DMD/BMD - Part 3 Flashcards
Why is genetic testing for BMD/DMD important?
- Genetic testing for DMD/BMD is important because it confirms a clinical diagnosis.
- It is important to know specific mutations for some drug trials.
- Knowing the mutations also enables carrier testing and confirmatory testing in other family members.
- It should be noted that 2 dystrophin mutations have been known to occur in the same family so confirmation testing in all affected individuals in the family should ideally be performed.
- Carrier testing allows recurrence risk to be calculated and prenatal diagnosis to be offered.
- Diagnostic referrals are often received from neurologists or clinical genetics.
- Carrier testing and prenatal diagnosis should be referred through clinical genetics.
- Initial test is MLPA using both kits.
What software is used for DMD/BMD MLPA analysis?
- Analysis can be performed using different software. May use Manchester T-test spreadsheet, Coffalyser or GeneMarker.
If a normal result is shown by MLPA does this exclude a diagnosis of DMD?
- If a normal result is show by MLPA this does not exclude a diagnosis as deletions/duplications only account for approximately 72% of dystrophin mutations.
- If a clinical diagnosis is confirmed by CK levels/muscle biopsy then can offer point mutation analysis/linkage studies.
How would we proceed if we obtained a positive result on dystrophin MLPA analysis?
- A positive result is when a deletion or duplication is detected by MLPA.
- If a single exon deletion is detected then we need to confirm by another method prior to reporting. Often single exon duplications are confirmed too as it is possible to get spurious results due to DNA quality.
- If a deletion or duplication is detected then we need to check whether it is in frame or out of frame. The Leiden Muscular Dystrophy database can be used to check this. It should be noted on the report that this is only a prediction because it is based on the DNA level rather than the RNA level.
- Once you have checked the reading frame you can report a deletion or duplication confirming a diagnosis of DMD/BMD.
- If the first or last exon is deleted then the deletion could extend into neighbouring genes and alter the phenotype.
- Carrier testing and prenatal diagnosis can be offered to the family.
- If the individual has children all daughters will be obligate carriers.
Describe how diagnostic referrals for DMD/BMD for females should be reported.
- Diagnostic referrals are sometimes received for females.
- If a deletion or a duplication is found in a female with symptoms the result can be reported as being consistent with the clinical phenotype of a manifesting DMD/BMD carrier.
- Further testing should be offered in these cases.
- Cytogenetic analysis is advisable to detect whether a translocation or Turner syndrome is present.
- X-inactivation studies can also be offered if appropriate.
- All male and female positive patients should be referred to the clinical genetics department for counselling and cascade screening.
Describe carrier testing for dystrophinopathies.
Do we always know the familial mutations for carrier testing?
What do we do if we don’t have this information?
What do we do if a mutation is found? What do we do if no mutation is found?
- For carrier tests when the familial mutation is known the MLPA set covering the familial deletion/duplication or sequence analysis will be carried out, ideally including the familial positive control to ensure the mutation can be picked up.
- Can get carrier test referrals where the familial mutation is not identified yet. This is often because the affected individual is deceased. In these cases testing should be done using both MLPA kits and should be performed on the woman in the family with the highest prior risk of being a carrier. This will normally be the mother of the index case.
- If the familial mutation has not been identified after testing linkage analysis can be performed if appropriate family members are available.
- In a carrier test if a mutation is found we report that the individual is a carrier of a dystrophinopathy. Prenatal diagnosis can then be offered to male preganacies and carrier testing to male relatives when referred through clinical genetics.
- If a mutation is not found and the familial mutation is known then the individual is unlikely to be a carrier of a dystrophinopathy except if they are the mother of an affected child where there is still the risk of germline mosaicism.
- If the familial mutation is not known and the test is negative the carrier risk is reduced and there is still a residual risk. This depends on the detection rate of the test performed. Baysean risk calculation can be performed to calculate the residual carrier risk.
How would we progress if we identified a DMD/BMD mutation on carrier testing?
- In a carrier test if a mutation is found we report that the individual is a carrier of a dystrophinopathy.
- Prenatal diagnosis can then be offered to male pregnancies and carrier testing to male relatives when referred through clinical genetics.
How would we progress if we identified no DMD/BMD mutation on carrier testing?
- If a mutation is not found and the familial mutation is known then the individual is unlikely to be a carrier of a dystrophinopathy except if they are the mother of an affected child where there is still the risk of germline mosaicism.
- If the familial mutation is not known and the test is negative the carrier risk is reduced and there is still a residual risk. This depends on the detection rate of the test performed. Baysean risk calculation can be performed to calculate the residual carrier risk.
Outline Bayes analysis for DMD/BMD.
1). Need to start off with the prior risk based on the individuals position within the pedigree.
4u (where u = mutation rate) is the risk that a female is a carrier of DMD taking into account no other information.
18u is the prior risk of BMD due to biological fitness.
The prior risk is 2/3 for the mother of a sporadic DMD/BMD case.
The prior risk is 1/3 for the grandmother of a sporadic case.
2) . Need to modify the prior risk by the conditional risk. This includes factors such as whether there are affected or unaffected sons, the results of genetic tests and CK levels.
3) . Joint risks can then be calculated and a posterior risk can be determined.
What is the risk that a female is a carrier of DMD taking into account no other information.
4u (where u = mutation rate) is the risk that a female is a carrier of DMD taking into account no other information.
What is the prior risk of BMD due to biological fitness?
18u is the prior risk of BMD due to biological fitness.
What is the prior risk of DMD for the mother of a sporadic case?
The prior risk is 2/3 for the mother of a sporadic DMD/BMD case.
What is the prior risk of DMD for the grandmother of a sporadic case?
The prior risk is 1/3 for the grandmother of a sporadic case.
Describe prenatal diagnosis for DMD/BMD.
- For prenatal diagnosis usually only male foetuses are tested. This is because it is not possible to predict the phenotype of a female and there are ethical issues concerning identifying a carrier female before the age of consent.
- To find out the sex of the foetus free fetal DNA sexing is often performed prior to prenatal test. Due to the possibility of a false result sexing is often confirmed via QF-PCR or FISH when the prenatal sample arrives.
- MCC in the prenatal sample can interfere with the results of prenatal analysis. Therefore it is important to exclude MCC in any prenatal sample. This testing is performed alongside the prenatal and usually involves the use of a number of microsatellite markers to ensure both maternal alleles are not present in the prenatal sample.
- Prenatals may be performed when the mother is not known to be a carrier due to the risk of germline mosaicism.
- If mutation is unknown prenatal diagnosis may be performed using linkage analysis.
- Preimplantation genetic diagnosis is available for DMD.
If the mutation is not known how may prenatal diagnosis be carried out?
If mutation is unknown prenatal diagnosis may be performed using linkage analysis.
