Noninvasive prenatal an overview

Authors: Alice Poulton, Lisa Hui

Abstract

Australian health authorities recommend offering prenatal screening for fetal chromosome conditions, also known as aneuploidies (e.g. Down syndrome [trisomy 21]), to all pregnant individuals to support informed decision-making.

Noninvasive prenatal testing (NIPT) is one of 3 types of prenatal aneuploidy screening tests available in Australia. NIPT requires a maternal blood test after 10 weeks gestation. Although it doesn’t require an ultrasound, a 12- or 13-week ultrasound is recommended as it provides an opportunity for early diagnosis of major structural anomalies. NIPT is not subsidised by Medicare.

It is important to take a patient-centred approach when discussing screening options. Patients should be encouraged to consider whether knowing the test result will impact their pregnancy decision-making or preparations.

There are two main NIPT genome-wide and targeted. All currently available NIPT platforms perform well for detecting the common autosomal aneuploidies (trisomy 21, 18 and 13).

NIPT has the highest true-positive rate (highest sensitivity) and lowest false-positive rate (highest specificity) among aneuploidy screening methods, however false-positive results can occur. Genetic counselling and confirmatory invasive diagnostic testing are recommended for patients with a high-probability NIPT result, especially if they are considering pregnancy termination.

Introduction

Fetal chromosome conditions, also known as aneuploidies, occur when a fetus has an extra chromosome (trisomy), a missing chromosome (monosomy), an extra chromosome segment (duplication) or a missing segment (deletion). The most common fetal aneuploidies include Down syndrome (trisomy 21), Edwards syndrome (trisomy 18), and Patau syndrome (trisomy 13). In Australia, Down syndrome is estimated to have a population prevalence of 1 in 800. These conditions can be detected during pregnancy through prenatal screening.

Australian health authorities recommend offering prenatal screening for fetal aneuploidy to all pregnant individuals.^1^^,^^2^ Information about screening should be provided as early as possible, preferably in the first trimester. Screening outcomes help guide pregnancy decision-making and management, including preparing for the birth of a child with additional needs, managing a high-risk pregnancy, or making decisions surrounding termination of pregnancy.

This article provides an overview of the current prenatal screening tests available in Australia, with a particular focus on noninvasive prenatal testing (NIPT), a maternal blood test to screen for fetal aneuploidy. It explores the clinical indications, the different types of tests available, the conditions they can detect, and their limitations. Key terms used in the article are explained in Box 1.

Prenatal screening tests available in Australia

There are 3 types of aneuploidy screening tests available in Australia (Table 1)^3^^-^^5^:

• noninvasive prenatal testing (NIPT),
• combined first trimester screening (CFTS), and
• second trimester maternal serum screening (2TMSS).

These tests provide results as either a ‘high’ or ‘low’ probability of the fetus having a specific condition.

Among available tests, NIPT is the most sensitive (highest true-positive rate) and specific (lowest false-positive rate). NIPT analyses cell-free DNA (cfDNA) found in maternal plasma. Cell-free DNA consists of short DNA fragments released from maternal cells and the placenta, as part of normal cell turnover (Figure 1).^6^

It is essential to take a patient-centred approach when discussing screening options. Patients should be encouraged to consider what the results may mean for their pregnancy and whether knowing the outcome will impact their decision-making or preparations. Cost to the patient also needs to be discussed; NIPT is not subsidised by Medicare and may cost up to $800.

If NIPT is chosen as the preferred aneuploidy screening test, a 12- or 13-week ultrasound remains highly recommended. This scan provides an opportunity to detect major structural anomalies, confirm fetal viability, determine if there is more than one fetus (plurality), establish gestational age, and screen for other obstetric complications.^7^

What does the NIPT result mean?

Although NIPT has the highest true-positive rate (sensitivity) and the lowest false-positive rate (specificity) among aneuploidy screening methods,^3^ false-positive results can still occur and it remains a screening test rather than a diagnostic one. Additionally, sometimes NIPT may fail to provide a result.

NIPT results are typically reported as a high probability suggests an increased chance of the identified chromosomal condition, while a low probability indicates a lower likelihood. Genetic counselling and confirmatory invasive diagnostic testing – such as amniocentesis or chorionic villus sampling – are strongly recommended for all patients with a high-probability NIPT result, especially if they are considering pregnancy termination.

What are the clinical indications for NIPT?

NIPT can be used as a first-line screening test for fetal aneuploidy. It can also serve as a second-line screening test for individuals who receive an increased probability result from CFTS. If a low-probability NIPT result follows an increased probability CFTS result, a patient may opt to forgo invasive testing, avoiding the small risk of miscarriage associated with these procedures. However, as NIPT is a screening test rather than a diagnostic test, there remains a possibility that a fetal chromosomal condition is present but was not detected by NIPT. Therefore, patients with an increased probability CFTS result should have the opportunity to discuss the benefits and limitations of proceeding with NIPT, undergoing invasive diagnostic testing, or opting for no further testing. Invasive diagnostic testing is recommended for patients with a very high probability CFTS result (e.g. greater than 1 in 100), or a fetal structural anomaly, as these cases carry a high likelihood of a chromosomal condition that NIPT may not detect.^8^

What are the different types of NIPT?

Different techniques have been developed for analysing cfDNA, each with distinct capabilities. The 2 main approaches are genome-wide NIPT and targeted NIPT.^9^ Genome-wide NIPT examines cfDNA from all chromosomes, while targeted NIPT focuses on selected chromosomes (Table 2).

All currently available NIPT platforms perform well for detecting the common autosomal aneuploidies (trisomy 21, 18 and 13), with no significant differences in accuracy. Sex chromosome screening may also be offered alongside autosomal aneuploidy screening, but this requires specific pretest counselling and informed consent.

The Royal Australian and New Zealand College of Obstetricians and Gynaecologists does not recommend routine population-based screening for rare autosomal aneuploidies or segmental aneuploidies (genome-wide NIPT), or microdeletion syndromes.^2^ Patients considering NIPT for these additional conditions should be informed of the potential benefits and harms, including the prevalence of the conditions, the higher false-positive rate, and the possibility of unexpected findings.^10^

What conditions can be identified on NIPT?

NIPT does not screen for all genetic conditions. If a genetic condition is suspected, based on either ultrasound findings, reproductive carrier screening or family history, genetic counselling is recommended and prenatal diagnostic testing for the specific genetic indication should be considered.

Below is a summary of chromosome conditions that can be screened using NIPT. Table 2 compares the conditions that can be identified with genome-wide versus targeted NIPT.

Common autosomal aneuploidies

NIPT for common trisomies, including Down syndrome (trisomy 21), Edwards syndrome (trisomy 18) and Patau syndrome (trisomy 13), is well validated and highly reliable, with high sensitivity, specificity and positive predictive value.^3^

Sex chromosome aneuploidies

NIPT can screen for sex chromosome aneuploidies (X and Y chromosome anomalies), such as Turner syndrome (monosomy X), Klinefelter syndrome (XXY), triple X syndrome (XXX) and Jacobs syndrome (XYY). However, the positive predictive value for these conditions is lower compared with common autosomal aneuploidies.^11^ This is partly due to biological factors such as confined placental mosaicism (Figure 2).

Rare autosomal aneuploidies

Rare autosomal aneuploidies involve trisomy or monosomy of chromosomes other than 13, 18, 21, X and Y. Since the rare aneuploidies often lead to early pregnancy loss, they are rarely observed at birth.^12^

When a rare autosomal aneuploidy is identified on NIPT, it may be due to confined placental mosaicism or, less commonly, true fetal mosaicism (Figure 2).^13^^,^^14^ True fetal mosaicism occurs when the fetus has 2 or more genetically distinct cell lines. This contributes to the lower positive predictive value for these conditions.

Segmental aneuploidies

Segmental aneuploidies are deletions or duplications of chromosome material of 7 Mb or more in size; they can be associated with fetal structural anomalies and clinically significant syndromes. Segmental aneuploidies are identified by some genome-wide NIPT platforms. Individuals who are carriers of a balanced translocation may be particularly interested in screening, as it may identify the unbalanced form of their translocation.

Microdeletions and microduplications of chromosomes

Some targeted NIPT platforms can detect submicroscopic deletions or duplications (~1 to 3 Mb in size) along a chromosome. Regions associated with specific syndromes

• 22q11.2 deletion syndrome (DiGeorge syndrome)
• 15q11.2 (Angelman syndrome and Prader–Willi syndrome)
• 5p deletion (Cri du chat syndrome)
• 4p deletion (Wolf–Hirschhorn syndrome).

The positive predictive value of NIPT for 22q11.2 deletion syndrome is reported to be 52.6%.^15^ Data on NIPT performance for other microdeletions and microduplications are limited.^16^

Triploidy

Triploidy is a severe chromosomal anomaly that occurs when a fetus has an entire extra set of chromosomes in each cell. The only type of NIPT that can detect triploidy are those that are based on single-nucleotide polymorphism analysis. Triploid conceptions are usually detectable on ultrasound and have a high miscarriage rate, so the utility of NIPT for this condition has not been robustly demonstrated.

Other considerations when offering NIPT

Test failures

A ‘failed’ or ‘no call’ NIPT result occurs when the test is unable to provide any result. The most common cause of NIPT failure is insufficient placental cfDNA in the sample, also known as a low fetal fraction. The amount of placental cfDNA in maternal plasma can vary due to factors such as gestational age, maternal weight and maternal medical conditions. Low placental cfDNA levels are also associated with fetal aneuploidy, meaning a ‘no call’ result could increase the likelihood that a fetal chromosomal condition is present but was undetectable at the time or by that particular test.^17^

Individuals with a ‘no call’ result should receive counselling on their options, which include repeat NIPT (with success rates around 60%), invasive diagnostic testing, or other aneuploidy screening methods such as CFTS.^7^

Multiple gestations

Recent data show NIPT performs comparably in twin and singleton pregnancies, and it is the recommended aneuploidy screening method for twin pregnancies.^18^^,^^19^

Reasons for false-positive or false-negative NIPT results

Confined placental mosaicism

There may be differences in genetic makeup between the fetus and placenta, leading to an abnormal maternal plasma cfDNA profile in the presence of a euploid fetus (Figure 2). Confined placental mosaicism occurs in approximately 1% of pregnancies. Since pregnancy-derived cfDNA in maternal blood comes from the placenta, confined placental mosaicism can result in discrepancies between NIPT findings and the true fetal karyotype.^20^

When confined placental mosaicism is suspected – such as when there is a discrepancy between the NIPT result and other clinical findings, including ultrasound results – amniocentesis may be preferred over chorionic villus sampling, as it is a more direct assessment of the fetal karyotype. However, for the majority of patients with a high probability NIPT result, proceeding to chorionic villus sampling would be appropriate.

Demised twin

A demised twin can continue to contribute placental cfDNA to the maternal bloodstream for some weeks. If the demised twin was aneuploid, this may impact the accuracy of NIPT for the surviving twin, potentially leading to a false-positive result.^21^

Maternal neoplasms

Maternal neoplasms release cfDNA into maternal blood.^22^ Tumours commonly have chromosomal variations and their cfDNA can interfere with NIPT, leading to unusual results, such as multiple aneuploidies.^23^ Genome-wide NIPT is the only type of NIPT that can detect a maternal neoplasm, as tumour DNA typically involves aberrations in multiple chromosomes.^24^

Fibroids are common benign tumours that usually do not interfere with NIPT performance. However, if the fibroids are very large or numerous (e.g. total fibroid volume exceeds 400 mL), there is a higher chance of a false-positive result for segmental aneuploidy. The presence of fibroids does not increase the false-positive rate for common autosomal or sex chromosome aneuploidies.^24^

Undiagnosed maternal malignancies have been incidentally detected through NIPT. Diagnoses include lymphoma, colorectal cancer and breast cancer.^25^ However, it is important to note that NIPT is not designed as a cancer screening tool, and the performance of NIPT for detecting cancer is not well established. The frequency of undiagnosed maternal cancer being identified through genome-wide NIPT is approximately 1 in 10,000.

Patient resources

The Royal Australian and New Zealand College of Obstetricians and Gynaecologists has developed a patient information pamphlet about prenatal screening for chromosomal genetic conditions.

Murdoch Children’s Research Institue and James Cook University have developed Your Choice – a decision aid to help patients understand prenatal testing most suitable for them.

Conclusion

NIPT is a first-line screening test for fetal aneuploidies due to its high sensitivity and low false-positive rate. However, a high probability screening result should still be confirmed with invasive diagnostic testing to ensure accuracy. While NIPT can identify a range of conditions, it does have limitations that should be discussed with patients when offering these tests. Clinicians should be well informed about the scope, capabilities and cost of the specific NIPT platform they are providing, to ensure the best care.

This article is peer reviewed.