The ACOG/SMFM practice bulletin that addresses prenatal screening for fetal chromosomal anomalies clearly states that both aneuploidy screening and diagnostic testing “should be discussed and offered to all patients regardless of maternal age or risk for chromosomal abnormality”

• Aneuploidy screening
  • Serum screening with or without NT ultrasound or
  • cfDNA screening
• Diagnostic testing (CVS or amniocentesis)

Notes

• Standard serum screening remains a first-line option along with cell-free DNA screening, also known as noninvasive prenatal testing (NIPT), because additional chromosomal and single gene defects may be picked up with NT and performance may be more comparable to NIPT if the higher NIPT test failure rates are considered
• While younger women may have a higher risk for fetal microdeletion syndromes than aneuploidy, ACOG does not recommend NIPT for microdeletions | Women who want information regarding microdeletions should be offered microarray testing using CVS or amniocentesis  

What Is NIPT?

NIPT is a blood test that utilizes cell-free DNA technology (cfDNA) to predict the risk for fetal genetic disorders during pregnancy. In 2011, NIPT was introduced as a screen for T21 (trisomy 21 or Down syndrome). Today, NIPT cover the most common aneuploidies (T21, T13 and T18), as well as sex chromosomes and may also include some microdeletions and single gene genetic disorders

How Does It Work?

• DNA fragments (outside of cells) can be found floating in the blood of all individuals
• In every pregnant woman, there are fragments from
  • Her own DNA
  • Fragments of placental DNA (generally thought to derive from the outer cytotrophoblast rather than the inner mesenchymal layer)
• A maternal blood sample is obtained ≥10 weeks (with some labs offering testing beginning at 9 weeks) of pregnancy
  • Usually performed at 11 to 13 weeks
• Fetal Fraction
  • The percentage of fetal DNA found in maternal blood is known as the fetal fraction | Typical range 3 to 13% of maternal cfDNA
  • The fetal fraction is critical to the success of NIPT | Minimum required approximately 2 to 4%
  • Sensitivity drops with lower fetal fraction and if too low, test failure will result

Technologies

• Generally, NIPT utilizes next generation sequencing and bioinformatics algorithms to interrogate DNA fragments that have been extracted from maternal samples
  • SNP-based: Bioinformatic algorithms combine risk from individual targeted single nucleotide polymorphisms (SNPs) to differentiate maternal from placental DNA fragments
    • SNP is the only analysis that can report on zygosity and individual fetal fractions in the case of twins (dependent on laboratory)
  • Quantification method: Bioinformatic algorithms determine the amount of DNA from chromosomal regions of interest | E.g. if too much chromosome 21 DNA is detected, then a ‘screen positive’ result for T21 will be generated

What Disorders Are Included?

• T13, T18 and T21
  • Most effective at screening for T21
• Chromosomes X and Y (fetal sex usually available)
• Sex chromosome conditions (depending on the lab)
  • Monosomy X (Turner syndrome)
  • 47,XXX (Triple X syndrome)
  • 47,XXY (Klinefelter syndrome)
  • 47,XYY (Jacob’s syndrome)
• Microdeletions (also known as copy number variants or CNVs) are available with more extensive panels | CNVs occur in 0.4% of pregnancies and are not related to maternal age
  • 22q11.2 deletion syndrome (DiGeorge syndrome) | 1,3000 to 1,4000 live births | May be more prevalent prenatally and cases may be missed at birth (see ‘Related ObG Topics)
  • 1p36 deletion syndrome | 1 in 5,000 to 1 in 10,000 live births
  • 4p16.3 deletion syndrome (Wolf-Hirschhorn syndrome) | 1 in 50,000 live births
  • 5p15.2 deletion syndrome (Cri du Chat syndrome) | 1 in 20,000 to 1 in 50,000 live births
  • 15q11.2 deletion syndrome (Angelman syndrome/Prader-Willi syndrome) |1 in 12,000 to 20,000 (AS) | 1 in 10,000 to 30,000 (PWS)

SYNOPSIS:

NIPT is a screening test only and not diagnostic. Cell-free DNA (cfDNA) is also commonly used with an understanding that the DNA is derived from placenta and not the fetus. NIPT utilizes next generation sequencing and bioinformatics algorithms to look at the DNA fragments in the mother and fetus, as a way of determining the likelihood of certain genetic conditions in the fetus.  While there are multiple panels available, there is consensus regarding the clinical utility of NIPS screening for T13, T18 and T21. Patient education, especially around the concept of positive predictive value (PPV) is a priority. Calculator tools are available from professional societies (see ‘Learn More – Primary Sources’ below) or ideally laboratories should be able to provide obstetric professionals with real world test performance results.

NIPT Screening Performance

Detection rates

• The following detection rates are based on recent meta-analysis (see ‘Learn More -Primary Resources’ below)
  • T21: >99% detection rate for T21
  • 98% detection rate for T18
  • 99% detection rate for T13
• Combined false positive rate of 0.13%
  • cfDNA is the most sensitive and specific screen for T21, T18 and T13
• Sensitivity and specificity are superior to standard screening for T21 and other common aneuploidies

Positive Predictive Value (PPV)

Trisomies

• NIPT generally has very high negative predictive values (NPV) | From ages 20 to 45, NPV is >99% (NSGC GSF calculator)
• PPV is also generally high but can vary based on age | Lower background risk will lower PPV
  • PPVs have been reported up to approximately 90% for T21 but tend to be lower for T18, T13 and monosomy X
  • Abnormal ultrasound will increase PPV

Microdeletions

• NIPT for microdeletion syndromes can have high sensitivity and specificity but still have very low PPV because the particular syndrome is a priori so rare | For example
  • Cri du Chat Syndrome (Using NSGC GSF calculator – see ‘Learn More – Primary Sources’ below)
    • Even if sensitivity and specificity are both >99%, the chance of a positive results being a true positive (i.e. PPV) is ≤1%
    • Negative predictive value will generally always be high for a rare disorder
  • 22q11.2 deletion syndrome (22q11.2DS) is the most common microdeletion syndrome, found in 1 in 3000 to 4000 live births | Appears to be more common prenatally with a prevalence of approximately 1 in 1000 of otherwise normal pregnancies (see ‘Learn More – Related Entries)
    • PPV in a low risk population (see ‘Learn More – Primary Sources’) has been reported to be approximately 1 in 20 and higher in pregnancies at higher risk (e.g. congenital heart disease consistent with 22q11.2DS)

Note: NIPT is a screening test and not diagnostic | Regardless of PPV, screen positive results require patients be offered invasive diagnostic testing to confirm results

• Some women may opt for a cfDNA screen after a positive serum analyte screen instead of an invasive test. This is a valid option for patients who do not want an invasive test
  • However, patients should be counseled that a cfDNA screen will not give a diagnosis and may delay a formal diagnosis
  • cfDNA will not identify all babies with chromosomal abnormalities
  • Residual risk of a chromosomal abnormality after an abnormal traditional screen followed by a normal cfDNA screen is around 2%

Reasons for a False Positive Result

• There is a high likelihood that a positive T21 NIPT screen is truly positive (approximately 90%)
• However, confirmatory testing is necessary because false positive results are possible
  • Confined placental mosaicism (since NIPT only looks at the placental DNA)
  • Vanishing twin that was aneuploid but surviving twin is normal
  • Maternal condition such as cancer (NIPT results will usually show multiple chromosomal aneuploidies)
  • Unknown

NIPT Test Failure

• Low fetal fraction
  • BMI: 10% of patients >250 lbs will have a fetal fraction <4% | May be caused by (1) dilutional effect or (2) elevation in maternal DNA due to inflammatory processes
  • Early Gestational age <9 weeks
• Other reasons for no result include
  • Laboratory failure | IVF pregnancy | Maternal drug exposure to LMWH | Racial background (e.g. Black and South Asian vs white) | Fetal aneuploidy (e.g. T13 or T18 and sex chromosome aneuploidies)

Follow-Up for ‘No Call Result’

• Inform patients that there is an increased risk of aneuploidy | Effect may be due to smaller placentas  
• Offer genetic counseling, detailed ultrasound evaluation and diagnostic testing
• Repeat screening “may be considered’
  • Success rate 75 to 80% (less with high BMI)
• Repeat screening is not advised for the following
  • Ultrasound anomalies present
  • Later gestational age where further delay may complicate access to reproductive options

Note: It is preferred that the laboratory report the fetal fraction

Single Gene and NIPT

• NIPT also has the capability to identify single gene disorders
  • Ability to identify single gene variants that would likely not be detected with carrier testing
  • Generally serious autosomal dominant disorders that may be absent in the parents, but pathogenic variants may spontaneously occur in the fetus (de novo) such as Cornelia de Lange syndrome or Achondroplasia
  • Some of these conditions are associated with advanced paternal age (≥40 at time of conception)
• Single gene screening is clinically available, although currently not recommended by ACOG

Note: ACMG provides healthcare professionals with open access ‘ACT Sheets’ to guide next steps following a positive NIPT report (see ‘Learn More – Primary Sources’ below)  

Learn More – Primary Sources: 

ACOG Practice Bulletin 226: Screening for Fetal Chromosomal Abnormalities

ACOG Practice Advisory: Cell-free DNA to Screen for Single-Gene Disorders

ACOG Statement on FDA Warning on Genetic Non-Invasive Prenatal Screening Tests | ACOG

ACMG ACT Sheets and Algorithms

Analysis of cell‐free DNA in maternal blood in screening for aneuploidies: updated meta‐analysis

Screening for trisomies by cfDNA testing of maternal blood in twin pregnancy: update of The Fetal Medicine Foundation results and meta‐analysis

NIPT/Cell Free DNA Screening Predictive Value Calculator

GHR: What is noninvasive prenatal testing (NIPT) and what disorders can it screen for?

Locate a Genetic Counselor or Genetics Services:  

Genetic Services Locator-ACMG

Genetic Services Locator-NSGC  

Genetic Services Locator-CAGC  

Locate a Maternal Fetal Medicine Specialist  

Maternal Fetal Medicine Specialist Locator-SMFM   

Related ObG Topics:

ACMG Guidance on the Use of Noninvasive Prenatal Screening in Low Risk Women

ACMG Recommendations for Preconception and Prenatal Carrier Screening

cfDNA vs. Routine Screening – How Do They Compare?

Should Amniocentesis or Chorionic Villus Sampling Be Offered to All Pregnant Women?

22q11.2 Deletion Syndrome – Key Clinical Highlights

What Is the Best Follow-Up Diagnostic Test After a High-Risk NIPT Result?

Counseling Women with NIPT Results Suggesting Cancer

Do Women at Increased Risk for Down Syndrome Following Standard Screening Prefer NIPT or Invasive Testing?