Funded by the NIH • Developed at the University of Washington, Seattle
Funded by the NIH • Developed at the University of Washington, Seattle
[Includes: Fragile X Syndrome, Fragile X-Associated Tremor/Ataxia Syndrome (FXTAS), FMR1-Related Premature Ovarian Failure (POF)]
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Authors:
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Robert A Saul, MD, FACMG
Jack C Tarleton, PhD, FACMG |
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Initial Posting:
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Last Revision:
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Disease characteristics. FMR1-related disorders include fragile X syndrome, fragile X-associated tremor/ataxia syndrome (FXTAS), and FMR1-related premature ovarian failure (POF). Fragile X syndrome occurs in individuals with an FMR1 full mutation and is nearly always characterized by moderate mental retardation in affected males and mild mental retardation in affected females. Because FMR1 mutations are complex alterations involving nonclassic gene-disrupting alterations (trinucleotide repeat expansion) and abnormal gene methylation, affected individuals occasionally have an atypical presentation with an IQ above 70, the traditional demarcation denoting mental retardation. Males with an FMR1 full mutation accompanied by aberrant methylation may have a characteristic appearance (large head, long face, prominent forehead and chin, protruding ears), connective tissue findings (joint laxity), and large testes after puberty. Behavioral abnormalities, sometimes including autism spectrum disorder, are common. FXTAS occurs in males who have an FMR1 premutation and is characterized by late-onset, progressive cerebellar ataxia and intention tremor. FMR1-related POF (age at cessation of menses <40 years) occurs in approximately 20% of females who have an FMR1 premutation.
Diagnosis/testing. The diagnosis of FMR1-related disorders rests on the detection of an alteration in the FMR1 gene. More than 99% of individuals with fragile X syndrome have a loss-of-function mutation in the FMR1 gene caused by an increased number of CGG trinucleotide repeats (typically >200) accompanied by aberrant methylation of the FMR1 gene. Other mutations within FMR1 that cause fragile X syndrome include deletions, point mutations that disrupt RNA splicing, and a missense mutation. All individuals with FXTAS have FMR1 premutation trinucleotide repeats ranging from 59 to approximately 200. Females with FMR1-related POF have FMR1 trinucleotide repeats from high normal (35 repeats) into the premutation range. Both increased trinucleotide repeats and methylation changes in FMR1 can be detected by clinically available molecular genetic testing.
Management. Treatment of manifestations: Fragile X syndrome: early developmental intervention, special education (individual attention, small class size, and avoiding sudden change and excessive stimulation), and vocational training; individualized pharmacologic management of behavioral issues that significantly affect social interaction; routine treatment of medical problems. FXTAS: supportive care for gait disturbance and/or cognitive deficits. POF: reproductive endocrine evaluation for treatment and counseling for reproductive options. Agents/circumstances to avoid: folic acid in individuals with poorly controlled seizures.
Genetic counseling. All mothers of individuals with an FMR1 full mutation (expansion >200 CGG trinucleotide repeats) are carriers of an FMR1 gene expansion. They and their family members are at increased risk of having offspring with fragile X syndrome and FXTAS. Males with FXTAS will transmit their FMR1 premutation expansion to none of their sons and to all of their daughters, who will be premutation carriers. Carrier testing and prenatal testing are possible for pregnancies at increased risk if the diagnosis of an FMR1-related disorder has been confirmed in a family member.
Fragile X syndrome. A definite diagnosis of fragile X syndrome requires the presence of a loss-of-function mutation in FMR1, usually in a male with moderate mental retardation or a female with mild mental retardation.
Note: Because FMR1 mutations are complex alterations involving nonclassic gene-disrupting alterations (trinucleotide repeat expansion) and abnormal gene methylation, affected individuals occasionally have an atypical presentation with an IQ above 70, the traditional demarcation denoting mental retardation.
Affected individuals have normal growth and stature and no associated malformations.
Fragile X-associated tremor/ataxia syndrome (FXTAS)
A definite diagnosis of FXTAS requires the presence of a premutation in FMR1 and white matter lesions on MRI in the middle cerebellar peduncles and/or brain stem (the major neuroradiologic sign) with either intention tremor or gait ataxia (the two major clinical signs).
Other minor neuroradiologic criteria include MRI white matter lesions in the cerebral white matter or moderate to generalized atrophy.
Other minor clinical criteria include parkinsonism, moderate to severe working memory deficits, or executive cognitive function deficits.
A probable diagnosis of FXTAS requires either one major neuroradiologic sign and one minor clinical sign or two major clinical signs.
A possible diagnosis of FXTAS is based on one minor neuroradiologic sign and one major clinical sign [Grigsby et al 2005].
FMR1-related premature ovarian failure (POF). FMR1-related POF is defined as cessation of menses before age 40 years in a woman with an FMR1 premutation.
Chromosome analysis. Chromosome analysis using modified culture techniques to induce fragile sites is no longer used for diagnosis of fragile X syndrome because it is less sensitive and more costly than molecular genetic testing (see Molecular Genetic Testing).
Protein testing.
Although protein testing is not performed routinely in most clinical laboratory settings, a few laboratories provide assays measuring the production of the product of
FMR1, fragile X mental retardation 1 protein (FMRP) [Willemsen et al 1997]. See
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Situations in which FMRP testing may be useful include screening of mentally retarded populations and characterization of cellular production of FMRP in individuals having unusual phenotypes. Because severity of the fragile X syndrome phenotype appears to correlate with FMRP expression, assessment of FMRP production in some affected individuals has been proposed as a potential prognostic indicator of disease severity [Tassone et al 1999].
GeneReviews designates a molecular genetic test as clinically available only if the test is listed in the GeneTests Laboratory Directory by either a US CLIA-licensed laboratory or a non-US clinical laboratory. GeneTests does not verify laboratory-submitted information or warrant any aspect of a laboratory's licensure or performance. Clinicians must communicate directly with the laboratories to verify information. —ED.
Gene. FMR1 is the only gene known to be associated with FMR1-related disorders.
Allele sizes. FMR1 alleles are categorized according to the trinucleotide repeat number contained in exon 1 and the methylation status of the repeat region. However, the distinction between allele categories is not absolute and must be made by considering both family history and repeat instability. The boundary between intermediate and premutation categories listed below differs slightly from the American College of Medical Genetics (ACMG) guidelines for diagnostic and carrier testing [Sherman et al 2005]. The ACMG guidelines describe the intermediate range as 41-60 repeats and the premutation range as 61-200 repeats, but state that "the definitions of premutation and intermediate alleles are blurred." The demarcation used here is based on there being no reports of maternal alleles and fewer than 59 repeats transmitted from mother to offspring who have fragile X syndrome. In support of the categorization used here, recent data describing the FMR1 testing experience from a large commercial laboratory found no expansions of alleles containing fewer than 59 repeats during a single meiosis and suggested that the upper end of the intermediate or "gray zone" could be expanded to 58 repeats [Strom et al 2007].
Normal alleles: Approximately 5-40 repeats
Alleles of this size are stably transmitted without any increase or decrease in repeat number.
In stable normal alleles, the CGG region is interrupted by an AGG triplet after every nine or ten CGG repeats. These AGG triplets are believed to maintain repeat integrity by preventing DNA strand slippage during replication. (See Molecular Genetic Pathogenesis for a more complete discussion of pure repeats.)
Normal alleles with more than 35 CGG repeats are associated with increased risk of POF (see FMR1-related premature ovarian failure).
Intermediate alleles (also termed "gray zone"): No consensus exists regarding the precise size of intermediate alleles, but it may be broadly defined as 41-58 repeats. Historically, the largest repeat included in the intermediate range has been 54; however, the intermediate range may extend slightly higher as no transmission of alleles with 58 or fewer repeats is known to have resulted in an affected individual to date.
Note: (1) Because most clinical testing laboratories state that repeat measurements are plus or minus two to three repeats, it may be wise to consider reported test results with 55-58 repeats as potential premutations. (2) The risk for instability of alleles with 41-49 repeats when transmitted from mother to child is minimal. Any changes in repeat number are typically very small (±1 or 2 repeats). (3) The frequency and magnitude of repeat instability increases with alleles containing more than 50 repeats [Sherman 2005]. (4) Nolin et al (2003) reported two instances of maternal transmission of 59-repeat alleles producing offspring with full mutations; 59 is the smallest repeat known to expand to full mutation in a single transmission. (5) An important predictor of repeat instability in large intermediate alleles (>50 repeats) is the number of "pure" CGG repeats without interrupting AGG repeats. Increasingly longer pure repeats, especially those with more than 35 uninterrupted CGG repeats, are more likely to become unstable [Eichler et al 1994].
Premutation alleles: Approximately 59-200 repeats. Alleles of this size are not associated with mental retardation but do convey increased risk for FXTAS and POF. Because of potential repeat instability upon transmission of premutation alleles, women with alleles in this range are considered to be at risk of having children affected with fragile X syndrome.
Note: The upper limit of the premutation range is sometimes noted as approximately 230. Both numbers (200 and 230) are estimates derived from Southern blot analysis, in which repeat size can only be roughly estimated.
Full mutation alleles: More than 200 repeats, with several hundred to several thousand repeats being typical. Aberrant hypermethylation of the FMR1 promoter region typically occurs in alleles in which the number of CGG repeats exceeds approximately 200.
Clinical testing
Polymerase chain reaction (PCR) specific for the trinucleotide repeat region of FMR1 has high sensitivity for FMR1 repeats in the normal and lower premutation range (≤100 to 120 repeats) (see Table 3).
Note: (1) While the PCR assay alone yields accurate estimates of many FMR1 allele sizes, it may fail to detect alleles in the upper premutation range and full mutation alleles with a high repeat number. (2) When PCR of the FMR1 repeat segment reveals a normal or premutation allele in males, or two alleles of different size within the normal, intermediate, or premutation range in females, further testing may not be indicated. (3) PCR analysis of rare individuals who have cellular mosaicism for the FMR1 repeat may give a false negative result. In cellular mosaicism, PCR detects alleles in the normal, intermediate, or low premutation ranges but fails to detect alleles in the high premutation or full mutation range [Orrico et al 1998 , Schmucker & Seidel 1999]. Southern blot analysis detects these complex alterations in most cases.
Southern blot analysis detects all FMR1 alleles including normal, larger-sized premutations, and full mutations, allowing a low-resolution estimation of trinucleotide repeat number. PCR analysis can then be used to obtain a more precise repeat estimate of normal and smaller premutation alleles.
Methylation status. Southern blot analysis may be used to determine the methylation status of the FMR1 promoter. PCR techniques that determine methylation status ("methylation PCR") have been developed in several clinical laboratories and may offer a more rapid test turnaround time [Das et al 1997 , Weinhausel & Haas 2001].
Sequence analysis. More than 99% of FMR1 mutations detected in individuals with fragile X syndrome are repeat expansions. Although FMR1 would be expected to have a mutation rate similar to other genes, there is a surprising paucity of reports of individuals with point mutations or other small alterations in FNMR1 resulting in fragile X syndrome. To address the possibility that there may be a small subset of undetected individuals with fragile X syndrome who have rare intragenic mutations, a few clinical laboratories offer DNA sequence analysis of the FMR1 promoter region and the 17 coding exons and flanking intronic regions.
Note: (1) Small exonic deletions/duplications, insertions, and intragenic inversions can also be detected by sequence analysis. (2) Sequence analysis may fail to detect FMR1 deletions of an entire allele in females.
Deletion analysis. Deletions are typically detected in FMR1 as a secondary finding when the trinucleotide repeat region is being interrogated by Southern blot analysis. Therefore, it is likely that FMR1 deletions downstream of the repeat region, as well as other gene rearrangements, are underascertained. Nevertheless, both small deletions near the repeat region in exon 1 and large-scale deletions that completely remove FMR1 continue to be reported regularly in the literature.
FISH analysis. Fewer than 1% of individuals with fragile X syndrome have a partial or full deletion of the FMR1 gene (reviewed in Hammond et al 1997). Deletions not located in the repeat region of the gene may be missed on routine clinical testing for the trinucleotide repeat expansion; FISH may detect such deletions.
X-chromosome inactivation. Although not routinely performed, DNA testing for nonrandom X-chromosome inactivation may be useful in evaluating females who have a full mutation. Random X-chromosome inactivation in a female predicts a 50:50 ratio of cells inactivating the maternally derived X chromosome and cells inactivating the paternally derived X chromosome. However, population studies find a distribution of ratios. Typically, a greater-than 90:10 ratio indicates unusual skewing of the X-chromosome inactivation ratio. In females who have an FMR1 full mutation, nonrandom X-chromosome inactivation may result in more or less FMRP production with a resulting effect on the severity of the phenotype.
FMRP assessment by immunohistochemical staining of a lymphocyte smear may be used as an indicator of X-chromosome inactivation ratios. Cells in which the active X chromosome has a full FMR1 mutation should produce FMRP. In addition, FMRP assessment may be useful to determine protein production in males with methylation mosaicism, point mutations, or other mutation types that may not completely inactivate FMR1. An important caveat to X-chromosome inactivation studies is that such testing is typically performed using lymphocytes rather than brain tissues, where the mental and behavioral phenotype is manifested.
Research testing. The number and position of AGG repeats are known to be important in the overall stability of the CGG repeat sequence [Eichler et al 1994], but this analysis is currently available only in research settings.
Table 1
summarizes molecular genetic testing for this disorder.
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Test Method
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Mutations Detected
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Mutation
Detection Frequency by Test Method |
Test Availability
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PCR. CGG expansion in
FMR1 (allele sizes in the normal and lower premutation range)
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>99%
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Clinical
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Southern blot. CGG expansion in
FMR1 (all repeat ranges); methylation status
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FMR1 sequence variants
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<1%
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Large (partial or whole-gene)
FMR1 deletions
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<1%
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Deletion analysis
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<1%
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Research only
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Interpretation of test results
For issues to consider in interpretation of sequence analysis results, click here .
If the clinical phenotype is very consistent with fragile X syndrome and blood analysis is normal, testing of a second tissue type (e.g., skin fibroblasts) should be considered [MacKenzie et al 2006].
Establishing the diagnosis in a proband. Molecular genetic testing is appropriate for the following (see Figure 1):
Individuals of either sex with mental retardation, developmental delay, or autism, especially if they have (a) any physical or behavioral characteristics of fragile X syndrome, (b) a family history of fragile X syndrome, or (c) male or female relatives with undiagnosed mental retardation
Individuals who have a cytogenetic fragile X test result that is discordant with their phenotype, including (a) those who have a strong clinical indication (including risk of being a carrier) and who have had a negative or ambiguous cytogenetic test result and (b) those with an atypical phenotype who have had a positive cytogenetic test result
Note: Chromosome analysis using modified culture techniques to induce fragile sites is no longer used for diagnosis of fragile X syndrome because it is less sensitive and more costly than molecular genetic testing.
Males and females older than age 50 years who have progressive cerebellar ataxia and intention tremor with a positive family history of FMR1-related disorders in whom other common causes of ataxia have been excluded (see Ataxia Overview)
Women with unexplained POF [Corrigan et al 2005]
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Figure 1. Testing algorithm for FMR1-related disorders. The boxes identified with asterisks (*) identify individuals to be considered for FMR1 molecular testing. See Testing Strategy, Establishing the diagnosis in a proband for further discussion [Maddalena et al 2001 , Sherman et al 2005]. |
Clarification of the genetic status of women seeking reproductive counseling who have a family history of FMR1-related disorders requires prior confirmation of the presence of an expanded (or altered) FMR1 allele in the family or the presence of undiagnosed mental retardation.
Prenatal diagnosis for at-risk pregnancies requires prior confirmation of the presence of an expanded (or altered) FMR1 allele in the family.
Note: Results from chorionic villus sampling (CVS) testing must be interpreted with caution because often the methylation status of FMR1 is not yet established in chorionic villi at the time of sampling. CVS, while a standard technique for prenatal diagnosis, may lead to a situation in which follow-up amniocentesis is necessary to resolve an ambiguous result.
Preimplantation genetic diagnosis (PGD) for at-risk pregnancies requires prior confirmation of the presence of an expanded (or altered) FMR1 allele in the family.
No phenotypes other than fragile X syndrome, FXTAS, and POF are known to be associated with mutations in FMR1.
However, preliminary studies of the correlation of FMR1 allele size variations in the normal and premutation range suggest a possible relationship to mild cognitive impairment in females [Allen et al 2005] and males [Loat et al 2006]. Varied results [Ennis, Murray et al 2006] demonstrate the need for further research.
A male with complex FMR1 mosaicism (full mutation, premutation, and deletion) with only learning disability [Han et al 2006] also raises issues concerning gene and protein expression in FMR1-related phenotypes.
Males with full mutation alleles (fragile X syndrome). The phenotypic features of males with a full mutation and, hence, the fragile X syndrome, vary in relation to puberty (see Clinical Features in Males with Fragile X Syndrome).
Prepubertal males tend to have normal growth but large occipitofrontal head circumference (>50th percentile). Hypotonia, gastroesophageal reflux, and recurrent otitis media are problems in infancy that require medical attention [Hagerman & Hagerman 2002]. Other physical features not readily recognizable in the preschool-age child become more obvious with age. These involve the craniofacies (long face, prominent forehead, large ears, and prominent jaw) and genitalia (macro-orchidism), delayed attainment of motor milestones and speech, and abnormal temperament (hyperactivity, hand flapping, hand biting, temper tantrums, and occasionally autism).
Behaviors in postpubertal males with fragile X syndrome often include tactile defensiveness, poor eye contact, perseverative speech, problems in impulse control, and distractibility. The behaviors tend to become more obvious over time. The comorbid diagnosis of autism occurs in nearly 25% of affected individuals [Hatton et al 2006].
Note: Recent evidence suggests an increased risk of autism spectrum disorder and/or attention deficit disorder in premutation carriers as well [Farzin et al 2006].
Ophthalmologic (strabismus), orthopedic (joint laxity), cardiac (mitral valve prolapse), and cutaneous (excess softness and smoothness) abnormalities have also been noted. Except for the strabismus, these issues typically do not require significant intervention.
Periventricular heteropia and other neuroradiologic abnormalities [Moro et al 2006] are consistent with abnormal neuronal migration and development suggested by the metabotropic glutamate receptor (mGluR) theory of fragile X mental retardation (see Molecular Genetic Pathogenesis).
Clinical Features in Males with Fragile X Syndrome (adapted from Tarleton & Saul 1993)
Delayed developmental milestones (*)
* = usual age of attainment for affected boys
Prepubertal features
Postpubertal features
Other features
Females heterozygous for full mutation alleles (fragile X syndrome). The physical and behavioral features seen in males with fragile X syndrome have been reported in females heterozygous for the full mutation, but with lower frequency and milder involvement.
Fragile X-associated tremor/ataxia syndrome (FXTAS) is characterized by late-onset progressive cerebellar ataxia and intention tremor in persons who have an FMR1 premutation [Jacquemont et al 2004 , Jacquemont et al 2006]. Other neurologic findings include short-term memory loss, executive function deficits, cognitive decline, dementia, parkinsonism, peripheral neuropathy, lower-limb proximal muscle weakness, and autonomic dysfunction [Loesch et al 2005 , Bacalman et al 2006 , Grigsby et al 2006 , Louis et al 2006].
Both males and females with a premutation are at increased risk for FXTAS. The prevalence of FXTAS is estimated at 40% overall for males with premutations who are over age 50 years [Grigsby et al 2005]. Penetrance in males is age related (see Table 2).
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Age in Years
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Risk
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50-59
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17%
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60-69
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38%
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70-79
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47%
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≥80
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75%
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Although the precise risk for females has not yet been defined, it appears to be lower than that for males [Hagerman et al 2004 , Biancalana et al 2005].
A retrospective longitudinal review of 55 males with premutations provides early natural history information of FXTAS [Leehey et al 2007]. The first sign to appear is usually tremor at approximately age 60 years. Ataxia tends to develop two years later, leading to increased tendency to fall and subsequent dependence on walking aids. Life expectancy after onset of symptoms ranged from five to 25 years.
Neuroradiologic signs (decreased cerebellar volume, increased ventricular volume, increased white matter hyperdensity) appear to correlate with premutation CGG repeat length [Cohen et al 2006].
FMR1-related premature ovarian failure (POF), defined as cessation of menses before age 40 years, has been observed in carriers of premutation alleles [Murray et al 1999 , Uzielli et al 1999 , Hundscheid et al 2000 , Bussani et al 2004 , Machado-Ferreira et al 2004]. Ovarian failure has occurred as early as age 11 years. The diagnosis of POF does not eliminate the possibility of subsequent conception. A premutation carrier woman had a child with fragile X syndrome after her diagnosis with POF [Corrigan et al 2005 , Nelson et al 2005]. It is estimated that 5%-10% of women may conceive after the diagnosis of POF is established [Nelson et al 2005].
An increased risk for POF and FMR1 alleles containing trinucleotide repeats in the high normal (≥35 repeats) and intermediate ranges has been reported [Bretherick et al 2005 , Bodega et al 2006]. Currently, no consensus exists for estimating an absolute risk for POF when a woman has high normal or intermediate repeat alleles. Sherman (2005) concluded that the risk for POF was 21% (estimates ranged from 15% to 27% in various studies) in premutation carriers, compared to a 1% background risk. In this review an odds ratio of 2.5 was estimated for intermediate repeat sizes of 41-58 [Wittenberger et al 2007]. (See Genotype-Phenotype Correlations, Premutation for additional risk estimates.)
Sullivan et al (2005) suggest that variation in the age at menopause in the general population might be related to FMR1 CGG repeat size of less than 80, a finding further supported by data from Ennis, Ward et al (2006). A significant increase of alleles in the 35 to 54 range was found in women with POF [Bretherick et al 2005]. In all three studies, larger premutations (>80 CGG repeats) carried lower risk for POF.
Women with full mutation alleles are not at increased risk for POF.
The phenotype of males with an FMR1 mutation depends almost entirely on the nature of the mutation; the phenotype of females with an FMR1 mutation depends on both the nature of the FMR1 mutation and random X-chromosome inactivation (see Table 3) .
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MR = mental retardation
1. Both males and females with premutations and manifestations of some symptoms of fragile X syndrome have been reported [Riddle et al 1998]. 2. FMR1 mutations are complex alterations involving nonclassic gene-disrupting alterations (trinucleotide repeat expansion) and abnormal gene methylation. This complexity at the gene level affects production of the FMR1 protein and may result in an atypical presentation in which affected individuals occasionally have an IQ above 70, the traditional demarcation denoting mental retardation. |
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Premutation. Males and females who have a fragile X premutation have normal intellect and appearance. As noted in Table 3 , footnote 1, a few individuals with a premutation have subtle intellectual or behavioral symptoms including learning difficulties or social anxiety. The difficulties are usually not socially debilitating, and these individuals may still marry and have children.
It is estimated that 21% of premutation carriers will have POF [Sherman 2005]. The odds ratios for POF in premutation carrier females increases with increasing repeat sizes [Sherman 2005] (see Table 4). Although the numbers vary slightly, other studies confirm that these increased risks tend to plateau above 80-100 repeats [Bodega et al 2006 ; Ennis, Ward et al 2006].
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Full mutation. Males who have a full fragile X mutation generally have moderate to severe mental impairment and may or may not have a distinctive appearance.
Approximately 50% of females who have a full fragile X mutation are mentally retarded; however, they are usually less severely affected than males with a full mutation. Conversely, approximately 50% of females who are heterozygous for the full mutation are intellectually normal. The variability among females is believed to result from the ratio in the brain of active X