Update Non Invasive
Prenatal TestingLauren Lynch, MD
Maternal Fetal Medicine and Clinical Genetics
Agenda
Risk of Aneuploidy
First Trimester Screening
Second Trimester Screening
Cell Free DNA in Maternal Blood
Expanded carrier screening
Risk Of Aneuploidy
Risk related to
maternal age
gestational age
prior history
ART?
Risk not related to paternal age
Risk of Down Syndrome by
maternal age
First Trimester Screening
Screens for Trisomy 21 and 18, although other
aneuploidies may be detected
Utilizes Nuchal Transluscency measurement ,
PAPP-A and HCG
10+3-13+6 weeks (CRL 39-84mm)
Detects 85% of Trisomy 21 and 90% Trisomy
18
Nuchal Transluscency
Nuchal Transluscency
Nuchal Translucency
The hypoechoic region located between the skin
and soft tissues behind the cervical spine
Presumed to represent mesenchymal edema
and is often associated with distended jugular
lymphatics
Although NT increases with gestational age, an
NT >3.5mm is abnormal at any GA
NT increases the risk of aneuploidy, cardiac
anomalies and other genetic defects
NT Measurements
Only NT measurements made by
practitioner/sonographer certified by
Nuchal Transluscency Quality Review
Quality (NTQR)accepted by qualified
labs (ntqr.org)
Ongoing quality monitoring by NTQR
First Trimester Screening
Serum Screening: PAPP-A/free B-HCG
(Ultrascreen®)
65% detection Down Syndrome
Combined Test: NT and PAPP-A/ B-
HCG (First Screen®)
85% detection Down Syndrome
Second Trimester Screening
15+0 to 20+6 weeks
Triple Screen: AFP, HCG, Estriol
Quadruple Screen: AFP, HCG, Estriol,
InhibinA
Down Syndrome Detection
Rate at a 5% FPRFIRST TRIMESTER
Serum alone (Ultrascreen) 65%
Combined Test(First Screen) 85%
SECOND TRIMESTER
Triple Screen 65%
Quadruple Screen 80%
First and Second Trimester
Screening: Possible Test
CombinationsIntegrated Testing: Combined test, no disclosure
regardlessQuadruple test (MOST SENSITIVE AT
LOWEST FALSE POSITIVE RATE)
Sequential Testing: Combined testHigh riskoffer CVS,
Not high riskquadruple test
Contingent Testing:Combined test
High riskoffer CVS,low riskno more testing,
intermediate riskquadruple test
Fetal DNA in maternal blood
Cell free DNA (cfDNA)
3-13% of cell free DNA in maternal blood is fetal (“fetal fraction”)
Mostly placental in origin
Half life is minutes
Can be detected as early as 5 weeks and consistently by 9 weeks
≈ 98% accurate in predicting fetal sex
cfDNA in Maternal Blood
Screening only for Trisomy 21, 18 ,13
Optional screening for sex chromosome abnormalities (and sex determination)
Sensitivity Tri 21 >98%
False Positive rate <0.5%
Obesity (BMI>30): less cfDNA : more often test not possible
Multiple pregnancies: less reliable
cfDNA: Positive Predictive
Value
PPV depends on prevalence of the disease
Ex: A 40y/o patient with a positive test for Tri 21
has a 87% chance of having a fetus with Tri 21 vs.
33% if she is 25y/o
Positive predictive value should be reported for
each patient with a positive result (some labs do
not)
TOP should never be done based on cfDNA results
without confirmation by CVS or amniocentesis
cfDNA: False Positives
May be caused by:
Limitations of the technology
Vanished twin
Confined placental mosaicism
Maternal malignancy
cfDNA: “No Result”
If fetal fraction is low the test is less accurate
Most labs will not report the results if fetal
fraction is less than 4%
Reasons for low fetal fraction are: high BMI,
early GA and aneuploidy
Women with “no results” are at higher risk of
having a fetus with aneuploidy (as high as 23%)
and should receive genetic counseling
cfDNA vs. First Trimester
ScreenFirst Screen
Sensitivity Tri 21 85%
False Positive 5%
May detect other
abnormalities such as
cardiac defects
May detect other
chromosome abnormalities
Cost: ≈ $250
cfDNA
Sensitivity Tri 21 > 98%
False Positive <0.5%
Only detects Trisomy
21,18,13 (optional sex
chromosomes)
Only detects aneuploidies
sought for
Cost: >$500
cfDNA vs. Invasive testingcfDNA
Sensitivity Tri 21/18/13
98%
50% of aneuploidy are other
than the above
False Positive< 0.5%
No testing related losses
Decreases number of invasive
tests required
Cost: similar to invasive
CVS/Amniocentesis
Sensitivity all aneuploidy 100%
False positive: 0%
Procedure related risk: 1:1000
n/a
Cost: similar to cfDNA
ACOG Committee Opinion
NIPTDecember 2012
cfDNA is an option as primary screening for women at increased risk of aneuploidy
Counseling regarding limitations of cfDNA recommended
cfDNA is not currently recommended for low risk women
ACOG &SMFM Committee
Opinion
September 2015“Given the performance of conventional
screening methods, the limitations of cell-free
DNA screening performance, and the limited data
on cost- effectiveness in the low-risk obstetric
population, conventional screening methods
remain the most appropriate choice for first-line
screening for most women in the general obstetric
population”
Committee Opinion (cont)
All women with positive cell free DNA testing should
be offered diagnostic testing
Cell free DNA testing not recommended in multiple
gestations
Routine cell free DNA testing for microdeletion
syndromes not recommended
Women with “uninterpretable” or “inconclusive”
results should be counselled and offered diagnostic
testing
SMFM Dec 2015
Expanded Carrier Screening
Expanded Carrier Screening
Each one of us is a carrier of 4-5 recessive
mutations
If a couple carries a mutation in the same gene
their offspring have a 25% of being affected
Until now carrier screening was based on
ethnicity
Ethnicity is increasingly difficult to determine
Expanded Carrier Testing
Current technology allows for screening of a large
number of conditions simultaneously
Most labs test for >100 diseases mostly autosomal
recessive and some X-linked
Targeted mutations are sought: not all mutations
There is a residual risk
Ideally performed before pregnancy
Can be done sequentially
Pretesting counseling should be offered
Expanded Carrier TestingExamples of diseases tested
Cystic fibrosis
Congenital adrenal hyperplasia
Sickle Cell and thalasemias
Familial dysautonomia
Fragile X
PKU
Expanded carrier testing
Carrier screening does NOT
replace newborn screening for
genetic diseases
Expanded carrier screening
Carrier Frequencies>23,000 patients
Counsyl
108 recessive diseases
24% individuals carriers for at least 1 mutation
6% carriers for 2 or more mutations
Most common mutations Alpha antitripsin, cystic fibrosis, SMA, Smith Lemli Opitz, Sickle/B thal
76% of mutations would have been missed by ACOG carrier screening guidelines
Lazarin,et al:GenetMed,2013
ACOG,SMFM &ACMG
Statement ECS
March 2015Does not replace current screening guidelines published
by individual organizations
Patients with a positive family history of a genetic
condition, genetic counseling is indicated for accurate
risk assessment and to ensure the most specific carrier
test is offered
Exceptions: Hemoglobinopathy screening should still be
done by MCV and Hgb electrophoresis and Tay Sachs
by hexosamidase A
ACOG Statement cont.Providers should choose a lab that tests for diseases that
are:
Associated with Cognitive disability
Need for surgical or medical intervention
Effect on quality of life
Prenatal diagnosis possible
May choose not to screen for:
Adult onset diseases
Mild phenotypes (ex MTHFR)
Non Invasive Testing for fetal
aneuploidy: Summary
First Trimester testing is preferred
For high risk patients First Screen, cfDNA, and
Invasive testing are options
Low Risk patients First Screen or if FS not possible
,Quadruple Screen ,cfDNA may be offered
Invasive testing should be available irrespective of age
and insurance coverage
First trimester serum screen (Ultrascreen) and Triple
Screen are no longer adequate
Summary (cont)
cfDNA is a screening test (NOT diagnostic)
which is better than First Screen in detecting
Trisomy 21,18 &13, but does NOT replace
invasive testing since 50% of aneuploidies are
not the common trisomies and would not be
detected by cfDNA
Expanded Carrier Screening:
Summary
Patient with family history of genetic disease(a
priori risk) should receive genetic counseling
Patients without family history may be tested by
ACOG guidelines or expanded screening
although there are some advantages to the later
Exception: hemoglobinopathies and Tay Sacks
2nd Trimester Screening for DS
AFP HCG Estriol Inhibin
Tri 21 ↓ ↑ ↓ ↑
Tri 18 ↓ ↓ ↓ ↓
NTD ↑
First trimester screening for DS
PAPP-a B-HCG NT
Tri 21 ↓ ↑ ↑
Tri 18 ↓ ↓ ↑