Prenatal Screening for Down
Syndrome: Past, Present and
Emerging Practices
Short Presentation on Emerging Concept
(SPEC)
Down
Syndrome
Euploid
Euploid
Down
Syndrome
15
20
25
30
35
40
45
Maternal age (completed years)
50
1.30
1.35
1.40
1.45
Chromosome 21%
1.50
1.55
Down syndrome screening
• The first test was the question ‘How old are you?’
• If answered “35 or older”, the woman was offered
invasive testing (amniocentesis or CVS) and
diagnostic testing (karyotype).
Down
Syndrome
Euploid
‘Screen Positives’ located to the
right of the red line at 35 years
Detection rate 50%
False positive rate 15%
Figure Source: Canick JA, Palomaki GE.
J Med Screen. 2012 Jun;19(2):57-9.
15
20
25
30
35
40
45
Maternal age (completed years)
50
Down syndrome screening
• Maternal age was inexpensive, reliable and
available early in pregnancy, but had relatively low
detection and high false positives
• Today, 2nd trimester ‘quadruple’ testing is common
Euploid
‘Screen Positives’ located to the
left of the red line at risk of 1:270
Detection rate 80%
False positive rate 5%
Down
Syndrome
10 - 3 10 - 2 10 - 1 10 0 10 1 10 2 10 3 10 4 10 5 10 6 10 7
Figure Source: Canick JA, Palomaki GE.
J Med Screen. 2012 Jun;19(2):57-9.
Down's syndrome risk (1:n)
Down syndrome screening
• 1st trimester ‘combined’ testing has similar
performance to ‘quadruple testing’
• ‘combined’ + ‘quadruple’ = an ‘integrated’ test
‘Screen Positives’ located to the
left of the red line at risk of 1:100
Detection rate 90%
False positive rate 2%
Euploid
Down
Syndrome
10 - 3 10 - 2 10 - 1 10 0 10 1 10 2 10 3 10 4 10 5 10 6 10 7
Figure Source: Canick JA, Palomaki GE.
J Med Screen. 2012 Jun;19(2):57-9.
Down's syndrome risk (1:n)
Prenatal screening in the US in 2012
Type of test
Labs
Median N
1st trimester
34
3,000
565,692 (19)
2nd trimester
122
2,538
1,770,024 (60)
Integrated
30
4,176
583,416 (21)
All
123
3,660
2,963,592 (100)
Palomaki GE et al.,
Archives Path Lab Med 2013
Number (%)
Represents about 70%
of all US pregnancies
Current status
• Combinations of maternal age and multiple markers
– Serum markers (AFP, uE3, hCG, PAPP-A, inhibin-A)
– Ultrasound markers (nuchal translucency)
• But, these tests
–
–
–
–
are complex
still miss 10-20% of Down syndrome cases
lead to offering diagnostic tests to 2-6% of women
can only identify
• Down syndrome
• and to some extent, trisomy 18 and trisomy 13
• Would like even better performance for a wider
range of prenatal disorders
Circulating cell free (ccf) DNA
• First reported in 1997 by Dr. Dennis Lo, Chinese
University of Hong Kong (Lancet, 350:485)
• Used Y chromosome probes in maternal plasma to
identify male fetuses
• Both maternal and fetal (actually placental) DNA are
found in maternal circulation
• DNA in small fragments (150 to 200 bp)
• Reliably represents the entire genome of the mother
and fetus
• Fetal ccfDNA quickly cleared after birth
• Ratio of fetal to total ccf DNA is 10% (range <4% to 40%)
Current methodologies
• The current commercial laboratory-developed
tests (LDT) in the US can be divided by (not FDA
cleared/approved)
– Sequencing methodology: any fragment sequenced
(Shotgun) versus selectively amplified sequences
(Targeted)
– Interpretation methodology: comparing observed
percentage of aligned fragments from chromosome
of interest to expected (Counting) versus modeling
observed SNP genotype to specific models
(Genotyping)
External clinical validation studies
All
Study
False Positive
Rate (%)
Palomaki 2011
3/1,471 (0.2)
Ashoor 2012
Trisomy 21
No-calls
Down Syndrome
Detection Rate (%)
No-call
13/1,697 (0.8)
209/212 (98.6)
0
0/ 300 (0 )
1/ 400 (0.7)
50/ 50 (100)
0
Bianchi 2012
0/ 311 (0 )
23/ 532 (4.3)
89/ 89 (100)
1
Norton 2012
1/2,887 (0.1) 148/3,228 (4.6)
81/ 81 (100)
3
Nicolaides 2013
0/ 204 (0 )
25/ 25 (100)
2
All
4/4,173 (0.1)
454/457 (99.3)
6
13/ 242 (5.4)
Technology is advancing and ‘real world’ experience is being gained.
The performance of current commercial testing may differ
Down syndrome screening
• ccf DNA testing of maternal plasma
• Tests involve next generation sequencing (NGS)
‘Screen Positives’ located to the
left of the red line at risk of 1:100
Detection rate 99%
False positive rate 0.2%
1.30
Figure Source: Canick JA, Palomaki GE.
J Med Screen. 2012 Jun;19(2):57-9.
Euploid
Down
Syndrome
1.35
1.40
1.45
Chromosome 21%
1.50
1.55
Practice guidelines
• Sequencing of cell free DNA is considered sensitive and
specific for the common trisomies, such that
• Multiple professional organizations have written practice
guidelines relevant to ccfDNA plasma testing for
common trisomies
American Congress of Obstetricians and Gynecologists (ACOG)
American College of Medical Genetics and Genomics (ACMG)
International Society of Prenatal Diagnosis (ISPD)
National Society of Genetic Counselors (NSGC)
Society of Obstetricians and Gynecologists of Canada (SOGC)
Practice guidelines
• Testing for chromosomes 13, 18, and 21 should be
offered to ‘high risk’ pregnancies
• Testing should not be offered to the general pregnancy
population (‘low risk’) until more information is available
• Positive results followed up by offer of invasive testing
– ccfDNA currently is still is considered a screening test
• Patient and provider education is important
• Insufficient data for twins
“High Risk” Pregnancies
• Maternal age 35 years or older at delivery
• Fetal ultrasonographic findings indicating an increased
risk of aneuploidy
• History of a prior pregnancy with a trisomy
• Positive test result for aneuploidy, including first
trimester, sequential, or integrated screen, or a
quadruple screen
• Parental balanced robertsonian translocation with
increased risk of fetal trisomy 13 or trisomy 21
American College of Obstetricians and Gynecologists Committee on Genetics (2012). Committee Opinion No. 545: Noninvasive
prenatal testing for fetal aneuploidy. Obstet Gynecol, 120(6): 1532-1534.
Limitations
• Data is currently only sufficient to warrant use for
aneuploidy of chromosomes 13, 18, 21, and sex
chromosome aneuploidies
• Patients at risk for other genetic abnormalities, including
single gene disorders, should be counseled to utilize
other testing modalities
• Does not replace α-fetoprotein testing for neural tube
defects
• Testing turn-around-time should be considered when
testing later in pregnancy
ccfDNA testing in ‘High risk’ women
6,000
‘High risk’
(1:19)
300
DS
284
pos
3
neg
11
pos
3
fail
5,700
Euploid
5,632
neg
57
fail
295 (4.9%)
60 (1.0%)
5,635 (93.9%)
Offer Dx testing
26:1 (284:11)
Offer Dx testing
1:19 (3:57)
Routine care
1:1,900 (3:5,632)
Figure Source: Palomaki, GE.
Commercial LDTs: further disorders
• Trisomy 18 and trisomy 13
– All commercial LDTs provide an interpretation
– > 95% detection
• Sex chromosome aneuploidies
– Most commercial LDTs provide
– 45X, 47XXY, 47XXX, 47,XYY and fetal sex
– >90% detection
• Twin pregnancies
– Two commercial LDT provide
– Limited data
• Deletion/duplication syndromes
– Available for one commercial LDT
– Validation data not yet presented
Selected Resources
•
•
•
•
•
•
•
•
•
Lo YMD, et al. Presence of fetal DNA in maternal plasma and serum. Lancet , 1997;350:485.
Bianchi WE et al. Fetal gender and aneuploidy detection using fetal cell in maternal blood:
Analysis of NIFTY data. Prenat Diagn 2002;22:609-15.
Fan HC et al. Noninvasive diagnosis of fetal aneuploidy by shotgun sequencing DNA from
maternal blood. PNAS, 2008;105:16266.
Chiu RWK et al. Noninvasive prenatal diagnosis of fetal chromosomal aneuploidy by
massively parallel sequencing of NDA in maternal plasma. PNAS 2008; 105:20458.
Palomaki GE et al. DNA sequencing of maternal plasma to detect Down syndrome: An
international clinical validation study. Genet Med, 2011;13:913.
Bianchi DW et al. Genome-wide fetal aneuploidy detection by maternal plasma DNA
sequencing. Obstet Gynecol, 2013;119:890
Ashoor G et al. Chromosome-selective sequencing of maternal plasma cell-free DNA for firsttrimester detection of trisomy 21 and 18. Am J Obstet Gynecol, 2012;206:322 e1-5.
Norton ME et al. Non-Invasive Chromosome Evaluation (NICE) Study: results of a
multicenter prospective cohort study for detection of fetal trisomy 21 and trisomy 18. Am J
Obstet Gynecol 2012;207:137 e1-8.
Nicolaides KH et al. Validation of targeted sequencing of single-nucleotide polymorphisms for
non-invasive prenatal detection of aneuploidy of chromosomes 13, 18, 21, X and Y. Prenat
Diagn 2013;33:575-9.
Additional Free Resource for CAP Members
NOTE: please remove this page before
presenting.
CAP Member Exclusive: CAP Pathology Resource Guides
Focused on a specific hot-topic technology, these
comprehensive guides highlights current resources, select
journal articles, as well as CAP and non-CAP educational
opportunities. And don’t miss the “Insights From Early
Adopters” section in each guide to gain perspective from
pioneering colleagues.
AVAILABLE NOW:
• Molecular Pathology (single gene test, small panel)
• Genomic Analysis (large panel, exome, genome)
Learn more: go to cap.org and type Pathology Resource Guides in the
“search” field located at the top of your screen.
“An outstanding overview
of basic materials,
including the technology
and links to a number of
individuals and centers
that can assist.”
“Extremely well done,
of high practical and
educational value.”