Fragile X syndrome; Premature Ovarian Insufficiency; Fragile X Associated Tremor/Ataxia Syndrome; Martin Bell syndrome

FMR1-related disorders include fragile X syndrome, fragile X-associated tremor/ataxia syndrome (FXTAS), and FMR1-related premature ovarian insufficiency (POI). Fragile X syndrome is characterized by moderate intellectual disability in males and mild intellectual disability in affected females.  Males may also display a characteristic appearance, macroorchidism after puberty and behavioral abnormalities.  FXTAS may occur in males and, rarely, in females who have an FMR1 premutation, and is characterized by late-onset, progressive cerebellar ataxia and intention tremor.  FMR1-related POI occurs in approximately 20% of females who have an FRM1 premutation.

The FMR1-related disorders are caused by mutations in the FMR1 gene on the X-chromosome, the most common mutation being expansion of the CGG repeat in the 5′ untranslated region of exon 1. Repeat alleles in the FMR1 gene are classified in our lab as:

• Normal: ~5 to ~54 repeats
• Premutation: ~55 to ~200 repeats and unmethylated
• Full mutation: >200 repeats and methylated

More than 99% of individuals with fragile X syndrome have a loss-of-function mutation in the FMR1 gene caused by the expansion of CGG trinucleotide repeats into the full mutation range, which results in aberrant methylation of the FMR1 gene.

Other mutations in FMR1 that cause fragile X syndrome include deletions and point mutations that disrupt RNA splicing, and missense mutations. All individuals with FXTAS or FMR1-related POI have an FMR1 premutation.

1. Confirmation of diagnosis:
   1. Fragile X Syndrome: Individuals of either sex with global developmental delay (GDD) or intellectual disability (ID) of unknown etiology , or autism spectrum disorders (ASD).  Testing females with learning disabilites may also be considered.
   2. FXTAS: Patients over 50 years of age who have progressive cerebellar ataxia and intention tremor in whom other common causes of ataxia have been excluded.
   3. Premature Ovarian Insufficiency: Women with unexplained premature ovarian insufficiency or reproductive or fertility problems associated with an elevated follicle stimulating hormone (FSH) level in the postmenopausal range before the age of 40.
2. Carrier testing. 

   NB: Carriers have the potential for health problems (FXTAS or FMR1-related POI) in addition to the ability to transmit disease to offspring, therefore this testing in an asymptomatic individual is presymptomatic testing.

   1. Adults with a family history of fragile X syndrome, fragile X tremor/ataxia syndrome, or premature ovarian failure (in more than one family member) if the pedigree structure is consistent with X-linked inheritance and the individual is at risk of inheriting the mutated gene. Referral to a medical geneticist for counselling and assessment should be considered in these cases. 
   2. Adults who have at least one male relative with autism or mental retardation/developmental delay of an unknown etiology within a three-generation pedigree, if the pedigree structure is consistent with X-linked inheritance and the individual is at risk of inheriting the mutated gene. 
3. Prenatal testing (prenatal diagnosis requests are not normally accepted from physicians other than Medical Geneticists):
   1. Pregnancies of females known to have an FMR1 mutation.

Population-based carrier screening (i.e., screening in the absence of any other indication) is not covered by Health Insurance BC (BC MSP). Please contact MGL to discuss.

PCR amplification is performed across the CGG repeat region of the FMR1 gene to determine the repeat size.  In some cases, triplet-primed (tp) PCR (Amplidex PCR/CE FMR1 Reagents, Asuragen, Inc) is performed to assess for the presence of expanded alleles. This assay does not assess methylation status; however, in most cases the repeat is sized well into the full mutation range and, thus, hypermethylation can be assumed.  In rare cases, a repeat collection and testing by Southern blot analysis will be recommended.

 For more information, see FAQ

 Please note: MGL reports repeat sizes only when relevant for risk estimate counselling (i.e. premutation range from 55 – ~120 repeats); otherwise, repeats are categorized as normal, premutation, and full mutation only.

Pregnancy-related/Prenatal

If pregnancy management will be altered, 3 weeks; otherwise, routine TAT.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays).  

Prenatal Specimens
Prenatal testing REQUIRES LABORATORY CONSULTATION PRIOR TO THE PROCEDURE and can only be ordered by a Medical Geneticist. Contact the laboratory at 604-875-2852 and choose the appropriate option for the Molecular Geneticist on service.
Chorionic Villi: 20 mg.
Direct Amniotic fluid: 25 mL collected in two separate tubes of equal volume.
Cultured Amniocytes: Two (2) 100% confluent T-25 flasks.
DNA extracted from prenatal specimens: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth. Ship samples by overnight courier with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays) as follows:

• Villi – on wet ice or in media at room temperature
• Amniocytes, Amniotic fluid, DNA – at room temperature

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used.  This method will not detect all mutations (e.g., point mutations in the coding region of the gene, large genomic deletions, promoter mutations, regulatory element mutations). 

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member.  Ideally, this information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In certain scenarios of repeat size mosaicism, false negative results may occur.  If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

Rare single nucleotide variants or polymorphisms could lead to false-negative results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype.  Consult the on-service Molecular Geneticist for approach to testing in such individuals. 

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing.  However, if there is no clinical urgency, the cautious approach is to wait one week post-packed red cell transfusion before collecting a sample for genetic testing.  Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.