DNA testing for Down syndrome

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DNA testing for Down syndrome

• Dr Nerine Gregersen
• Division Human Genetics
• NHLS and Wits University

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DOWN SYNDROME

• Commonest genetic cause of intellectual
  disability worldwide
• All ethnic groups
• Increased risk with advancing maternal age
• Age 25 1/1350
• Age 35 1/400
• Age 40 1/100
• Age 45 1/30

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Down syndrome

• Birth prevalence
• 1/700 births worldwide
• 1/500 in RSA
• Cause
• Non-disjunction / trisomy 95
• Translocation 4
• About 50 de novo
• Mosaicism 1

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Photo used with parental consent
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Down syndrome clinical features

• Craniofacial
• Brachycephaly, upslanted eyes, epicanthic folds,
  flat facial profile, flat nasal bridge,
  (protruding tongue), (dysplastic ears)
• Limbs
• Brachydactyly, single palmar crease,
  clinodactyly, sandle-gap
• Cardiac (40)
• ASD, VSD, AVSD, PDA
• CNS
• Hypotonia, developmental delay, MR
• Other
• GIT abn (15), short stature

Features common in normal black neonates Features
useful for diagnosis in black patients
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Down syndromecomplications

• Atlanto-axial instability
• Transient neonatal leukaemia
• ALL (20 X greater risk)
• Hypothyroidism (antibodies) in 30
• Alzheimer disease
• 8 by 49 yrs
• 75 by 60yrs

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TESTING
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100 base pairs
105
109
DNA sequencing
FISH Chromosome banding
CGH and MLPA
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CHROMOSOMES
DNA

• Live, dividing cells Any tissue
• Screen all chromosomes Test specific regions
• TAT 2 4 weeks TAT 72 hours
• Labour intensive Can be automated

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Short tandem repeats (STR)

• STR microsatellite markers
• 10 000s across genome
• Stretches of DNA of units of 2-4 nucleotides
• TGTGTG
• CAACAA.CAA
• Different alleles exist for each STR in popn.
• Each allele differs in repeat length

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(CC)12
(CC)7
(CC)11
(CC)8
(AAC)18
(AAC)26
(AAC)10
(AAC)15
(CC)8
(CC)11
(CC)12
(CC)7
(AAC)10
(AAC)15
(AAC)26
(AAC)18
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Quantitative fluorescent PCR aneuploidy screen
(QF-PCR)

• Principle
• Test STR markers on chromosomes
• 4-5 on autosomes of choice (13, 18, 21)
• Fewer on X and Y
• Used to detect numerical chromosome abnormalities
• Can test blood, amniotic fluid, CVS, post-mortem
  tissue etc.

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QF-PCR

• Advantages
• Rapid result (48 72 hours)
• 99 accuracy
• No live cells required
• Disadvantages
• Wont detect mosaicism (lt30)
• Wont detect other chromosome abnormalities
• Mechanism of aneuploidy remains unknown
• Blood-stained amnio may make testing impossible
• Risk of maternal contamination if CVS not
  carefully prepared/dissected

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Past
Pre-and postnatal testing for DS by karyotyping
Postnatal testing for DS by karyotyping
Prenatal testing for DS in AMA women by QF-PCR
January 2007 May 2008 563 requests for DS
testing
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Past
Pre-and postnatal testing for DS by karyotyping
Postnatal testing for DS by karyotyping
Prenatal testing for DS in AMA women by QF-PCR
January 2007 May 2008 563 requests for DS
testing
307 confirmed DS (54.5) 79 confirmed DS on
specimens not requested as DS 386 confirmed DS
185 not DS (33)
67 no result
4 other chromosome abnormalities
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Past
Pre-and postnatal testing for DS by karyotyping
Postnatal testing for DS by karyotyping
Prenatal testing for DS in AMA women by QF-PCR
January 2007 May 2008 563 requests for DS
testing
307 confirmed DS (54.5) 79 confirmed DS on
specimens not requested as DS 386 confirmed DS
185 not DS (33)
67 no result
4 other chromosome abnormalities
366 trisomy 95
6 mosaic 1.5
14 translocation 3.5
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present
Indication for DS testing
Prenatal testing for DS in AMA women by QF-PCR
Postnatal testing for DS by QF-PCR
Confirmed DS
2 RR relates to inherited translocation
98 RR relates to trisomy
Offer parental karyotype
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present
Indication for DS testing
Prenatal testing for DS in AMA women by QF-PCR
Postnatal testing for DS by QF-PCR
Negative QF-PCR
Confirmed DS
Dysmorphic features Unknown diagnosis
2 RR relates to inherited translocation
Strongly suspect DS ?mosaic
98 RR relates to trisomy
Request karyotype
Genetic counselling
Refer to Genetics
Offer prenatal testing and/or parental karyotype
if recurrence risk perceived as high
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Baby with multiple congenital abnormalities
Alive
Stillborn
Macerated
Skin snip (saline)
QF-PCR T13, 18, 21, X, Y
Refer to Genetics
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Baby with multiple congenital abnormalities
Alive
Stillborn
Fresh SB
Macerated
Skin snip (saline)

• Cardiac blood (heparin)
• Skin snip (saline)

QF-PCR T13, 18, 21, X, Y
Karyotype
Refer to Genetics
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Baby with multiple congenital abnormalities
Alive
Stillborn
Fresh SB
Macerated
MCA - unknown
Suspect trisomy 13 or 18
Skin snip (saline)

• Cardiac blood (heparin)
• Skin snip (saline)

Blood (heparin)
Blood (EDTA)
QF-PCR T13, 18, 21, X, Y
Karyotype
Karyotype
QF-PCR
Negative
Positive
Refer to Genetics
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Queries and Referrals

• Division Human Genetics
• 011 489 9223 / 4

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Acknowledgments

• Prof Arnold Christianson
• Dr Tony Lane
• Ms Julie Lampret
• Ms Brenda Kruger