Institut für Medizinische Genetik

Test prenatale non invasivo – NIPT –

vantaggi e limiti - Infertilità: passato, presente e futuro - 30 anni dalla prima nascita in Ticino –

- 23.10.2015 -

- Beatrice Oneda-

Institut für Medizinische Genetik

Fetal cells

– 1 in a billion of total cell population

Cell-free DNA (cfDNA)

– 2–20% of total cfDNA is fetal

NIPT: Two Sources of Fetal DNA in Maternal Blood

- Beatrice Oneda-

Cell-free fetal DNA in Maternal Blood

– Released through apoptosis

– Fetal cfDNA arises from cytotrophoblastic cells of

placenta

– Released into bloodstream as small DNA fragments

(150-200bp)

– Reliably detected after 10+ weeks gestation

– Undetectable within hours postpartum

• A Reliable Analyte During Pregnancy

Institut für Medizinische Genetik

- Beatrice Oneda-

NIPT: high sensitivity and high specificity

comparison of commercial products

NITFY- BGI

• It can be performed very early during pregnancy (from the 10th week of

gestation)

• Highly accurate (detection rate >99.9%)

• No risks for the fetus

• Relatively simple to perform

- Beatrice Oneda-

Positions of International Societies

• International Society of Prenatal Diagnosis (Benn et al 2013)

• the National Society of Genetic Counselors (NSGC) (Devers et al 2013)

• the American College of Obstetricians and Gynecologists (Committee Opinion 545)

• Society for Maternal-Fetal Medicine (Committee Opinion 545)

NIPT is considered to be the most effective screening test

for aneuploidy in high-risk women (Committee Opinion

545)

- Beatrice Oneda-

Limitations of Non-Invasive Prenatal Testing

(NIPT)

• NIPT cannot:

― Detect chromosome differences other than aneuploidy of chromosomes 13, 18, 21, X and Y

• some companies are now adding screening for other trisomies and certain microdeletion syndromes

― Detect single gene conditions (25 genome copies/mL between 11 and 17 weeks of gestation, 166 bp average:

very problematic template for analysis by conventional diagnostic approaches)

• Failed results

― 2.2%-11.1% failed to give results (Zhang H et al. Ultrasound Obstet Gynecol 2015, Beamon CJ et al. Genet

Med 2014, Porreco RP et al. Am J Obstet Gynecol 2014).

― 2% of results may not be available in less than 3-4 weeks, especially in those requiring repeated sampling

(Gil MM et al. Fetal Diagn Ther 2014).

• cfDNA decreases with increasing maternal BMI, twin pregnancies and when trisomies 18 or 13 are present

• NIPT does not completely rule out aneuploidy (screening test - it needs invasive confirmation)

- Beatrice Oneda-

Specificity vs Positive predictive value

92.2% 76.6% 32.8%

Limitations of Non-Invasive Prenatal Testing (NIPT)

• False positives (and negatives) are possible

― Confined placental mosaicism

― Low fetal fraction (>8% fetal fraction adequate to achieve best performance)

― Maternal contribution (sex chromosome mosaicism - 8.56% FP - and comorbidity)

― Vanishing twin (7-8% after ART, Human Reproduction 2013; 28:924–928)

- Beatrice Oneda-

Institut für Medizinische Genetik

Positive predictive value from our routine diagnostic….

• Results in a cohort of 75 consecutive cases (Prendia Test-Genesupport, Panorama

test-Natera, Prenatest-Lifecodexx, Harmony test-Ariosa)

Positive NIPT – pathologic cytogenetic: 61.5% (40/65): 28/30 cases of

trisomy 21, 3/3 triple X, 4/5 cases of trisomy 13, 4/4 case of trisomy 18, 1/1 trisomy 22.

Positive NIPT – normal cytogenetic: 35.4% (23/65): 1 trisomy 21, 1

trisomy 13, 1 monosomy 21, 11 monosomies X, 1 XXY, 1 trisomy 7, 1 trisomy 16, 1 monosomy X +

trisomy 12,1 trisomy 17, 1 trisomy 2, 1 microduplication 16p13.12p12.3 ,1 microduplication

17q24.3q25.1, 1 microdeletion 22q11.2

(or ->13/51: 25.5%)

Inconclusive NIPT - pathologic cytogenetic: 3: 1 triploidy in a

spontaneous abortion at the 15 week of gestation, 2 trisomy 21 (one diagnosed prenatally and one

postnatally)

Other NIPT results which differ from cytogenetic: 7: 1.2 Mb terminal

deletion 1pter-p36.33 with normal NIPT (which covers the classic microdel.), 1 partial monosomy

21 with NIPT positive for trisomy 21 + 5 more cases which ended with the birth of a severely

affected child

- since April 2013 -

NIPT + cytogenetic analysis for confirmation

- Beatrice Oneda-

Case 1– Other NIPT results which differ from cytogenetic

-> birth of a severely affected child

• NT = 3.5 mm and MSS 1/274

• NIPT normal

• 24 weeks right kidney agenesis, 30 week tricuspid insufficiency, hydrops and corpus callosum agenesis

• baby was born at the 36+1 weeks with several malformations and died at the age of 1 month. Hydrops, ascites, hypoplastic corpus

callosum, pericardial effusion, agenesis of the right kidney, facial dysmorphism (blepharophimosis, periorbital puffiness, retrognathia, dysplastic ears), proximal insertion of the thumb,

bilateral singular palmar crease, bilateral camptodactily of the 5th finger, wide anterior sutures, tricuspidal insufficiency, persistent ductus Botalli, nystagmus, stenotic external meatus

acusticum, short neck, sacral dimple. Weight and length on the 50th percentile. Drinking difficulties.

46,XX,del(18)(pter->q21.2:)

28.3 Mb – 108 genes deleted

• LIKELY FALSE NEGATIVE NIPT

• INAPPROPRIATE GENETIC COUNSELLING the were not against pregnancy termination, they asked for invasive procedure, but got

reassured by the negative NIPT result

- Beatrice Oneda-

• diagnosed right after birth with very large chromosomal imbalances which were detected by

conventional karyotyping alone

• couples were counseled towards NIPT testing after abnormal US findings during pregnancies, and

no additional tests had been performed after the negative NIPT results (Increased 1st trimester

abnormal NT followed by 3rd trimester onset IUGR, 1st trimester onset of IUGR, and 2nd trimester

IUGR followed by cleft palate in the 3rd trimester

4 additional births of severely affected

babies following negative NIPT

INAPPROPRIATE GENETIC COUNSELLING

- Beatrice Oneda-

Current guidelines: NIPT in high risk

pregnancies (International Society for Prenatal Diagnosis, the National Society of Genetic Counselors, the American College of Obstetrics and Gynecology with

the Society for Maternal-Fetal Medicine and the Society of Obstetricians and Gynecologists of Canada)

• Advanced maternal age (35 years or older at

expected delivery)

• Abnormal maternal serum screening

• Personal family history of aneuploidy

• Ultrasound abnormalities indicative of

chromosomal aneuploidy (including IUGR and NT ≥ 3.5

mm)

- Beatrice Oneda-

Possible delay in establishing the final

diagnosis

• In 95-98% of cases, a result from NIPT is available within 14 days. In 2% of

cases, a result may not be available in less than 3-4 weeks (Gil et al 2014)

• 2 cases: MSS finding of >1:100, + US abnormalities first weeks second

trimester. NIPT positive, confirmed cytogenetically on invasive material. Due

to repeated NIPT final diagnosis 22nd - 24th WG, -> limit for pregnancy

termination and prohibits termination via curettage with an increase in

emotional burden

- Beatrice Oneda-

Summary

• Birth of 5 severely affected babies with very big chromosomal aberrations visible with

conventional karyotyping

• From our routine: results confirmed in 61.5% (40/68) of the cases (28/30 trisomy 21, 4/5 trisomy 13, 4/4 case

of trisomy 18)

6.2% of women who received an abnormal prenatal DNA

blood test result terminated their pregnancies without

having CVS or amnio centesis.

Am J Obstet Gynecol. 2014 Nov;211(5):527.e1-527

- Beatrice Oneda-

Conclusion

• NIPT OK for low risk pregnancy

• Maternal Wish

• Advanced maternal age

• MSS with low risk (i.e. <1:100)

• Couple against pregnancy termination

- Beatrice Oneda-

Conclusion

• More caution in offering NIPT in very high risk pregnancy (if

to be offered in such cases questionable):

• US abnormalities (including NT>3.5 mm and IUGR)

• MSS with a very high risk (i.e. >1:100 – should be

supported by data)

• In very high risk cases NIPT does not add benefit to the diagnostic

procedure:

• if positive -> invasive testing is recommended for confirmation

• if negative -> invasive testing is still recommended

• More education of health care providers/counselors

- Beatrice Oneda-

- Beatrice Oneda-

Anita Rauch

Katharina Steindl

Rosa Baldinger

Regina Reissmann

Rahim Masood

Pavel Krejci

Irina Reshetnikova

Malgorzata Taralczak

Pascal Joset

Markus Zweier

Michael Papik

Ela Papuk

Alessandra Baumer

Eva Wey

Rocchina Abbas

Jolanda Brecher

Martina Darms