Researchers report that genetic tests for Friedreich’s ataxia (FA) are complex and may require additional screening in cases of typical clinical presentation.
The study, “Pitfalls in molecular diagnosis of Friedreich ataxia,” was published in the European Journal of Medical Genetics.
Friedreich’s ataxia is caused by mutations in the FXN gene, which provides instructions to build frataxin, a protein with a key role in the proper functioning of nerves and muscle cells. People with FA have reduced levels of frataxin and typically show difficulty walking due to loss of muscle coordination, which spreads to other parts of the body.
Frataxin is a protein found in mitochondria, which produces energy in cells. The most common mutation in FA is an abnormal expansion of a GAA triplet repeat in the FXN gene. G and A are nucleotides — the building blocks of DNA — and stand for guanine and adenine, respectively.
The vast majority of Friedreich’s ataxia patients have this GAA expansion in both FXN gene copies.
Molecular diagnosis in FA, either to confirm clinical diagnosis or to identify mutation carriers, assesses the length of the GAA expansion, which can go as high as 1,700 triplets, resulting in impaired FXN gene expression.
Specifically, a genetic diagnosis of FA includes a two-step RNA analysis called long-range (LR) and triplet-repeat primed (TP) PCR, which spots the presence of a normal or expanded gene. However, this method cannot differentiate between individuals with one or two copies of the defective FXN gene.
The research team presented the case of a 15 year-old girl who had cerebellar ataxia (damage in a brain area called the cerebellum), sensory neuropathy (nerve disease), and cardiomyopathy — diseases of the heart muscle — since age 10. The patient showed cerebellar atrophy through magnetic resonance imaging.
Genetic analysis showed that the patient had no GAA expansion. But one year later, DNA sequencing revealed that she had mutations in the FXN gene. Specifically, the patient showed deletions of exons 4 and 5 of FXN.
“This deletion has already been reported in severe and early-onset [FA] cases,” the investigators wrote.
The same alterations were found in her mother’s DNA. Exons are the bits of DNA that contain the information that generates mature RNA molecules and proteins.
Researchers then repeated the LR-PCR, which now revealed a barely detectable expansion of 830 triplets. However, the other analysis step, TP-PCR, still led to normal results. This discrepancy was caused by the normal size of the gene copy, despite the presence of GAA expansions.
Modification of this TP-PCR step finally showed the same GAA expansion profile as LP-PCR, as well as other alterations.
“This case highlights the complexity of FXN molecular diagnosis,” the scientists wrote.
“Genetic tests using TP-PCR should be interpreted with caution, and clinicians in charge of [FA] patients, should be aware of these limitations, and ask for additional molecular testing when clinical presentation is typical,” they added.