Identification of pathogenic expansions within FXN in individuals with suspected Friedreich Ataxia diagnoses
Laboratory Genetics and Genomics
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Primary Categories:
- Laboratory Genetics
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Secondary Categories:
- Laboratory Genetics
Introduction:
Friedreich Ataxia (FA) is an autosomal recessive neurodegenerative disorder caused by biallelic pathogenic variants in the FXN gene. Major clinical features of FA typically present before the age of 25, and include progressive ataxia of the gait and limbs, dysarthria, dysphagia, decreased proprioception, distal muscle weakness, spasticity, scoliosis, pes cavus, and hypertrophic cardiomyopathy. An estimated 96% of individuals with FA have biallelic pathogenic GAA trinucleotide repeat expansions (greater than 65 repeats) within intron 1 of FXN, while the remaining 4% are typically compound heterozygous for one expanded allele and one pathogenic sequence variant on the second allele. FXN GAA repeats of 33 or less are considered normal and have no risk of disease. Premutation alleles (34-43 repeats) are not associated with disease but are at risk for expanding into the pathogenic range in the subsequent generation. Borderline alleles (44-65 repeats) are also at risk for expansion in the next generation, but have mixed evidence as to whether they can cause a mild form of FA if the opposite allele has a pathogenic expansion. A diagnosis of FA can be confirmed with genetic testing, however, there are barriers to receiving testing and diagnosis, including cost. To address this need, Biogen’s FA Identified program offers no-cost Friedreich Ataxia testing for patients 16 years of age or older who are suspected of having FA, or have a clinical diagnosis of FA.
Methods:
In sponsorship with Biogen’s FA Identified program, we have developed a PCR-based assay capable of detecting pathogenic FXN GAA repeat expansions and accurately sizing up to 65 repeats in length. Our test involves two amplicon-length PCR assays from both the 3’ and 5’ ends of the GAA repeat region, and a 5’ repeat-primed PCR assay with a locus-specific primer. Testing for sequence variants and copy number variants within the FXN gene is performed via NGS reflexively automatically for patients with a single positive GAA repeat expansion, to assess the other allele for a second causative variant.
Results:
From May 3rd to October 8th, 2024, 71 individuals participated in Biogen’s FA Identified program. Of the 71 individuals tested, 18 were positive for FA, 4 were carriers for one expanded FXN allele, and 49 were negative for any expanded FXN alleles. Of the 18 individuals positive for FA, 17 had two expanded GAA repeats, and one individual had one expanded GAA repeat and one loss-of-function sequence variant. The average age of those receiving a positive molecular FA diagnosis was 42.2 years of age, with ages ranging from 17 to 75. 77.8% of individuals given a positive molecular FA result were previously clinically diagnosed with FA, while 5.6% of individuals were previously diagnosed with hereditary ataxia, 5.6% had been given a dual diagnosis, and 16.7% of individuals had no prior diagnosis. Of the individuals who reported the age of onset at which their symptoms began (N=12), the average age was 20.3 years. All individuals given a positive FA diagnosis reported comorbidities, with the most common being ataxia (100%), muscle weakness (77.8%), absent reflexes (55.6%), and dysarthria (50%). Examining provider demographics, we found that 72.2% of providers ordering testing through the FA Identified program are neurologists.
Conclusion:
Our PCR and NGS-based test designed to detect pathogenic GAA repeat expansions and sequence variants within the FXN gene was successful in providing 18 positive test results to individuals known to have, or suspected of having, Friedreich Ataxia. The FA Identified sponsored testing program has presented a unique opportunity to understand the milieu of clinical characteristics of individuals with a positive FA diagnosis, and the health care providers seeking such testing for their patients.
Friedreich Ataxia (FA) is an autosomal recessive neurodegenerative disorder caused by biallelic pathogenic variants in the FXN gene. Major clinical features of FA typically present before the age of 25, and include progressive ataxia of the gait and limbs, dysarthria, dysphagia, decreased proprioception, distal muscle weakness, spasticity, scoliosis, pes cavus, and hypertrophic cardiomyopathy. An estimated 96% of individuals with FA have biallelic pathogenic GAA trinucleotide repeat expansions (greater than 65 repeats) within intron 1 of FXN, while the remaining 4% are typically compound heterozygous for one expanded allele and one pathogenic sequence variant on the second allele. FXN GAA repeats of 33 or less are considered normal and have no risk of disease. Premutation alleles (34-43 repeats) are not associated with disease but are at risk for expanding into the pathogenic range in the subsequent generation. Borderline alleles (44-65 repeats) are also at risk for expansion in the next generation, but have mixed evidence as to whether they can cause a mild form of FA if the opposite allele has a pathogenic expansion. A diagnosis of FA can be confirmed with genetic testing, however, there are barriers to receiving testing and diagnosis, including cost. To address this need, Biogen’s FA Identified program offers no-cost Friedreich Ataxia testing for patients 16 years of age or older who are suspected of having FA, or have a clinical diagnosis of FA.
Methods:
In sponsorship with Biogen’s FA Identified program, we have developed a PCR-based assay capable of detecting pathogenic FXN GAA repeat expansions and accurately sizing up to 65 repeats in length. Our test involves two amplicon-length PCR assays from both the 3’ and 5’ ends of the GAA repeat region, and a 5’ repeat-primed PCR assay with a locus-specific primer. Testing for sequence variants and copy number variants within the FXN gene is performed via NGS reflexively automatically for patients with a single positive GAA repeat expansion, to assess the other allele for a second causative variant.
Results:
From May 3rd to October 8th, 2024, 71 individuals participated in Biogen’s FA Identified program. Of the 71 individuals tested, 18 were positive for FA, 4 were carriers for one expanded FXN allele, and 49 were negative for any expanded FXN alleles. Of the 18 individuals positive for FA, 17 had two expanded GAA repeats, and one individual had one expanded GAA repeat and one loss-of-function sequence variant. The average age of those receiving a positive molecular FA diagnosis was 42.2 years of age, with ages ranging from 17 to 75. 77.8% of individuals given a positive molecular FA result were previously clinically diagnosed with FA, while 5.6% of individuals were previously diagnosed with hereditary ataxia, 5.6% had been given a dual diagnosis, and 16.7% of individuals had no prior diagnosis. Of the individuals who reported the age of onset at which their symptoms began (N=12), the average age was 20.3 years. All individuals given a positive FA diagnosis reported comorbidities, with the most common being ataxia (100%), muscle weakness (77.8%), absent reflexes (55.6%), and dysarthria (50%). Examining provider demographics, we found that 72.2% of providers ordering testing through the FA Identified program are neurologists.
Conclusion:
Our PCR and NGS-based test designed to detect pathogenic GAA repeat expansions and sequence variants within the FXN gene was successful in providing 18 positive test results to individuals known to have, or suspected of having, Friedreich Ataxia. The FA Identified sponsored testing program has presented a unique opportunity to understand the milieu of clinical characteristics of individuals with a positive FA diagnosis, and the health care providers seeking such testing for their patients.