Learning point for physicians:
A panel approach using next generation sequencing can provide a genetic diagnosis in a timely and financially efficient manner, rapidly interrogating many genes for a fraction of the cost associated with the traditional gene-by-gene approach. Thorough clinical phenotyping is not redundant, remaining critical for the interpretation of genetic variants identified.
Background
The autosomal recessive cerebellar ataxias (ARCA) are a group of clinically and genetically heterogeneous disorders. Friedreich’s ataxia (FA) is the most common form, although many patients with an apparently recessive or sporadic ataxia have no identifiable genetic aetiology. For these patients, diagnostic testing has traditionally followed a slow gene-by-gene analysis using Sanger sequencing to exclude one of the many dominantly inherited Spinocerebellar ataxias (SCAs) or alternative rarer forms of ARCA such as ataxia telangiectasia.1 In some countries financial constraints or strict clinical criteria preclude more expansive genetic testing.
The absence of a genetic diagnosis leaves patients without an explanation for their disability, families without genetic counseling and the loss of opportunity to identify potential therapies. The increasing availability of next generation sequencing (NGS) in the past five years offers more extensive genetic testing with greater time and financial efficiency2. We describe an Irish family with ARCA investigated over 15 years, traversing the traditional approach and NGS, providing a compelling case for the funding of targeted gene panels through specialty services.
Family history
Three affected siblings in a family of 11 were initially investigated independently through neurology clinics from 1999 to 2015. Their non-consanguineous parents were unaffected. The three siblings developed a slowly progressive spastic ataxia with onset between 32 and 43 years. All had early and prominent dysarthria, frontal cognitive impairment, one had a single seizure. Two siblings had downbeat nystagmus in the primary position and ocular telangiectasia with tortuous conjunctival vessels. MRI imaging demonstrated moderate-to-marked cerebellar atrophy and marked frontal atrophy in the most severely affected sibling.
Initial investigations included standard biochemical assays, all of which were negative. Genetic testing for the most common triplet repeat disorders: FA, SCA1-3,6 and 7 were excluded. Between 1999 and 2015, 15 different single genes were assessed for disease causing mutations by Sanger sequencing in the UK, Germany and the Netherlands without a positive result. Some individual genetic tests were performed at considerable expense, such as €5,162 for SCA15 and €2,527 for the ATM gene (Table 1).
Table 1. Genetic tests performed and associated costs in the ANO10 pedigree
| Test | Year | Patienta,b,c | Cost |
|---|---|---|---|
| FRDA | 2000 | A | €200.00 |
| 2000 | B | €200.00 | |
| 2010 | C | €200.00 | |
| SCA1 | 1999 | A | €184.10 |
| 2000 | B | €184.10 | |
| SCA2 | 1999 | A | €184.10 |
| 2000 | B | €184.10 | |
| SCA3 | 1999 | A | €184.10 |
| 2000 | B | €184.10 | |
| SCA6 | 1999 | A | €184.10 |
| 2000 | B | €184.10 | |
| SCA7 | 2011 | A | €159.6 |
| 2013 | B | €87.75 | |
| SCA12 | 2012 | B | €120.00 |
| SCA14 | 2010 | A | €1795.97 |
| 2012 | B | €1147.50 | |
| SCA15 | 2010 | A | €5162.23 |
| Fragile X | 2007 | A | €200.00 |
| 2007 | B | €200.00 | |
| AOA1 | 2008 | B | €400.00 |
| 2010 | C | €400.00 | |
| AOA2 | 2011 | B | €923.00 |
| ATM | 2004 | B | €2527.00 |
| POLG | 2012 | B | €243.00 |
| SACS | 2012 | C | €1036.00 |
| HSP Panel | 2014 | C | €1023.00 |
| ATX Panel | 2014 | A | €1264.80 |
| ANO10 | 2015 | C | €236.83 |
Abbreviations: FRDA, Friedreich’s Ataxia; ATM, ataxia telangiectasia; SACS, Spastic ataxia; HSP, Hereditary spastic paraplegias; ATX, Alpha-Thalassemia X-Linked.
arepresents tde eldest and first patient tdat presented witd symptoms.
brepresents tde middle sibling and second patient tdat presented witd symptoms.
crepresents tde youngest and tdird patients presented witd symptoms here in tdis case.
In 2015 the 91-gene Oxford Ataxia panel was requested on the eldest sibling (www.ouh.nhs.uk/geneticslab). This revealed homozygous c.132dupA p.(Asp45fs) mutations in ANO10, recently identified as a disease-causing gene in spinocerebellar autosomal recessive ataxia type 10 (SCAR10).3 The total cost of negative genetic testing in all three siblings prior to the ataxia panel was €17 497.85, whereas the cost of a diagnostic single ataxia panel was relatively small at €1,264.80, with confirmatory genetic testing of the specific ANO10-mutation in the other siblings costing just €237.
Discussion
The main limitations of traditional Sanger sequencing have been the restricted parallel sample throughput and difficulties with automation of the process. NGS addresses these limitations, opening up an exciting era in genetic diagnostics that has started with syndrome-specific gene panels and is moving towards more routine exome and whole genome sequencing.4
This family started in a general outpatient clinic where they were independently investigated before moving to a subspecialty ataxia clinic. For 15 years the pursuit of individual genetic tests, limited by commercial availability of particular tests and insufficient clinical experience with rare ARCAs, was unsuccessful and ultimately very costly. The identification of mutations in the ANO10 gene has also offered the opportunity to try a co-enzyme Q10 therapy, based on a single observation in another family.5 Successful diagnosis and the potential, albeit very small, for the identification of a therapeutic strategy further underpins the value of a NGS approach.
Conflict of interest: None declared.
References
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