mQlicker
This session will utilise the mQlicker
interactive audience program
 Questions will be presented and you
will be able to respond electronically
through your mobile device
 Enable the internet on your mobile device
 Open the web browser
 Go to https://respond.cc
 When a question is presented, enter the
Session Key for that question
 Select one of the multiple choice answers
and click Submit

Non-Invasive Prenatal Testing:
Increasing Detection of Down’s
Syndrome to >99%
Dr Shian Miller
Obstetrician & Gynaecologist
BSc (Hon), MBBS, FRANZCOG
Disclaimer



This session is not funded by a
pharmaceutical nor medical device company
Dr Shian Miller is an independent
obstetrician and gynaecologist practicing
general obstetrics and general gynaecology
Dr Shian Miller currently has no involvement
with any of the companies offering NIPT and
does not collect, market, nor perform NIPT
– any promotion of NIPT is done only as a
practising obstetrician
mQlicker
This session will utilise the mQlicker
interactive audience program
 Questions will be presented and you
will be able to respond electronically
through your mobile device
 Enable the internet on your mobile device
 Open the web browser
 Go to https://respond.cc
 When a question is presented, enter the
Session Key for that question
 Select one of the multiple choice answers
and click Submit

mQlicker check
SESSION KEY: 47255
 Q1- I am a:
1. GP
2. Specialist
3. Nurse or nurse practitioner
4. Student
5. Non-medical layperson
6. Other

(mQlicker answers)
mQlicker Question
SESSION KEY: 27230
 Q2 – My current knowledge about
NIPT is:
1. What is it?
2. I’ve heard of it
3. I know a little bit about it
4. I have a good knowledge about NIPT
5. I’m an expert

(mQlicker answers)
mQlicker Question
SESSION KEY: 5530
 Q3 – The number of times I have
recommended, counselled, or
ordered NIPT for a patient before is:
1. Never
2. Once
3. A few times
4. More than 10 times
5. All the time

(mQlicker answers)
Prenatal screening




Prenatal screening & diagnosis aims to detect
conditions before a fetus is born
Can be non-invasive: maternal serum
screening, ultrasound
Or can be invasive: amniocentesis, CVS,
fetal blood sampling
Genetic testing of fetuses previously
required invasive techniques – now NonInvasive Prenatal Testing (NIPT) can
indirectly sample fetal DNA from maternal
blood
Overview
Biology of NIPT – cfDNA
 Clinical applications of NIPT
 Use of NIPT for aneuploidy screening
 Discordant results
 NIPT in practice
 Currently available NIPT options
 The future of NIPT

Cell-free DNA in maternal blood


In 1997, Lo et al published the discovery of
circulating cell-free fetal DNA in the maternal
serum
Presence of fetal DNA in maternal plasma and serum.YM Dennis Lo et al. The Lancet 1997;
350: 485-87
Fragments of DNA are released from placental
cells through apopotosis into the maternal
circulation
 Generally, cell-free DNA (cfDNA) in maternal
blood is 90% from maternal bone marrow and
10% from placenta
 Placental cfDNA is detectable before placental
circulation is developed – present in useful
amounts from 8-9 weeks gestation

Placental cell-free DNA
Increased levels with abnormal
placentation – placenta accreta, preeclampsia – and Trisomy 21 (more
apoptosis)
 Decreased levels in obese women –
thought to be due to apoptosis of adipose
tissue increasing maternal fraction

Clearance of placental cfDNA
As the DNA fragments are not contained
within a cell, they are unstable and have a
short half-life of 4-30 minutes
 Placental cfDNA is undetectable in
maternal serum within hours of delivery
 Thus, placental cfDNA detected
during a pregnancy is considered to
be representative of the current
fetus

Non-Invasive Prenatal Test (NIPT)
NIPT involves collecting maternal blood
and analysing it for placental cfDNA
 DNA fragments need to be multiplied
millions of times in a short period of time
 Although cfDNA discovered in 1997, not
until the technology became commercially
available in 2005, could NIPT be realised
 The technology was ‘Massively Parallel
Sequencing’ or Next Generation
Sequencing – complex molecular biology

Overview of NIPT process
Sequencing cfDNA





The simplified explanation is:
The cfDNA is multiplied millions of times
(both maternal and placental/fetal)
The relative quantity of each chromosome is
compared to a reference genome
Then look for over-representation of a
chromosome
Eg. Excess chromosome 21 compared to
other chromosomes suggest Down’s
syndrome
Excess Chromosome
Not just chromosomes

Using targeted sequencing, can also detect
single gene mutations and microdeletions
Clinical applications of NIPT





Aneuploidy, typically Trisomy 21 (Down’s),
Trisomy 18 (Edward’s), Trisomy 13 (Patau’s)
and XY abnormalities
Main reason for screening these trisomies is
that, unlike other trisomies, these often
progress to a live birth
Gender determination, particularly for
history of X-linked disorders
Single gene disorders – cystic fibrosis,
myotonic dystrophy
Fetal RhD typing to direct management of
pregnant women with Anti-D antibodies
mQlicker Question
SESSION KEY: 27902
 Q4 – I think the vast potential of
NIPT is:
1. Scary
2. Troubling
3. Neither concerning nor exciting
4. Promising
5. Going to open up many terrific
opportunities

(mQlicker answer)
Use of NIPT in aneuploidy screening
Since introduction in Australia in late
2012, more than 2000 women have used
NIPT for aneuploidy screening
 Generally targeted to high-risk women
but appears to be as effective in a low-risk
population
 Currently, the most appropriate
implementation strategy for NIPT in the
Australian context is still to be
determined

Current prenatal aneuploidy screening
In Australia, high uptake of combined first
trimester screening (CFTS)
 Involves a blood test for bHCG and PAPP-A
 Also an ultrasound scan at 11-13+6 weeks
 Apart from aneuploidy, screening also
beneficial for:

◦
◦
◦
◦
◦
Correct dating of pregnancy
Anatomy assessment
Increased NT is a marker for other abnormalities
Biochemistry also predicts pre-eclampsia, IUGR
Information about multiple pregnancies
NIPT compared to CFTS
Sensitivity
Specificity
False
positive
False
negative
PPV
CFTS
90%
95%
3-5%
NIPT
>99%
>99%
<0.5%
10%
<1%
4%
45%
NIPT advantages
High negative predictive value – reduces
need for invasive testing
 High detection rate of >99% for Trisomy
21 and >90% for Trisomy 18 & 13
 Only a maternal blood test
 No risk to the fetus
 Gender determination
 Available from 10 weeks gestation
onwards – useful for those who have
missed CFTS

NIPT disadvantages
Anatomical abnormalities not detected –
cfDNA even with anencephaly
 Still considered a screening test – if a
result is ‘positive’, still need invasive
testing (CVS/amniocentesis) to confirm
 Does not target atypical chromosome
abnormalities – account for 30% of
abnormal karyotypes
 Some types of NIPT not suitable for twins
 Cost – no Medicare rebate

Discordant results


Low false positive rate of 0.2%
Causes include:
◦
◦
◦
◦
◦
◦
◦
◦
◦
Placental mosaicism
Co-twin demise
Organ transplant eg kidney transplant; male donor
Bone marrow transplant
False negative 45XO – cryptic mosaic – fetus XO,
placenta XX
Maternal chromosomal abnormality
Increasing maternal age – white cells can ‘lose an X’
and become ‘XO’ – somatic mosaicism
Laboratory error
Occult cancer – sheds chromosomes chaotically
Multiple aneuploidy in occult cancer
No result from NIPT
Test failure rate <5%
 Obese women have a significantly
reduced fetal fraction of cfDNA
 For a maternal weight of 160kg, 50% of
samples will have fetal fraction <4%
 If sample taken too early in the pregnancy,
may have false negative results for Y
chromosome or Rhesus status

NIPT for Twin pregnancies
Currently there is limited experience with
performance of NIPT for twins
 Results to date are promising
 Lower sensitivity for detecting aneuploidy
 Fetal fraction is not double that of a
singleton but less than that – may affect
test performance

NIPT in practice







NIPT can be performed from 10 weeks gestation
Currently no controls or guidelines for use
Women have taken it up in an ad hoc way
As it involves only a maternal blood test and poses
no risk to the fetus, very attractive to mothers
Previously prohibitively expensive but now easily
obtained at under $500 (becoming comparable to
CFTS ~$300)
Ease of access may mean implications of test results
may not be considered adequately
Risk of commercialisation – marketing direct to
consumer
Models for the use of NIPT in
aneuploidy screening
CFTS – if high risk >1:300, offer NIPT or
invasive testing
CFTS – tiered management (contingent)
1.
2.
◦
◦
◦
◦
Very low risk <1:1000 – no further testing
Low risk 1:1000 to 1:300 – offered no
further testing or NIPT
High risk >1:300 – offered NIPT or invasive
testing
Very high risk >1:10 – invasive testing
Other models for using NIPT
3.
4.
5.
Routine NIPT at 10 weeks gestation
then ultrasound at 12 weeks (for dating,
anatomy assessment) – if no result,
recommend CFTS
Routine NIPT, ultrasound, and
biochemistry
NIPT only for high risk women: maternal
age >35, previous history of aneuploidy,
known parental balanced translocation
mQlicker Question
SESSION KEY: 81114
 Q5 – The model that appeals to me is:
1. CFTS then if high-risk, NIPT or invasive
testing
2. CFTS with contingent screening
3. Routine NIPT then USS
4. Routine NIPT then USS and biochemistry
5. NIPT only for high-risk women; CFTS for
low-risk women

(mQlicker answers)
Ethical considerations with NIPT





Gender determination used for gender
selection
Use in paternity testing
Discovery of previously unknown maternal
genetic disorder, eg Turner mosaicism
Screening non-specifically for genetic
alterations may lead to findings of uncertain
significance
Termination decision based on NIPT result
only (trend in the USA)
Current availability of NIPT in Australia
Previously required shipping samples
overseas in a Streck tube (contains a
preservative that allows DNA to remain
stable even in ambient temperatures)
 Now, laboratories available in Australia
 Turnaround time for overseas labs 7-10 days
– Australian labs 3-4 days
 Available at major pathology labs, obstetric
ultrasound places, fertility specialists and
private obstetricians
 Different labs & technology but very similar
NPV and sensitivity and specificity

GPs will be the front-line
Increasing public awareness of NIPT
 Consultations regarding NIPT are likely to
be initiated by pregnant women
themselves
 GPs will be the first point of contact
 GPs should be able to provide counseling
and guidance to inform appropriate
decision making
 Likely, GPs will order NIPT for patients
before contact with obstetrician/hospital

mQlicker Question
SESSION KEY: 63987
 Q6 – Now I feel confident counseling
a patient about NIPT:
1. Strongly disagree
2. Disagree
3. Neutral
4. Agree
5. Strongly agree

(mQlicker answers)
mQlicker Question
SESSION KEY: 83696
 Q7 – Now, I think the vast potential
of NIPT is:
1. Scary
2. Troubling
3. Neither concerning nor exciting
4. Promising
5. Going to open up many terrific
opportunities

(mQlicker answers)
CASE ONE
19yo G1P0 at 10 weeks gestation
 Planned pregnancy; excited
 No significant history

CASE ONE – mQlicker Question
SESSION KEY: 65943
 What aneuploidy screening do you
recommend?
1. None required
2. Offer CFTS
3. Offer NIPT with anatomy USS
4. Offer invasive testing (CVS, amnio)

(mQlicker answers)
CASE TWO
40yo G1P0 at 10 weeks gestation
 Planned pregnancy; excited
 Fit and well, no significant history

CASE TWO – mQlicker Question
SESSION KEY: 32294
 What aneuploidy screening do you
recommend?
1. None required
2. Offer CFTS
3. Offer NIPT with anatomy USS
4. Offer invasive testing (CVS, amnio)

(mQlicker answers)
CASE THREE
30yo G3P2 at 10 weeks gestation
 First child had Down’s syndrome
 Parental karyotypes normal
 No other history of note

CASE THREE – mQlicker Question
SESSION KEY: 18373
 What aneuploidy screening do you
recommend?
1. None required
2. Offer CFTS
3. Offer NIPT with anatomy USS
4. Offer invasive testing (CVS, amnio)

(mQlicker answers)
CASE FOUR
28yo G1P0 at 10 weeks gestation
 Known maternal balanced translocation
of chromosome 21
 No other history of note

CASE FOUR – mQlicker Question
SESSION KEY: 91700
 What aneuploidy screening do you
recommend?
1. None required
2. Offer CFTS
3. Offer NIPT with anatomy USS
4. Offer invasive testing (CVS, amnio)

(mQlicker answers)
The future of NIPT
Still some doubt about the place of NIPT
in prenatal screening in Australia
 The USA has taken it up enthusiastically –
uptake of CFTS has always been poorer
 Prices will continue to fall
 Professional organisations are under
pressure to establish guidelines for use
 NIPT is very likely to take a central role
in the future of Australian prenatal
screening

mQlicker Question
SESSION KEY: 74249
 Q8 – My feelings about NIPT are:
1. I worry Gattaca (eugenics) is going to
become the reality
2. I think NIPT probably won’t live up to the
hype
3. My feelings about NIPT are neutral
4. I think NIPT is going to be useful
5. I agree with Shian – NIPT is going to be
the future!

(mQlicker answers)