GAA Expansion Research Articles

The 57th Annual Meeting of the American Academy of Neurology, Miami, Florida, April 9-16, 2005

The 57th Annual Meeting of the American Academy of Neurology, Miami, Florida, April 9-16, 2005

Late-Onset Friedreich Ataxia

Friedreich ataxia (FA), the most common hereditary ataxia, is caused by pathological expansion of GAA repeats in the first intron of the X25 gene on chromosome 9. Since the discovery of the gene, atypical features are increasingly recognized in individuals with FA, and up to 25% of patients with recessive or sporadic ataxia do not fulfill the Harding or Quebec Cooperative Study on Friedreich's Ataxia criteria for FA. Late-onset FA (LOFA) is defined as onset after age 25 years. To describe and further delineate the clinical and magnetic resonance imaging findings in patients with LOFA and to review the literature. Clinical evaluation and comparison of clinical data and investigations. Ataxia clinics at UCLA and Cedars-Sinai Medical Center. Thirteen patients with LOFA with 13 sex-matched and Inherited Ataxia Progression Scale-matched patients with typical FA. Gait and limb ataxias were seen in all the participants. Dysarthria, loss of vibration sense, and abnormal eye movements were also common in both groups. Patients with LOFA more often had lower limb spasticity (40% vs 0%; chi2 = 4.0; P = .04) and retained reflexes (46.1% vs 7.7%; chi2 = 3.46; P = .05). They had no complaint of sphincter disturbances, and there was no evidence of cardiomyopathy on echocardiograms (chi2 = 4.0; P = .04). Five of 9 patients with LOFA had cerebellar atrophy on neuroimaging. Patients with gait and limb ataxias, dysarthria, loss of vibration sense, and fixational instability after age 25 years should be considered for molecular testing for GAA expansion in the FA gene. In contrast to previous studies, cerebellar vermian atrophy is not an uncommon finding.

957. Accurate Humanized Mouse Models of Friedreich Ataxia

Friedreich ataxia (FA) is an autosomal, recessive neurodegenerative disorder caused by a GAA trinucleotide expansion in the first intron of the Friedreich ataxia gene (FRDA) that causes reduced synthesis of frataxin, a mitochondrial protein likely to be involved in biosynthesis of iron-sulfur clusters. Frataxin insufficiency leads to increased oxidative stress, progressive loss of large sensory neurons and hypertrophic cardiomyopathy. Knockout mouse models of FA do not recapitulate accurately the molecular basis of the disease and thus cannot be used to evaluate strategies for overcoming the effects of the GAA expansion. Our strategy is to generate accurate humanised mouse models of FA, which contain the entire human FRDA genomic locus and surrounding regulatory elements and harbors a long GAA expansion as found in FA patients. Such mice will not only manifest the main phenotypic traits or symptoms of a disorder, but also provide the correct underlying molecular cause of the disease as found in patients. A major obstacle in the generation of constructs containing long GAA expansions has been the instability of GAA expansions in bacterial multicopy vectors. By using single copy BAC vectors and optimised bacterial culturing techniques, we have recently successfully cloned and stably maintained GAA expansions containing up to 500 repeats over multiple bacterial subcultures and numerous cell doublings, without any sign of contraction or rearrangements. Using homologous recombination we have introduced a (GAA)500 expansion adjacent to an antibiotic-resistance gene into the first intron of the human FRDA gene present on a fully sequenced BAC clone harboring a 188 kb genomic DNA fragment containing exons 1-5b of the human FRDA gene and surrounding sequences. The modified BAC clone containing the human FRDA gene with a (GAA)500 expansion was used to generate two lines of transgenic mice with low and high transgene copy numbers. This is the longest GAA expansion to be stably maintained in transgenic mice. RT-PCR and Western blot analyses confirmed that the presence of the introduced (GAA)500 expansion results in decreased FRDA gene expression and in lower levels of frataxin protein. Breeding of the transgenic mice with heterozygous Frda knockout mice demonstrated that the human transgene is able to rescue the embryonic lethality of the homozygous KO mice when the transgene is in hemizygous or homozygous form in both lines. Rescue mice are being assessed by a series of behavioural, neurological and histological tests for phenotypic symptoms of FA. These transgenic mice should not only facilitate analysis of the progressive pathophysiology of FA but also permit the evaluation of potential new therapies specifically targeted to overcoming the molecular effects of the GAA expansion.

The Evaluation of Left Ventricular Systolic and Diastolic Functions in Patients With Friedreich Ataxia A Pulse Tissue Doppler Study

Friedreich's ataxia (FRDA), the most common subtype of early onset hereditary ataxia, is an autosomal recessive neurodegenerative disorder caused by unstable GAA expansions. Two-dimensional, pulse, and pulse tissue Doppler echocardiographic examinations were performed on 21 patients with GAA expansion. There was no association between left ventricle ejection fraction, tissue Doppler systolic s wave, and left ventricle diastolic functions examined by pulse and tissue Doppler. The septum thickness of patients with Friedreich's ataxia was significantly increased when compared with that of the control group and wall thickness was found to be associated with GAA repeats. In patients with FRDA, despite a correlation between genetic abnormality with left ventricular early and late diastolic parameters, global diastolic functions were preserved when examined by tissue Doppler.

Striking intrafamilial phenotypic variability and spastic paraplegia in the presence of similar homozygous expansions of the FRDA1 gene

We report on a Friedreich's ataxia (FA) family with 3 affected siblings with markedly different phenotypic presentations, including one with spastic paraplegia. Molecular analysis showed midsize GAA repeat expansion sizes in all 3 individuals. Gait spasticity in FA, although rare, has been described in a few patients who are compound heterozygotes for a point mutation, or who had GAA expansions of less than 200 repeats. The occurrence of spastic paraplegia in our family, in the presence of homozygous midsize GAA repeat expansions, is an unusual finding. Spasticity can be the main feature in both sporadic and familial patients with FA, either as an isolated finding, or in addition to other neurological abnormalities, and should be included as a rare feature in the clinical spectrum of FA. This family also demonstrates that in FA, marked intrafamilial phenotypic variability can arise in the presence of similar GAA expansion sizes. Therefore, in familial FA, the disease course in relatives therefore cannot be predicted solely from repeat length. Factors such as somatic mosaicism, repeat interruptions, modifying mutations and environmental factors must also be considered.

Expanded GAA repeats and clinical variation in Friedreich's ataxia.

One of the main features of Friedreich's ataxia (FA) is phenotypic variability that can now be explained by the molecular mechanism (GAA expansion) underlying the disease. We have analyzed genotype-phenotype correlations in a group of 40 patients homozygous for the GAA expansion. The smaller GAA expansion (GAA1 allele) size correlated with age at onset and progression disease rate, but we found no correlation between the larger GAA expansion (GAA2 allele) size and these clinical parameters. The frequency of pes cavus, scoliosis, axonal sensory neuropathy and areflexia increased with the size of GAA1, whereas some signs such as sphincter disturbances, cerebellar atrophy on MRI, amyotrophy, dysarthria and decreased vibration sense were associated with increased duration of the disease. GAA1 size is the main determinant of FA phenotype and GAA2 size is a poor predictor of clinical variation. Some clinical features are independent of GAA1 and GAA2 sizes and are determined by the duration of the disease.

Friedreich ataxia

There has been rapid progress in the understanding of several aspects of Friedreich’s ataxia (FA) since the gene mutation was identified in 1996. At the clinical level, now it is possible to confirm that the majority of patients fullfilling clinical criteria for classic FA have the FA gene mutation but some do not, indicating genetic heterogeneity. Also, the phenotype associated with the FA mutation is much wider than that defined by clinical criteria and includes ataxia with retained or brisk reflexes as well as late onset ataxia with or without retained reflexes. It is now clear that the unstable GAA expansion that underlies FA causes a deficiency of the mitochondrial protein frataxin, leading to potentially harmful oxidative injury associated with excessive iron deposits in mitochondria. In addition, pathogenesis may involve a primary defect in synthesis of iron-sulfur cluster containing enzymes. Therapeutic attempts are already using anti-oxidant strategies and such efforts are likely to be enhanced by the rapid availability of animal models of the disease.

La maladie de Friedreich chez l'enfant

Through the observation of seven cases we reported the epidemiological, clinical, paraclinical, evolutionary and therapeutical aspects of this disease. Epidemiologicaly, the sex ratio is of 0,75. The medium age is of six years and two months. Factors in favor of a heriditary origin are the consanguinity found in 28,6% of cases and the presence of similar case in the family observed in 42,3%. Clinically, the quasi constantly neurolgical signs are ataxy (100%), dysarthria (71,4%), areflexia of tenden reflex in both loner limles (57,14%), deep sensibility troubles are less frequent (28,6%). Vestibular affection with form of horizontal nystagmus is found in 14,3% of cases. The dysmorphic syndrom in the form of claw found in (57,14%) and of dorsal scoliosis in (14,3%). Cardiac affection in the ferme of cardiopathy is noted in two cases (28,6%). Paraclinically, we found cerebellar atrophy in 57,14% witch was confirmed by magnetic resonance imagery of one patient. The electromyography is perturbed in 28,6% of cases. The genetic study done to two cases showed an expansion of the triplet GAA at the level of the chromosome nine. Improvement under symptomatic treatment based on muscular and fonctional physiotherapy was progressively slow for all our patients.

Iron metabolism in mice with partial frataxin deficiency.

Friedreich ataxia (FRDA), the most common autosomal recessive inherited ataxic disorder, is the consequence of deficiency of the mitochondrial protein frataxin, typically caused by homozygous intronic GAA expansions in the corresponding gene. The yeast frataxin homologue (yfh1p) is required for cellular respiration. Yfh1p appears to regulate mitochondrial iron homeostasis and protect from free radical toxicity. Complete loss of frataxin in knockout mice leads to early embryonic lethality, indicating an important role for frataxin during development. Heterozygous littermates with partial frataxin deficiency are apparently healthy and have no obvious phenotype. Here we evaluate iron metabolism and sensitivity to dietary and parenteral iron loading in heterozygote frataxin knockout mice (Fx(+/-)). Iron concentrations in the liver, heart, pancreas and spleen, and cellular iron distribution patterns were compared between wild type and Fx(+/-) mice. Response to parenteral iron challenge was not different between Fx(+/-) mice and wild type littermates, while sporadic iron deposits were observed in the hearts of dietary iron-loaded Fx(+/-) mice. Finally, we evaluated the effect of partial frataxin deficiency on susceptibility to cardiac damage in the mouse model of hereditary hemochromatosis (HH), the Hfe knockout mice. HH, an iron overload disease, is one of the most frequent genetic diseases in populations of European origin. By breeding Hfe(-/-) with Fx(+/-) mice, we obtained compound mutant mice lacking both Hfe and one frataxin allele. Sparse iron deposits in areas of mild to moderate cardiac fibrosis were found in the majority of these mice. However, they did not develop any neurological symptoms. Our studies indicate an association between frataxin deficiency, iron deposits and cardiac fibrosis, but no obvious association between iron accumulation and neurodegeneration similar to FRDA could be detected in our model. In addition, these results suggest that frataxin mutations may have a modifier role in HH, that predisposes to cardiomyopathy.

Friedreich's ataxia: cardiac evaluation of 25 patients with clinical diagnosis and literature review.

Cardiac evaluation (clinical, electrocardiographic and echocardiographic) of 25 Brazilian patients with clinical diagnosis of Friedreich's ataxia (FA) related to the frequency and the size of GAA repeats (unstable expansion of trinucleotide repeats that results in the disease). Clinical and cardiac study including electrocardiogram and echocardiogram of all patients and molecular analysis to detect the frequency and the size of GAA expansion, by polymerase chain reaction analysis. Homozygous GAA expansion was detected in 17 patients (68%) - all typical cases. In 8 (32%) cases (6 atypical and 2 typical), no GAA expansion was observed, therefore it was not considered Friedreich's ataxia. All patients with GAA expansion (100%) had electrocardiographic abnormalities, and only 25% of the cases without GAA expansion had some abnormality on this exam. However, only 6% of all patients revealed some signals/symptoms suggestive of cardiac involvement. A molecular analysis is essential to confirm the diagnosis of Friedreich's ataxia; however, an adequate cardiac evaluation, including an electrocardiogram, was extremely useful to better screening the patients which should perform these molecular analysis.

Antioxidant enzymes in blood of patients with Friedreich's ataxia

Background and Aims: Increased generation of reactive oxygen species and mitochondrial dysfunction may underlie the pathophysiology of Friedreich's ataxia, the most common inherited ataxia, due to GAA expansion in a...

Clinical comparison between AVED patients with 744 del A mutation and Friedreich ataxia with GAA expansion in 15 Moroccan families

Fifteen Moroccan families with a phenotype resembling Friedreich Ataxia (FA) were studied. Seven families (13 patients) had the 744 del A mutation in the alpha-tocopherol transfer protein (α-TTP) gene, characteristic of ataxia with vitamin E deficiency (AVED). The other eight families (16 patients) had GAA expansions in the first intron of the frataxin gene. The clinical differences between the two groups differed. AVED caused by the 744 del A could be distinguished by head titubation, lower frequency of the neuropathy and slower disease progression, decreased visual activity and retinitis pigmentosa, which has also been associated with a His 101 Gln missense mutation in the α-TTP gene. The neurological disorder associated with vitamin E deficiency can be improved by the alpha-tocopherol treatment.

Friedreich ataxia with minimal GAA expansion presenting as adult-onset spastic ataxia

Around a quarter of Friedreich ataxia (FA) patients, despite being homozygous for GAA expansion within the FRDA gene, show atypical presentations. Our aim is to describe the case of three brothers with long-term follow-up suffering from late onset FA manifested with spastic ataxia. The three patients belong to a family with occipital dysplasia (OD) and Chiari I malformation previously reported by us. We have carried out serial examinations since 1977. Electrophysiological and neuroimaging studies, and molecular genetic analyses of hereditary ataxias are available in all three patients. Onset of symptoms occurred between 25 and 35 years. The clinical picture consisted of progressive spastic gait, truncal and limb ataxia, dysarthria, nystagmus, hyperreflexia with knee and ankle clonus and extensor plantar response, and mild hypopallesthesia. Ages at present vary between 50 and 59. One patient is wheelchair-bound but the other two are able to walk with support. Leaving OD aside, skeletal anomalies are not prominent. All three patients showed cardiomyopathy. MR imaging revealed atrophy of the cerebellum and spinal cord. Motor and sensory nerve conduction velocities were normal. Central conduction time of both motor and sensory pathways was delayed or unobtainable. All three patients were homozygous for the GAA expansion, the smaller expanded allele ranging between 131 and 156 repeats. Four heterozygotic carriers were detected among non-ataxic relatives including one with OD; furthermore, an asymptomatic OD patient showed normal genotype. We conclude that adult onset spastic ataxia is a distinctive FA phenotype associated with minimal GAA expansion. This phenotype represents a new cause of selective distal degeneration of central sensory axons. The present concurrence of OD and FA reflects coincidental cosegregation of two different inherited disorders.

Glutathione in blood of patients with Friedreich's ataxia.

Oxidative stress and mitochondrial dysfunction have long been considered to play a role in Friedreich's ataxia, a neurodegenerative disease due to a GAA expansion in a gene coding for a mitochondrial protein (frataxin), implicated in the regulation of iron metabolism. Since glutathione is an important antioxidant whose role has been recently proposed in the pathogenesis of some neurodegenerative diseases, we investigated glutathione metabolism in the blood of 14 patients with Friedreich's ataxia by measuring total, free and protein-bound glutathione concentrations. Blood samples were obtained from 14 unrelated patients with Friedreich's ataxia (nine males, five females) and 20 age-matched healthy controls (10 males, 10 females). Total and free glutathione concentrations were determined by reverse-phase liquid chromatography with fluorescence detection; the glutathionyl-haemoglobin separation from healthy and pathological subjects was obtained by electrospray ionization-mass spectrometry. We consistently found a reduction of free glutathione levels (0.55 +/- 0.06 nmol mg(-1) haemoglobin, vs. 8.4 +/- 1.79 nmol mg(-1) haemoglobin, P < 0.001) in the blood of patients with Friedreich's ataxia, a total glutathione concentration comparable to the controls (15 +/- 2.6 nmol mg(-1) haemoglobin, vs. 15.4 +/- 1.4 nmol mg(-1) haemoglobin), and a significant increase of glutathione bound to haemoglobin (15 +/- 1.5 vs. 8 +/- 1.8%, P < 0.05) in erythrocytes. Our findings give evidence of an impairment in vivo of glutathione homeostasis in Friedreich's ataxia, suggesting a relevant role of free radical cytotoxicity in the pathophysiology of the disease; this study may also prove useful in the search for an oxidative stress marker in neurodegeneration.

Friedreich's ataxia: clinical and molecular study of 25 Brazilian cases.

Friedreich's ataxia is a neurodegenerative disorder whose clinical diagnostic criteria for typical cases basically include: a) early age of onset (< 20 or 25 years), b) autosomal recessive inheritance, c) progressive ataxia of limbs and gait, and d) absence of lower limb tendon reflexes. We studied the frequency and the size of expanded GAA and their influence on neurologic findings, age at onset, and disease progression in 25 Brazilian patients with clinical diagnosis of Friedreich's ataxia - 19 typical and 6 atypical - using a long-range PCR test. Abnormalities in cerebellar signs, in electrocardiography, and pes cavus occurred more frequently in typical cases; however, plantar response and speech were more frequently normal in this group when the both typical and atypical cases were compared. Homozygous GAA expansion repeats were detected in 17 cases (68%) - all typical cases. In 8 patients (32%) (6 atypical and 2 typical), no expansion was observed, ruling out the diagnosis of Friedreich's ataxia. In cases with GAA expansions, foot deformity, cardiac abnormalities, and some neurologic findings occurred more frequently; however, abnormalities in cranial nerves and in tomographic findings were detected less frequently than in patients without GAA expansions. Molecular analysis was imperative for the diagnosis of Friedreich's ataxia, not only for typical cases but also for atypical ones. There was no genotype-phenotype correlation. Diagnosis based only on clinical findings is limited; however, it aids in better screening for suspected cases that should be tested. Evaluation for vitamin E deficiency is recommended, especially in cases without GAA expansion.

Serum transferrin receptor levels in Friedreich’s and other degenerative ataxias

Friedreich’s ataxia (FRDA) is the most common hereditary ataxia, with a birth incidence of 1 in 25,000. The most frequent cause, present in approximately 98% of FRDA chromosomes, is an abnormal GAA expansion in the first intron of the X25 gene. The presence of this expansion inhibits gene expression, resulting in decreased levels of the encoded protein frataxin. X25 missense, nonsense, and splice site mutations have been identified in the remaining FRDA chromosomes.1 Frataxin is located in mitochondria. The localization has been proved by immunocytofluorescence with monoclonal antibodies, great frataxin enrichment in the mitochondrial pellet obtained by differential centrifugation, and immunoelectron microscopy.2 Even though the exact function of frataxin remains unknown, it seems to be important in mitochondrial iron handling. Deletion of the yeast frataxin homologue, YFH1 , results in a 10-fold increased iron in mitochondria, a reduction in mitochondrial DNA, reduced respiration, increased sensitivity to oxidative stress, and decreased activities of iron-sulfur protein levels. Cytosolic iron is decreased with upregulation of high-affinity iron uptake proteins.3,4⇓ Evidences of iron accumulation in humans are …

Friedreich's ataxia and frataxin: molecular genetics, evolution and pathogenesis (Review).

Friedreich's ataxia is an autosomal recessive neuro-degenerative disorder involving both central and peripheral nervous system. Patients also show a systemic clinical picture presenting heart disease and diabetes mellitus or glucose intolerance. The disease is caused by mutations in the FRDA gene mapped on chromosome 9q13. The product of the gene is frataxin, an 18 kDa soluble mitochondrial protein with 210 amino acids. Crystal structure suggests a new, not previously reported, protein fold. The most frequent mutation is the expansion of a GAA trinucleotide repeat located within the first intron of the gene, and represents 98% of the mutations. Point mutations are described in compound heterozygous subjects with one expanded allele. A two-step model of GAA normal alleles towards premutation alleles, which might generate further full expanded mutations in the population with Indo-European ancestry, has been postulated. Clinical phenotype is variable and an inverse correlation with the GAA expansion size has been observed. Analysis of the GAA triplet is a strong molecular tool for clinical diagnosis, genetic counselling and prenatal diagnosis. Friedreich's ataxia patho-genesis is not solved yet. Substantial data from organism models, such the S. cerevisae yeast and more recently conditioned knock-outs in mouse, and studies in heart biopsies and fibroblast cultures from patients suggest an important role of mitochondrial iron in the development of the disease. Iron is accumulated in the mitochondrial matrix of both the yeast frataxin deficient mutant and the patient fibroblasts. It has been postulated that iron-induced oxygen radical affects the oxidative phosphorylation in frataxin deficiency states favouring the disease pathology. A second hypothesis postulates a direct role of frataxin in the mitochondrial energy activation and oxidative phosphorylation. Iron chelator drugs and antioxidant drugs have been postulated for Friedreich's treatment. No results from clinical trials are available yet, but idebenone, a short-chain quinone, seems to reduce the size of hypertrophic cardiomyopathy and levels of oxidative stress molecules in patients.

Electrophysiological contribution to understanding the pathophysiology of Friedreich's ataxia

Friedreich's ataxia (FA) is the most frequent early onset recessive ataxia. Ataxia clinical progression is slow, but the majority of patients is wheelchair-bounded before 30 years of age. Some studies suggested that the peripheral sensory neuropathy was a dynamic process that could explain the clinical worsening. In 1983 an electrophysiological and histological study showed peripheral nerve axonal degeneration unrelated to disease duration or severity. A follow-up study confirmed that symptom progression is independent from peripheral neuropathy that is not progressive. A defective developmment of the largest neurons of the dorsal ganglion was supposed. When the unstable GAA expansion in the X25 gene was identified as the responsible mutation in the majority of FA patients, genotype-phenotype correlation studies were performed. A direct correlation between GAA expansion and involvement of peripheral nerve sensory fibers and central somatosensory pathway was shown. Disease duration correlates with pyramidal, auditory and visual pathway involvement, while the GAA expansion size has no effect.

Friedreich's ataxia: from the patient to the gene

A GAA expansion in the first intron of the X25 gene is present in 98% of Friedreich's ataxia chromosomes. A point mutation is present in the remaining 2%. A direct correlation between the size of the smaller expanded allele and disease severity is present in patients homozygous for the GAA expansion. The abnormal GAA expansion inhibits the expression of the X25 gene, causing at homozygous state a severe decrease of frataxin. Frataxin has a mitochondrial localization and may be involved in iron handling. The expanded alleles show intergenerational instability with a trend to contraction and a sex bias. The mapping and cloning of the X25 gene have resulted in expansion of the phenotype, and forms with late onset and retained reflexes have been identified.

Increased levels of plasma malondialdehyde in Friedreich ataxia.

Friedreich ataxia is an autosomal recessive disease characterized by spinocerebellar degeneration and cardiomyopathy. It is caused by deficiency of frataxin, a highly conserved nuclear-encoded protein localized in mitochondria.1 Most patients (95%) are homozygous for the hyperexpansion of a GAA repeat sequence in the first intron of the frataxin gene, which inhibits transcription; a few are heterozygous for a GAA expansion and a frataxin point mutation.1 Data from yeast suggest that frataxin deficiency results in mitochondrial dysfunction and free radical damage. Yeast cells with a disrupted frataxin homolog gene ( YFH1 ) show a 10-fold accumulation of iron in mitochondria, lose mitochondrial DNA, and become unable to carry out oxidative phosphorylation.2 Iron in mitochondria can amplify the toxicity of reactive oxygen species (ROS) leaking from the respiratory chain by producing the free hydroxyl radical (OH•) through Fenton chemistry [Fe(II) + H2O2 → Fe(III) + OH• + OH−]. Occurrence of the Fenton reaction in ΔYFH1 yeast cells is suggested by their enhanced sensitivity to H2O2.2 Several lines of evidence support the hypothesis that similar mechanisms are involved in the human disease. First, patients with Friedreich ataxia have iron deposits in the heart …