Fragile X Syndrome

  • Preferred test to diagnose FXS, FXTAS, and for carrier screening in individuals with a positive family history
  • Test Description
    • Triplet repeat-primed polymerase chain reaction (PCR) followed by size analysis using capillary electrophoresis to determine FMR1 CGG repeat length
    • Methylation-specific PCR is performed for >100 CGG repeats to distinguish between premutation and full mutation alleles
  • Prenatal test for women with fragile X premutations or full mutations.
  • Contraindications
    • Prenatal testing for women with normal or intermediate allele sizes
    • Testing of chorionic villus (CVS) samples, because methylation patterns are not fully established in the first trimester of pregnancy
Related Tests
  • Carrier testing for cystic fibrosis (CF), fragile X syndrome (FXS), and spinal muscular atrophy (SMA) in expectant couples or those planning a pregnancy
  • Not recommended for diagnostic testing in patients with symptoms of CF, FXS, or SMA

Disease Overview

Fragile X syndrome (FXS), the most common heritable form of intellectual disability (ID) and autism, is caused by full FMR1 gene mutations. Individuals with an FMR1 gene premutation may develop fragile X-associated tremor/ataxia syndrome (FXTAS), FX-associated neuropsychiatric disorder, or primary ovarian insufficiency (POI).

Diagnostic testing should be offered to individuals with unexplained intellectual disability (ID), developmental delay, autism spectrum disorder, late-onset cerebellar ataxia and intention tremor, POI, or infertility associated with elevated follicle-stimulating hormone (FSH) levels. 

Screening for FXS should be offered to women with a positive family history of FXS, FXTAS, unexplained ID, or autism. Fetal testing should be offered to women who carry a fragile X premutation or full mutation.

Symptoms of FXS

  • Moderate ID (median IQ of 40-45 but ranges from <10 to normal range)
  • Autism in 50-70%
  • Perseverative speech
  • Behavioral issues: hand flapping/biting, attention deficit hyperactivity disorder (ADHD), social anxiety, poor eye contact, tactile defensiveness, aggressiveness, and irritability
  • Large ears, long face, large jaw, prominent forehead, and large testes

Symptoms of FXTAS

  • Cerebellar gait ataxia
  • Intention tremor
  • Parkinsonism; muscle rigidity, unbalanced shuffling gait, slowed movement and speech
  • Moderate to severe short-term memory loss
  • Executive function deficit
  • Incontinence/ impotence

Prevalence

FXS:

  • 1/4,000 males
  • 1/8,000 females

Premutation allele in U.S.:

  • 1/1,000 males
  • 1/350 females

Genetics

Gene

FMR1

Inheritance

X-linked

Structure/Function

FMR1 codes for fragile X mental retardation protein (FMRP), an RNA-binding protein expressed in many tissues

Mutations

Caused by FMR1 variants

  • 99% caused by expansion of the FMR1 gene CGG repeat​
  • Less than 1% caused by FMR1 sequence variants or partial/full FMR1 gene deletions
  • Risk for CGG repeat expansion is dependent on sex of transmitting parent and size of repeat

CGG repeat sizes:

  •  Normal alleles: 5-44 CGG repeats (unmethylated)
    • Stably transmitted
  • Intermediate alleles: 45-54 CGG repeats (unmethylated)
    • Unstable but will not expand to a full mutation in one generation
  • Premutations: 55 to ~200 CGG repeats (unmethylated)
    • Females:
      • At risk for having offspring with FXS-transmission of CGG repeats to offspring is unstable
        • Premutations of <56 repeats have not expanded to full mutation in a single generation  
        • Stability of alleles <90 CGG repeats is influenced by the number of AGG interspersions within the CGG repeat sequence
        • Premutations >90 repeats nearly always expand to full mutation in offspring
      • 21% risk for POI (before age 40)
      • At risk for fragile X-associated neuropsychiatric disorders
      • 17% risk for FXTAS in women>50 years of age
    • Males:
      • Transmission of CGG repeats is stable
      • All of their daughters and none of their sons will inherit the premutation
      • 47% risk for FXTAS in men >50 years of age
  • Full mutations: typically >200 CGG repeats (methylated)
    • Males are affected with FXS
    • 50% of females have moderate ID  
    • Disease symptom severity cannot be predicted based on:
      • Size of CGG repeat
      • Degree of methylation
      • Pattern of X-inactivation (in females)

Test Interpretation

Sensitivity/Specificity

  • Clinical sensitivity/specificity: 99% 
  • Analytical sensitivity/specificity: 99%  

Results

Result Number of CGG Repeats Clinical Significance

Full mutation

>~200 (methylated)

Male:

FXS with ID

Female:

Variable expression of FXS; 50% have ID

Premutation

55 to ~200 (unmethylated)

Male:

47% risk for FXTAS in men >50 years of age

Female:

At risk for POI

At risk for FX-associated neuropsychiatric disorders

17% risk for FXTAS in women >50 years of age

Indeterminate

45 to ~54

Offspring at risk for inheriting premutation

Negative

5 to ~44

Normal; not affected with nor a carrier of FXS

Limitations

  • Estimated CGG repeat number is not provided for full mutations (alleles with >200 repeats)
  • Sizing precision of CGG premutation alleles is within two-three CGG repeats
  • Methylation patterns are not fully established in the first trimester of pregnancy; thus, CVS is not recommended for prenatal diagnosis. A small, full mutation may be distinguished from a large premutation in amniocytes
  • Rare FMR1 variants unrelated to trinucleotide expansion will not be detected
  • Diagnostic errors can occur due to rare sequence variations
  • AGG trinucleotide interruptions within the FMR1 CGG repeat tract are not assessed

References

  1. GeneReviews - FMR1 Disorders

    Hunter JE, Berry-Kravis E, Hipp H, et al. FMR1 disorders. In: Adam MP, Ardinger HH, Pagon RA, et al, editors. GeneReviews, University of Washington; 1993-2020. [Last Update: Nov 2019; Accessed: Feb 2020]

    Online