NIPT: New tool for prenatal screening
NIPT is an advanced technique in detecting fetal chromosomal abnormalities that involves analyzing cell-free fetal DNA (cffDNA) in the maternal blood sample. cffDNA can be identified in the maternal blood as early as the second trimester from 5–7 weeks. Besides being abundant, stable, and remains in maternal circulation, it makes cffDNA the most suitable marker for pregnancy-specific testing. Recently, non-invasive NIPT has rapidly changed the prenatal paradigm in which it offers a middle step between combined tests and invasive methods (4). The ‘combined test,’ which includes an ultrasound imaging (ultrasonography) and a maternal blood test for serum marker screening, is routinely used as a part of prenatal care to detect chromosome aneuploidies and other congenital disabilities in the first and second trimesters (1). However, since both of these methods are highly prone to false-positive results, in women with increased risk of an affected pregnancy, invasive or non- invasive testing (NIPT) is recommended (2). NIPT is more accurate than serum screening and produces fewer false positives, but is not currently diagnostic like invasive testing. Meanwhile, the majority of mothers are uncomfortable with invasive testing (chorionic villus sampling (CVS) or amniocentesis) because of their physical distress and the 1%–2% risk associated with induced abortion (3). Therefore, currently, mothers are offered invasive testing only if a positive result from the NIPT test needs to be confirmed by amniocentesis for a definitive diagnosis.
History
The presence of fetal DNA in the maternal blood was firstly described in 1969, interpreting the chance of separating these cells in the pregnancy period. This event produced a novel noninvasive approach for detecting prenatal aneuploidies in fetal cells. Several fetal cells were detected in maternal circulation that erythroid cells take the most lines of studies since their existence in adult blood circulation is noticeably lower than their numbers in fetal blood. Fluorescence in situ hybridization (FISH) was the first technique to detect fetal chromosomal abnormalities; however, the low performance in generating consistent and enough fetal cells prevented the transition of this approach to routine clinical practice. Following that, the emergence of highly developed parallel sequencing systems makes shifting toward aneuploidy analysis of cfDNA. In 1997, cfDNA consistently detected in maternal circulation by Lo and colleagues, demonstrating the easier and time saving with the less labor-intensive method of using cfDNA instead of fetal cells.
Mechanism
The detection process via NIPT comprises three main stages: first, the maternal blood is collected and detection involving maternal and fetal cell-free DNA, followed by the massive parallel sequencing. In this step, circulating fetal DNA in maternal plasma is sequenced parallelly as well as maternal DNA, generating tens of millions of sequences reads across the entire genome. Next, fluorescent tags are aligned to cffDNA in maternal plasma, even without separating maternal and fetal cfDNA or enriching for fetal cfDNA. These tags can be uniquely mapped (tagged) to sites from a reference human genome to identify their chromosome of origin. Finally, the tags can be counted for determining the chromosome ploidy status by an increase (trisomy) or decrease (monosomy) in the relative number of tags on the affected chromosome compared to the euploid chromosomes. Counting millions of tags allows for very high sensitivity to detect aneuploidy in a given sample.
Current applicant in the world
Since 2011 NIPT has introduced in the United States and Western Europe and tests followed by rapidly becoming available in the Middle East, South America, South and Southeast Asia, and Africa as well. In 2014, clinical translation of NIPT technologies has advanced rapidly in the way that from 2014, NIPT has presented as detecting techniques in the most frequently observed chromosome aneuploidies. These are including Down syndrome (trisomy 21, or 47,+21), Edward syndrome (trisomy 18, or 47,+18), Patau syndrome (trisomy 13, or 47,+13), and common sex chromosome aneuploidies like Turner syndrome (45, X) and Klinefelter syndrome (47, XXY). Following that, the American Congress of Obstetricians and Gynecologists (ACOG), the National Society of Genetic Counselors (NSGC) and the American College of Medical Genetics (ACMG) recognized NIPT as a screening tool. ACOG and NSGC precisely recommend it as an opportunity for high-risk mothers in specific chromosome abnormalities associated with increased maternal age, family history, or positive serum or ultrasound screening tests. All these statements mentioned the need for pre-test counseling and for that positive NIPT results to be confirmed with either chorionic villus sampling (CVS) or amniocentesis. The current use of NIPT in the whole globe is most noticeable in Europe, where one-third of countries have adopted a national policy on NIPT. Among these, Belgium and the Netherlands suggest NIPT for all pregnant women. In contrast, most other countries have implemented NIPT as an offer for only higher-risk women after the first-trimester screening. Furthermore, in Australia, combined first-trimester testing and NIPT are both frequently used as an initial prenatal screening test. However, in the USA, despite having no national agreement policies on the use of NIPT present, it is widely implemented in the clinical trials. In Iran, the clinical application of NIPT has only an indication for mixed-risk mothers provided by private sectors with a post-invasive confirmatory.
Advantages
The application of NIPT in PND testing has several advantages over combined prenatal genetic testing methods limited to less-accurate results or invasive techniques with a risk of miscarriage. NIPT comprises several benefits like having no risk of miscarriage, being early detectable, less needing experts, and assessing sub-chromosomal aberrations. One of the NIPT testing’s main plus points is the early detection of chromosomal and other genetic disorders that provide better management of pregnancies. Therefore, less financial or medical resources required to manage the health of a newborn affected by a genetic abnormality. Early detection may also allow for more rapid and safer termination of second-trimester pregnancy in countries where abortion is legally available. Another important factor is that, NIPT is only based on maternal blood sampling and as a result it may limit the need for medical experts to perform invasive diagnostic procedures, resulting in more accessible PND testing even in less-developed areas. Furthermore, recent development in DNA sequencing systems, particularly shotgun massively parallel sequencing (MPS), provides faster and cheaper conditions for detecting circulating cffDNA in maternal blood. This way, not only is the selected region of common aneuploidies identified, but the chance for broadening the current paradigm of prenatal screening will also be provided. For instance, several recent studies reveal that other full trisomies, sex-chromosomal abnormalities, and sub-chromosomal aberrations associated with rare diseases can be the additional findings of NIPT testing in the future. In addition to developed sequencing methods, other molecular techniques, namely digital polymerase chain reaction (dPCR), are the central focus of recent research for detecting chromosomal aneuploidies via NIPT. These methods are aiming at assessing the DNA methylation and also epigenetic differences between fetal and maternal DNA.
Disadvantages
Despite potential benefits, NIPT raises several ethical, practical, and financial complications that require a comprehensive overview of the appropriate implementation.
• Ethical:
NIPT caused many ethical issues since it exists too fast in the clinic, and thus the ethical implications not fully explored. A significant concern is a fact that mothers do not understand the potential outcome of the test, since, in reality, the test accuracy is not 100% as there are false-negative and false-positive results. Therefore, NIPT may lead to difficult decision making for termination of the pregnancy or continuing it with the knowledge of aneuploid baby could be born. Therefore, adequate pre‐test counseling is principal, with clinicians understanding that their priority (test accuracy) is different from the patient’s preference (test safety for their fetus).
• Effects on medical training:
Since the introduction of NIPT in Europe and the USA, it hurts invasive tests like amniocentesis and CVS, resulting in a sharp decline in the number of these tests. Therefore, medical experts will potentially forward with insufficient training opportunities or even become de-skilled. This trend will affect doctors’ performance in invasive testing and has implications on their ability to perform other invasive fetal procedures such as fetoscopic laser ablation, which require similar entry techniques.
• Financial cost at present:
In countries that NIPT is currently available, only the limited private medical sectors providing the test, with tests costing £300–900. Hence, the patients with a higher socioeconomic status only afford to undergo testing. However, the cost of NIPT may decrease soon as the applied-technology becomes cheaper.
In general, reviewing the published data on NIPT as antenatal care indicated that professional societies consider NIPT a safe and effective primary screening test for fetal aneuploidy in high-risk pregnancies. NIPT can also be a complementary test for women who have a positive maternal serum screening test. Notably, NIPT’s brilliant benefit is related to detecting chromosomal abnormalities in the early weeks of pregnancy, hence reducing the anxiety of high-risk mothers. However, appropriate use of NIPT from the policy and practice vision must be assumed. For this mean, public concerns about possible eugenic applications of NIPT need to contemplate to guarantee their ethical and cultural acceptance. Moreover, certified policies are required to confirm that NIPTI technology only uses in the most functional condition from practice and economic point of view. Finally, practice guidelines are necessary to ensure that the testing is not accepted without understanding both the technique and the potential implications of the results.
