NIPT N ON – I NVASIVE P RENATAL T ESTING GENDIA Antwerp, Belgium.

NIPTNON – INVASIVE PRENATAL TESTING

GENDIA

Antwerp, Belgium

NIPT

NON – INVASIVE PRENATAL TESTING

Testing of cff DNA (cell free fetal DNA)

from maternal blood during pregnancy

for trisomy 21, 18 and 13

www.DOWNsyndromeNIPT.info

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Frequency aneuploidies

Aneuploidy Syndrome Frequency (live births)

Trisomy 21 Down syndrome 1 in 700

Trisomy 18 Edwards syndrome 1 in 5,000

Trisomy 13 Patau syndrome 1 in 16,000

Trisomy 21, 18, 13 screening

Trisomy 21 (Down syndrome)

Trisomy 18 (Edwards syndrome)

Trisomy 13 (Patau syndrome)

History Down syndrome screening

• 1980 : Amniocentesis (advanced maternal age)

• 1990 : Triple screening (T21, T18 and T13)

• 2000 : First trimester screening (T21, T18 and T13)

• 2012 : First trimester screening + NIPT (T21, T18 and T13)

• 2015 : NIPT (extensive genetic screening)

Risk Down syndrome versus Maternal Age

Age Frequency (live births)

< 35 < 0.3 %

37 0.5 %

40 1 %

50 10 %

Serum Down syndrome screening

• Triple screening ( > 1990)

– Maternal age

– Serum : AFP, HCG, free oestriol

• Combi test ( > 2000)

– Maternal age

– Nuchal translucency (NT)

– Serum : free B-HCG, PAPP-A

Classical Down syndrome screening First trimester serum screening (combi test)

Risk calculated from :

• Maternal age : the higher the age, the higher the risk of T21, T18, T13

• Nuchal translucency (NT) : the higher the NT, the higher the risk of T21, T18, T13

• Serum parameters PAPP-A and free B HCG

Classical Down syndrome screening

NT (mm)

Normal : 2.0

T21 : 3.4

T18 : 5.5

T13 : 4.0

B-HCG (MoM)

Normal : 1.0

T21 : 2.0

T18 : 0.2

T13 : 0.5

PAPP-A (MoM)

Normal : 1.0

T21 : 0.5

T18 : 0.2

T13 : 0.3

Nicolaides et al 2008

NIPT history

• 1997 : Lo et al. :cff DNA in maternal circulation

• 2001 : Fetal Rh(D) genotype

• 2006 : Sexing fetus for :

1. X-linked genetic disorders

2. Sexing (China)

• 2011 : Detection trisomy 21/18/13

• 2012 : > 100.000 patients screened in China / USA

• 2013 : Daily > 2000 NIPT tests worldwide

NIPT essentials

1. DESCRIPTION: NIPT is a DNA test on maternal blood to screen pregnancies for the most common fetal chromosome anomalies : trisomy 21 (Down syndrome), trisomy 18 (Edwards syndrome), and trisomy 13 (Patau syndrome), with optional sexing.

2. SAMPLE: Specific test kits provided by GENDIA are required: 20 ml blood in specific blood tubes is required from the mother. The samples have to be sent by Express mail to GENDIA's lab in Antwerp (Belgium), and arrive there within 2 days of withdrawal.

3. TIMING: From gestation week 10

4. TURNAROUND TIME: < 2 weeks

5. RELIABILITY: The reliability of NIPT is very high (more than 99% for trisomy 21).

6. INDICATIONS: Although NIPT can be performed in every pregnancy, it is especially indicated: • If the triple test or first trimester screening indicates an increased risk for Down syndrome or trisomy 18 • Advanced maternal age • Anxiety for invasive procedures

7. CONTRAINDICATIONS: NIPT is not the test of choice when there is: • Fetal anomalies on ultrasound • Known genetic anomalies that cannot be diagnosed by NIPT • A triplet pregnancy or vanished twin

8. PRICE: 690 Euro

Cell Free Fetal DNA (cff DNA) in Maternal Blood

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NIPT cff DNA

• < 1 % of total DNA in maternal circulation is fetal

• 5-30 % of cell-free DNA in maternal circulation is fetal

NIPT for trisomy 21

NIPT measures the ratio

of chromosome 21 sequence

versus control chromosome sequence

to exclude trisomy 21

NIPT cffDNA

• < 1 of

cffDNA DNA

Fetal

Maternal

Nl NlT21 T21

Importance of fetal fraction

Fetal Fraction

Expected ratio for Trisomy

4% 1.02

10% 1.05

20% 1.10

40% 1.20

Chromosome 21Reference

Chromosome

Fetal cfDNA

Maternal

cfDNA

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NIPT reliability

• Test : Trisomy 21, 18, 13

• Specificity : > 99 %

• Sensitivity :

T21 > 99 %

T18 > 97 %

T13 > 80 %

NO NIPT for sex aneuploidies

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• Phenotype for sex aneuploidies is highly variable

• Mosaicism in the fetus is a problem

• Mosaicism in the mother is a problem

• NIPT for sex aneuploidies is less accurate

NIPT Indications

NIPT is the test of choice when there is :

• Increased maternal age

• Increased risk on Combination or triple test

• Anxiety for invasive procedure (AC / CVS)

NIPT Contra indications

NIPT is NOT the test of choice when there is :

• Fetal anomalies on ultrasound

• A triplet pregnancy

• Vanished twin

• Known genetic anomalies that cannot be diagnosed by NIPT

NIPT Advantages versus combi test with AC / CVS

• High sensitivity (few false-negatives)

• High specificity (few false-positives)

• More than T21

• Non-invasive : no fetal risk• CVS : Risk of miscarriage : 1-2 %• AC : Risk of miscarriage : 0.5 %

NIPT Disadvantages

• Expensive (690 Euro)Combi test : 150 Euro

Combitest + AC + karyotype : 1000 Euro?

• Only testing 5 chromosomes

• Failure rate : 3 %

• Specific kits

• Not available everywhere

Companies offering NIPT

• ARIOSA (US)

• VERINATA (US)

• NATERA (US)

• SEQUENOM (US)

• BGI (China)

• LIFE-CODEXX (Germany)

GENDIA offers the HARMONY test from ARIOSA

• ARIOSA (Harmony)

NIPT results

1. Normal result : no specific follow up necessary,

unless ultrasound examination of the fetus reveals anomalies

2. Test failure : in 3 % pregnancies not enough fetal DNA :

NIPT repeated at no extra cost.

3. Abnormal NIPT result : amniocentesis or chorion biopsy

NIPT failures

If less than 4 % of cf DNA is fetal

1. High amounts of maternal cf DNA :

Maternal obesitas

2. Low amounts of fetal cf DNA :

• Trisomy 18• Triploidy ??

NIPT versus classical Down syndrome screening

Classical NIPT

False negatives 30 - 40 % 0.3 %

False positives 5 % (> 95 % of positives) < 0.1 %

Result > Week 13 > Week 12

Price 150 euro 690 Euro

NIPT versus classical screening in a country with 10 million inhabitants

Classical NIPT

Number screenings 100.000 100.000

Expected T21 200 (1/500) 200 (1/500)

Detection rate 73 % > 99 %

T21 146 198

False negatives 54 < 1

False positives 4990 < 100

Iatrogenic miscarriages 50 0

NIPT versus CVS / AC

CVS / AC

• High risk : 25-50 %(Monogenic disorder)

• Medium risk : 5-10 %

(chromosomal anomaly)

• Ultrasound anomaly (NT)

NIPT

• Low risk < 5 %

NIPT : the future

1. Array CGH – All chromosomes– Small deletions - duplications

2. Detection common monogenic mutations

- CF

3. Whole exome / genome sequencing

Message in a bottle

1. DESCRIPTION: NIPT is a DNA test on maternal blood to screen pregnancies for the most common fetal chromosome anomalies : trisomy 21 (Down syndrome), trisomy 18 (Edwards syndrome), and trisomy 13 (Patau syndrome), with optional sexing.

2. SAMPLE: Specific test kits provided by GENDIA are required: 20 ml blood in specific blood tubes is required from the mother. The samples have to be sent by Express mail to GENDIA's lab in Antwerp (Belgium), and arrive there within 2 days of withdrawal.

3. TIMING: From gestation week 10

4. TURNAROUND TIME: < 2 weeks

5. RELIABILITY: The reliability of NIPT is very high (more than 99% for trisomy 21).

6. INDICATIONS: Although NIPT can be performed in every pregnancy, it is especially indicated: • If the triple test or first trimester screening indicates an increased risk for Down syndrome or trisomy 18 • Advanced maternal age • Anxiety for invasive procedures

7. CONTRAINDICATIONS: NIPT is not the test of choice when there is: • Fetal anomalies on ultrasound • Known genetic anomalies that cannot be diagnosed by NIPT • A triplet pregnancy or vanished twin

8. PRICE: 690 Euro

www.DOWNsyndromeNIPT.info