Genetic testing is increasingly important for investigating suspected inherited neurological conditions. A genetic diagnosis can have a huge impact on patients and also their families. It is important for neurologists to appreciate the presymptomatic and prenatal testing options available to patients and their at-risk relatives once a genetic disorder is diagnosed. Asymptomatic family members can experience considerable psychological distress from the knowledge that they might have inherited a neurodegenerative condition. They may also be concerned about the risk of their children inheriting the condition. Information on reproductive options can provide hope and reassurance. This paper reviews the principles of genetic testing in neurological practice, and how they can be applied in prenatal and preimplantation genetic diagnosis. We explain the basis for direct and exclusion testing, use case examples to illustrate the process by which families are counselled and discuss the ethical implications of reproductive technologies.
- genetic testing
- pre-symptomatic testing
- pre-implantation genetic diagnosis
- prenatal testing
- direct and exclusion testing
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- genetic testing
- pre-symptomatic testing
- pre-implantation genetic diagnosis
- prenatal testing
- direct and exclusion testing
Many neurological disorders have a monogenic (single-gene) cause, and traditionally, genetic testing has been performed via a sequential candidate gene testing approach, which is both costly and time consuming. The advent of next-generation DNA sequencing techniques has meant that genetic testing can be performed by using large-scale gene panels and exome sequencing, leading to an accurate molecular diagnosis through rapidly identifying causative variants.1
For the practising neurologist, this means that genetic testing should be considered early in the diagnostic process if a genetic disorder is suspected. Where genetic testing is not available or declined, symptomatic patients should be offered the opportunity to have their DNA stored, to provide further options in future, both for the individual and their family as new diagnoses and treatments are developed.
Traditionally, there had been a divide between the services provided by clinical genetics and neurology, with neurologists being involved in the diagnosis of symptomatic individuals only. While clinical geneticists are cautious about opening access to presymptomatic testing, neurologists with some genetic training are increasingly becoming involved in the process. Multidisciplinary team working recognises individual expertise and ensures that everyone involved understands the downstream implications of a patient’s molecular genetic diagnosis for other family members.
Genetic diagnostic testing in neurological disease is now likely to involve sequencing of a range of genes associated with the clinical phenotype being evaluated. Identifying a clearly pathogenic variant in a candidate gene can result in a clear confirmation of diagnosis. However, complexities arise when the effect of a variant identified in an affected individual cannot be clearly determined: a so-called ‘variant of uncertain significance’. The more genes that are evaluated in an individual, the more likely a variant of uncertain significance will be found. Current diagnostic practice therefore involves using ‘panels’ of genes of known effect, rather than technologies such as exome sequencing looking at the entire coding sequence or genome sequencing looking at the entire genome. Panels can also be designed to ensure even coverage of sequencing, therefore minimising the risk of missing a pathogenic variant.
If a clearly pathogenic variant is identified in an affected family member, this opens a range of options using reproductive technologies, in particular prenatal or preimplantation diagnosis, to avoid the risk of the condition developing in future children. Asymptomatic first-degree relatives of people with a monogenic disorder may feel conflicted, worrying about the risk to any children they might have, while not wanting to know their own disease status. In such cases, preimplantation exclusion genetic diagnosis may be the answer. The decision-making process is not always easy, and genetic counselling is important to promote informed choice in an individual, as well as balancing an individual’s desire for genetic ignorance with the interests of family members.
ESTABLISHING AN ACCURATE GENETIC DIAGNOSIS
The aim of genetic testing in a suspected monogenic neurological disorder is to identify the underlying pathological variant. Some neurogenetic disorders are partly treatable, including those that resemble degenerative diseases. For instance, autosomal recessive ataxia with vitamin E deficiency is a rare neurological disorder that usually starts in childhood.2–4 The clinical presentation is of progressive ataxia, areflexia, dysarthria and sensory neuropathy. Genetic testing of a symptomatic individual as well as familial studies would be important in the early identification and treatment of the condition with vitamin E, which would improve prognosis, particularly at the early stages of the disease.
In conditions where there is no specific treatment, an accurate molecular diagnosis with sensitive detection and quantification of variants will determine recurrence risk, and therefore allow appropriate genetic counselling to an affected individual and their family.
There are two approaches to genetic testing, depending on whether the individual is symptomatic or at risk of developing the disease. Symptomatic or diagnostic testing is predominantly initiated by the neurologist, where a genetic disorder is suspected based on the patient’s phenotype. Presymptomatic or predictive testing is usually initiated in an asymptomatic individual over the age of 18 years who is deemed competent to consent, after a pathogenic variant has been identified in an affected family member.5
Establishing a diagnosis of an inherited neurological condition may help both in relieving uncertainty for the patient and family, as well as offering other family members the choice to undergo genetic testing. Furthermore, a genetic diagnosis can act as a precursor to decisions regarding reproductive choice. A correct phenotypic diagnosis is important so that the clinician can request analysis using the correct diagnostic gene panel. However, analysis using current technology may miss the causative variant if it is in a region of a gene that is poorly represented on the panel or if it is a large deletion or duplication; in such cases, complementary technologies such as multiplex ligation-dependent probe amplification may give more complete coverage. While more sensitive technologies and multigene panels improve sensitivity in detecting variants, they also increase the likelihood of detecting variants of uncertain significance.6 A key challenge is to try and determine the clinical and biological significance of any variant to the disease process. While further genetics testing in the family may sometimes help interpretation—for example, if an affected family member did not carry the candidate variant leading to the conclusion that the variant is not the cause for the phenotype—often the result remains of uncertain significance and therefore of no clinical use. In such cases, it is very important to avoid overinterpreting the finding and to base clinical decisions on a perceived low probability of finding a variant in a candidate gene and it not being linked to the phenotype.
SYMPTOMATIC (DIAGNOSTIC) TESTING
Pretest counselling requirements differ for diagnostic and presymptomatic testing. Diagnostic testing is performed to confirm or exclude a genetic condition. An accurate clinical diagnosis in a patient is key, as well as identifying any positive family history. In a symptomatic patient with a suspected genetic diagnosis, the clinician should discuss the nature of the test with the patient. Ideally, the patient’s family or caregiver should be included in these discussions due to the implications of a diagnosis for the wider family. However, a balance needs to be drawn between appropriate consent and the demystifying of the process to all first-degree relatives of the patient, which might delay diagnosis. In the UK, there are gene panels available for genetic testing for a wide array of neurological conditions. The UK Genetic Testing Network is an advisory organisation that provides commissioning support to the National Health Service, and a list of diagnostic services in the UK is detailed on their website.7
PRESYMPTOMATIC DIRECT AND EXCLUSION TESTING
Presymptomatic testing facilitates reproductive decision making, allowing an individual to plan a family and avoid the birth of a child at risk of developing a disorder. Direct testing is an assay to look for the pathogenic variant known to be causing the disease in the family. If the variant is present, then the individual found to be carrying it is at high risk of developing symptoms of the disease. Exclusion testing is used when an individual does not want to know his or her own disease status but would wish to exclude a child being born at a similar risk to themselves either through prenatal diagnosis or preimplantation genetic diagnosis.
For an exclusion diagnosis to be performed, DNA samples must be taken from both the at-risk individual and their partner as well as the at-risk person’s parents. Genotyping is carried out using polymorphic markers closely linked to the disease gene locus (see figure 1). The ‘high-risk haplotype’—the pattern inherited from the affected parent—is identified.8 Genetic testing via chorionic villus sampling can then determine whether the fetus has inherited the at-risk haplotype from the at-risk parent. Prenatal exclusion testing was originally developed to allow individuals at 50% risk of Huntington’s disease to terminate a pregnancy at the same risk as themselves. It is acceptable to a small minority of couples. The technique has become much more popular when applied to preimplantation genetic diagnosis cycles as embryos at 50% risk are identified and not implanted. The couple therefore does not need to make a decision on terminating a pregnancy at 50% risk.
Prenatal genetic testing has traditionally been an invasive procedure involving chorionic villus sampling or amniocentesis. Chorionic villus sampling can be undertaken at 11–12 weeks’ gestation under ultrasonographic guidance by aspiration of placental tissue (chorionic villi). It is the standard method used to diagnose single-gene disorders, but carries a miscarriage risk of 1–2%.9 Amniocentesis is typically carried out between 15 and 17 weeks of gestation and involves the aspiration of 10–20 mL of amniotic fluid through the abdominal wall under ultrasonic guidance. It has an associated risk of miscarriage of 0.5–1%.
Preimplantation genetic diagnosis
In contrast to prenatal genetic diagnosis, preimplantation genetic diagnosis is carried out before an established pregnancy. It involves testing biopsy material from embryos that are generated through in vitro fertilisation and intracytoplasmic sperm injection technologies (see figure 2). Preimplantation genetic diagnosis is now possible for several single-gene disorders and chromosome rearrangements.10 The genetic test is focused solely on detecting the haplotype around the pathological variant causing the disease of concern, and no genetic results are obtained for other conditions unless specifically sought. Box 1 11 12 gives current guidelines for preimplantation genetic diagnosis.
Criteria for Preimplantation Genetic Diagnosis
Preimplantation genetic diagnosis, like all reproductive technologies that involve in vitro human embryo manipulation, is strictly regulated by the Human Fertilisation and Embryology Authority (HFEA) under the terms of the Human Fertilisation and Embryology Act. Each condition for which testing is performed requires a treatment licence from the HFEA. A specific list of conditions in which testing is available is specified under the HFEA website http://guide.hfea.gov.uk/pgd/. National Health Service funded access to the technology is managed in a similar way to In-Vitro Fertilisation with treatment criteria and the NHS Scotland Pre-Implantation Genetic Diagnosis and Screening Service Framework for Decision Making11 specifies the following access requirements for National Health Service funded treatment:
accurate genetic test available and licensed by HFEA
referral made before woman’s 39th birthday
Body Mass Index of more than 18.5 and less than 30
Anti-mullerian hormone level >6 pmol/L
negative for HIV, Hep B and C
cohabiting in a stable relationship for more than 2 years
both partners should be non-smokers for 3 months prior to treatment and abstain from smoking and alcohol during treatment
no unaffected or untested children as a couple and one partner has no genetic child.
Broadly similar criteria are defined in the NHS England Clinical Commissioning Policy12
Preimplantation genetic diagnosis testing: advantages and disadvantages
The advantage of preimplantation genetic diagnosis over conventional prenatal testing is that couples do not need to contend with a decision regarding pregnancy termination. Its main disadvantage is the uncertain success of achieving a pregnancy, predominantly due to the limitations on the number of oocytes that can be harvested during each cycle. The technique can be problematic in older women due to the reduction in ovarian reserve that occurs naturally with increasing age. Preimplantation genetic diagnosis is time consuming and expensive and carries a risk of hyperstimulation, which can lead to hospitalisation.13 As with all forms of assisted conception access to UK National Health Service, funding for preimplantation genetic diagnosis is an issue with aspects of ‘postcode lottery’. Most UK regions will not offer the service to couples with an unaffected or untested child.
Reproductive autonomy describes the right for an individual to have children and to minimise the genetic risk of a severe inherited condition being passed on to the next generation. This concept of autonomy also covers an individual’s right to be ‘ignorant’ of their own genetic diagnosis, if that genetic knowledge were to cause them more harm rather than benefit. Genetic counselling promotes informed choice and is an important component in the management of monogenic neurological disorders.
Case 1: myotonic dystrophy
A 46-year-old woman was diagnosed with myotonic dystrophy type 1 following a cataract operation. Her 25-year-old daughter, who was 12 weeks pregnant, asked for referral to genetics as she was concerned about her own history of muscle weakness and the information she had found on the internet about babies with myotonic dystrophy being severely affected. Genetic testing showed that she carried a 250 CTG repeat expansion and she was diagnosed with myotonic dystrophy type 1. She chose to have chorionic villus sampling to exclude the myotonic dystrophy type 1 mutation in her pregnancy, but molecular genetic analysis showed the presence of a CTG expansion of >250 repeats. She was counselled that there was a significant risk that her pregnancy was affected by congenital myotonic dystrophy, but because of the overlap of phenotypes seen with larger repeats, not inevitable. She opted to continue the pregnancy with regular ultrasound scans to look for emerging complications that would confirm the diagnosis of congenital myotonic dystrophy. At 20 weeks, she was noted to have excessive amniotic fluid (polyhydramnios) and opted to terminate the pregnancy.
Congenital myotonic dystrophy
Myotonic dystrophy type 1 is a multisystem disorder characterised by distal muscle weakness and myotonia, cardiac conduction abnormalities, autonomic bowel dysfunction, respiratory compromise, endocrine dysfunction and cataracts. It is caused by an expansion of a CTG trinucleotide repeat in the non-coding region of the dystrophia myotonia protein kinase gene (DMPK). The normal range for the CTG expansion is 5–37 and adult onset of symptoms is seen in association with 50 or more repeats. A more severe form frequently associated with cognitive impairment can develop in childhood associated with 1000 repeats. Congenital myotonic dystrophy is associated with more than 1000 CTG repeats.14 It is the result of the ‘anticipation’ of an increase in the size of the CTG expansion on maternal transmission and can present as polyhydramnios during pregnancy, stillbirth, preterm delivery and in 30–40%, neonatal death. Survivors develop the childhood form of myotonic dystrophy type 1.15
Case 2: amyotrophic lateral sclerosis
A 37-year-old woman was referred to the clinical genetics service to discuss the implications of her brother’s recent diagnosis of amyotrophic lateral sclerosis (ALS). Genetic testing had revealed that he carried a pathogenic variant in the SOD1 gene, I113T. Their paternal uncle had developed ALS, which had presented as a progressive bulbar palsy, and had died of respiratory complications aged 68 years before genetic testing had been carried out. In addition, a female cousin had recently presented at the age of 35 years with features of a slowly progressive muscular atrophy. See figure 3 for family pedigree.
She was counselled that she had a 50% chance of having inherited the I113T variant from her father and that the clinical effect of the variant was variable with some carriers developing classical ALS symptoms and others remaining symptom free into old age. She decided to go ahead with presymptomatic testing, as she was keen to start a family and avoid the risk of a child developing ALS in future. Unfortunately, she tested positive for the I113T SOD1 mutation. The option of preimplantation genetic diagnosis was discussed and accepted. The first round of preimplantation genetic diagnosis produced 12 embryos, of which five carried the high-risk haplotype. Of the embryos with the low-risk haplotype, four had developed sufficiently for implantation and embryo number 3 was transferred. See table 1 demonstrating the different haplotypes.
ALS is a progressive neurodegenerative disease that usually presents with a mixture of upper and lower motor neurone features, although a pure upper motor neurone presentation (primary lateral sclerosis) or a pure lower motor neurone presentation (progressive muscular atrophy) can also occur.16 In most cases, there is a progressive paralysis and death from respiratory failure, typically within 2–3 years of symptom onset.17 Familial ALS, which can be defined as the occurrence of two or more cases of ALS in close blood relatives or cases in which an underlying causative variant has been found in an ALS-associated gene, accounts for 5–20% of cases.18 There is an X linked form of familial ALS (ALS15) caused by variants in the UBQLN2 gene19 as well as very rare recessive forms with onset in early childhood or teens, but most cases of familial ALS in whom a causative variant is found follow an autosomal dominant pattern. Until now, there have been over 20 genes implicated,20 the the most common being the C9ORF72 expansion, carried by over 10% of patients with familial ALS (familial and sporadic), SOD1 and TARDBP. The I113T missense variant in SOD1 accounts for approximately 20% of familial ALS and 3% of sporadic ALS21 22 and is associated with incomplete penetrance and marked phenotypic variability. In one large family, the penetrance (onset of symptoms) was 50% in carriers of the variant by age 60 with one female carrier asymptomatic at the age of 86.23
Case 3: Huntington’s disease
Huntington’s disease is an autosomal dominant neurodegenerative disease, characterised by a progressive motor dysfunction (chorea, dystonia, hypokinesia), cognitive decline and neuropsychiatric disturbance.24 The mean age of onset is 40 years, after which symptoms slowly progress with death occurring around 15 years after onset of disease. It is caused by an expanded and unstable trinucleotide repeat (CAG) in the Huntingtin gene (HTT).25 26 Huntington’s disease is a monogenic disorder with penetrance of alleles of 40 CAG repeats or more near complete by age 70.27
A 30-year-old asymptomatic woman had a family history of the Huntington’s disease with several family members affected. Her father had recently died of the condition, having tested positive for the HTT CAG repeat expansion. She was referred to the clinical genetics clinic with her husband to discuss prenatal exclusion preimplantation genetic diagnosis as she did not wish to know her own disease status, but wished to minimise the risk of a future child developing Huntington’s disease (see box 1). Elective single embryo transfer was used; a low-risk haplotype embryo was successfully implanted and pregnancy was established. She was then transferred to standard obstetric care, including first trimester Down’s syndrome screening at 12 weeks which identified increased nuchal translucency on ultrasound scan as well as positive blood markers. While preimplantation genetic diagnosis can reduce the risk of Huntington’s disease to less than 1%, other genetic conditions are not tested for before implantation. Preimplantation genetic screening of embryos using a chromosome microarray to detect chromosomal imbalances can provide more information, but the technique is not yet in routine preimplantation genetic diagnosis practice. The couple opted for chorionic villus sampling for a definitive diagnosis and the Quantitive Fluorescent-PCR test confirmed that the pregnancy was affected by trisomy 21. Very reluctantly the couple decided to terminate the pregnancy.
In this review, we have described the process of genetic testing and identification of pathological variants in individuals either through presymptomatic or diagnostic testing and how it can then open up a range of reproductive options for the individual, through either prenatal testing or preimplantation diagnosis. Presymptomatic individuals who are at risk of developing severe neurodegenerative disorder may not wish to know their own disease status and therefore may opt for exclusion testing. Through case examples, we illustrate this process. An appreciation of ethical and social issues involved in genetic testing and reproductive technologies is important to neurological practice, and collaborative working with the local clinical genetics team can improve the final outcome for patients.
Individuals at risk or symptomatic of single-gene neurological disorders now have a range of reproductive options, including prenatal diagnosis and preimplantation diagnosis.
An accurate molecular diagnosis in the affected relative is key to reproductive technologies; consider storing DNA on symptomatic individuals where there is a possibility of an underlying genetic cause.
Prenatal testing involves detecting a genetic abnormality before birth, whereas preimplantation genetic diagnosis tests for the genetic abnormality in vitro before transferring the unaffected embryo for pregnancy.
Presymptomatic individuals may opt for exclusion testing.
Promotion of informed choice and patient autonomy is at the heart of reproductive technologies; it is important to be aware of the ethical issues and the need for stringent control of these services.
Competing interests None declared.
Provenance and peer review Commissioned; externally peer reviewed. This paper was reviewed by Simon Hammans, Southampton, UK.
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