses to whole exome sequencing. There is no “best” test, just the right test for the right situation.
When there is a specific phenotype or known family history, there are several approaches to testing a suspected disease-related gene:
Gene sequencing can identify base substitutions or small insertions/deletions in the protein-coding regions (exons) of the gene. This test will generally not detect changes in non-coding regions, large deletions/duplications or trinucleotide repeats. Techniques include Sanger sequencing and next generation sequencing.
Deletion/duplication analysis (left) can detect larger DNA deletions and duplications (exon size and up). Disorders where such copy number changes may be seen include muscular dystrophy (dystrophin gene), epilepsy associated with the SCN1A gene and Charcot Marie Tooth disease type 1 (PMP22 gene). Methods include array-based comparative genomic hybridization (aCGH) and multiplex ligation-dependent probe amplification (MLPA).
Trinucleotide repeat testing identifies multiple, consecutive copies of certain codons (three-base sequences) in a section of DNA. There are more than a dozen trinucleotide repeat disorders, including Fragile X syndrome, Huntington disease, myotonic dystrophy and some spinocerebellar ataxias.
Methylation analysis looks at markers that regulate gene expression of certain genes. It can be useful in diagnosing certain neurogenetic imprinting disorders such as Angelman and Prader Willi syndromes.
Chromosome microarray (CMA) can pick up DNA deletions or duplications that encompass multiple genes. Examples include 1p36 deletion, 4p terminal deletion and 22q11.2 deletion. Some platforms can also pick up smaller, exon-sized losses or gains in DNA. However, they won’t pick up balanced rearrangements, which change the location or orientation of chromosome material without any measureable loss or gain of DNA. Some CMA platforms include SNP analysis, which can detect regions of homozygosity (in which maternal and paternal alleles are identical). Such analysis can facilitate identifying autosomal recessive disorders.
Karyotyping can identify abnormal numbers of chromosomes in a cell, chromosome rearrangements or very large losses/gains. Karyotyping can detect such abnormalities as trisomy 21, Turner syndrome, ring chromosome 20 and chromosome translocations and inversions. If applied to a large number of cells, karyotyping can detect mosaicism, or chromosome abnormalities affecting only a percentage of a patient’s cells. Unlike CMA, karyotyping can detect balanced chromosome rearrangements, but it will not detect smaller chromosome abnormalities (under 5 mega-base pairs in size).
Fluorescence in-situ hybridization (FISH) uses fluorescent probes that stick to specific DNA sequences on chromosomes. It can tell how many copies of that chromosome segment are present in the cell (normal is two) and can be used when a specific chromosome abnormality is suspected clinically. FISH can detect copy number changes as small as 40-250 kilobases.
Gene panels, typically next generation sequencing (NGS) panels, sequence a group of selected genes known to be associated with a given phenotype. Examples include panels for epilepsy, ataxia, dystonia and leukodystrophy. These tests typically take two to three months to return results, whereas single-gene sequencing typically takes about one month. Panels will not detect exon-level deletions or duplications in the included genes (unless they explicitly include deletion/duplication analysis). Since they test a large number of genes, panels often generate variants of uncertain significance.
Whole exome sequencing (WES) covers all 20,000+ protein-coding genes in a single test. Typically a sample is taken from the affected child and both parents. Consider WES when:
- the patient’s phenotype is not specific enough for single-gene or panel testing
- the phenotype is unique, possibly representing a previously undescribed disorder or a new or atypical form of a known disorder
- you suspect the patient may have more than one genetic disorder
- the patient has an undiagnosed (suspected) genetic disorder without a clear clinical diagnosis and is medically unstable, requiring rapid, comprehensive testing.
WES has an overall diagnostic yield of about 30 percent, and can lead to the discovery of a new disease gene. However, WES cannot detect exon level deletions or duplications, repeat disorders, mosaicism, disorders of genomic imprinting, or genetic alterations within non-coding regions.
Because WES tests all protein coding genes, it may pick up unexpected but medically actionable findings (such as changes in genes associated with cancer or arrhythmia). Families should receive genetic counseling on the limitations of WES and their right to opt out of receiving incidental findings.
Whole genome sequencing (WGS) adds coverage of noncoding DNA regions. Unlike WES, whole genome sequencing can also detect deletions/duplications, structural chromosome rearrangements (even balanced ones) and changes in non-coding regions such as gene promoters and introns. Only a few labs offer WGS clinically and it is not yet clear what benefit WGS adds if prior WES was negative.
The neurogenetic family history: Deciding which test to pursue
Careful phenotyping and a thorough patient and family history can guide testing. The following are a few general pearls:
Has the mother had three or more spontaneous miscarriages? This can indicate a possible chromosomal disorder, with one of the parents carrying a balanced chromosome rearrangement.
Are the parents older? Having a mother over age 35 increases a child’s risk for aneuploidy (trisomy 18, 21, etc.). Having an older parent (especially an older father) also increases the risk for de novo autosomal dominant disorders.
Are the parents related to each other or from a geographic isolate? If so, consider an autosomal recessive disorder; however, don’t automatically rule out other possible inheritance patterns!
Was the child conceived through IVF or intracytoplasmic sperm injection (ICSI), in which sperm is injected into the egg? If so, consider disorders of genomic imprinting.
Does the child’s condition span multiple organ systems? Are there dysmorphic features? If so, consider chromosome disorders like chromosome deletion syndromes.
Could the child have an inborn error of metabolism? The following features could suggest a potentially treatable genetic metabolic disorder:
- basic biochemical abnormalities like acidosis (especially increased anion gap), hypoglycemia or hyperammonemia
- urine ketones in a newborn
- medically refractory epilepsy without a clear cause
- neonatal seizures without a clear cause
- developmental regression
- multiple organ system involvement, including brain, nerve/muscle, liver, heart, kidney, eyes, hearing, etc.
If you suspect an inborn error of metabolism, I recommend you seek a consultation with a specialist in biochemical genetics.
Next steps: Determining where to send a genetic test
Once you’re ready to proceed with genetic testing, there are a number of helpful websites where you can look up appropriate laboratories, including NextGxDx and GeneTests. You can enter such search terms as “whole exome,” “epilepsy,” “neurology” or “Charcot Marie Tooth” and get a list of labs with prices and turn-around times.
Neurogenetics is complex, but pediatricians and child neurologists should have a basic understanding of the different genetic testing methodologies and awareness of clues in the family history that may suggest a particular inheritance pattern.
To request a consult or to refer a patient, contact the Neurogenetics Program.