Understanding Test Results
By Dr. Tara Sander Lee, Ph.D., Biochemistry; Charlotte Lozier Institute
Standard prenatal screening is often performed during the first and second trimester to calculate the risk of having a baby with a chromosomal disorder like Down syndrome. Maternal age, serum analyte screening for biochemical markers (such as the triple screen or quad screen), and fetal nuchal translucency (NT) measurement are considered first-line screening.[3] However, these standard screening tests do not accurately predict the risk of the child having a genetic disorder. For example, when predicting the risk of having a child with Down syndrome, there is a high false-positive rate of incorrect reporting (a negative result is reported as positive) ranging from 1-14% and incredibly low positive predictor values (PPV, the proportion of positive test results that are true positives) of 4.2%.[4]
To avoid these inaccuracies, traditional screening may be combined with other DNA screening and diagnostic testing, usually between 10-18 weeks gestation, to increase the chance of correctly predicting a risk of a genetic disorder. Diagnostic DNA tests can be performed using fetal samples obtained via amniocentesis and chorionic villus sampling (CVS). These tests are accurate, but the means to obtain fetal samples for DNA testing from the amniotic sac and placenta are invasive and carry their own risks for pregnancy loss.[5]
An advanced method of non-invasive prenatal screening (NIPS; also known as NIPT) is used to reduce the need for invasive techniques. NIPS uses cell-free fetal DNA (also known as cffDNA) found in the maternal circulation to screen for chromosomal aneuploidy such as trisomy 21. Scientists can detect cell-free fetal DNA from a mother’s blood sample as early as 4 weeks and 5 days after fertilization.[6] Cell-free fetal DNA is consistently detected from seven weeks[7], remains level between 10 and 21 weeks,[8] steadily increases after 24 weeks, peaks at birth, and then declines postpartum.[9] Therefore, most DNA screening is performed after 10 weeks gestation. NIPS is the predominant method used in both low- and high-risk patients and is endorsed by all major medical organizations to be used as the “primary test in all women.”[10]
Once the cell-free DNA sample is collected, NIPS uses advanced molecular techniques to determine a child’s genetic susceptibility to a genetic disorder.[11] The most common genetic screening includes the targets for trisomy 13, 18, 21, 22q11 deletion syndrome (i.e., DiGeorge syndrome) and others.Some platforms analyze cell-free fetal DNA fragments across the whole (or part) of the genome using next generation sequencing (NGS), targeted sequence analysis, and array-based techniques. NGS platforms that screen fragments from the entire genome can be reliable, specific, and sensitive with a reported failure rate of 0.1% (inconclusive result) and false-positive rate of <0.1%.[12] NIPS may be less invasive compared to amniocentesis and CVS, but it is far less accurate and is not diagnostic, because the cell-free fetal DNA that is collected is fragmented. Therefore, NIPS can only report whether the patient’s results are consistent with an increased risk for trisomy 21 that causes Down syndrome. Even with the most comprehensive molecular platform (i.e., NGS, array technology), NIPS will never be a diagnostic test that can definitively report a person’s known risk of having Down syndrome.
With any clinical laboratory test, especially NIPS, there are inherent limitations. No test or screen will always perform the way it should 100% of the time. From my own experience directing a genetic testing lab for almost 10 years—the DNA test is never 100% accurate every time. Underlying conditions can limit NIPS performance and interfere with test results including placental mosaicism, maternal chromosomal abnormality, vanishing twin, organ transplant, etc. Incorrect reporting due to erroneous results, technical problems, and lab errors (i.e., false positives, false negatives, mixed specimens, mislabeling, etc.) is also a possibility.
Past pregnancies may also interfere with the NIPS result. Some studies have shown that cell-free fetal DNA is rapidly cleared from the maternal blood, with 100% clearance within 1-2 days postpartum[13],[14], suggesting that fetal DNA from past pregnancies should not interfere with current tests. However, other studies have found the persistence of fetal DNA for decades in the mother.[15],[16]
These inherent limitations in DNA screening for genetic mutations will affect correct result reporting and interpretation. One widely utilized NIPT screening test on the market has a positive predictive value (PPV) of 81%, meaning that there is a significant chance that a positive test result is NOT a true positive.[17] But even this reported PPV value is deceiving, because PPV is based on test sensitivity, specificity, and the prevalence of the condition in the population being tested. Because the prevalence of Down syndrome increases with maternal age, PPVs will be higher in patients of advanced maternal age (>35 years old) and will likely increase when other aneuploidy risk factors are known (e.g., ultrasound abnormalities).[18]
A comprehensive study across 21 different centers in the United States, which included 1,914 women (mean age, 29.6 years), observed much lower positive predictor values of 45.5% for trisomy 21. This indicates that a significant proportion (over 50%) of “positive” test results for Down syndrome may not be truly positive when screening women mostly at low risk.[19] For this reason, the authors from this study highlight the “need for follow-up diagnostic testing to confirm true positive results before decisions are made about irrevocable clinical intervention.”[20] They know that some parents might tragically terminate the life of their child based on an erroneous and incorrect NIPS lab result.
Basics of Human Biology & Genetics
By Dr. Tara Sander Lee, Ph.D. Biochemistry; Charlotte Lozier Institute
At the moment of conception on day 1, when a woman’s egg is fertilized by man’s sperm to create a single-cell embryo, key steps are initiated that begin development of a new human life. This new human being contains the most basic, fundamental molecules it needs to grow, mature, and function, from the very beginning, including a very important molecule called deoxyribonucleic acid (or DNA) (FIGURE).[1] Every human being consists of DNA, which is a unique genetic code in all living things and is passed on from generation to generation, fashioning distinctive characteristics and distinct traits to each living thing. DNA is packaged in the form of chromosomes.
Human gametes (the sperm and egg) each contain 23 chromosomes, half the number of chromosomes needed to be a human being. The union of the male and female DNA during fertilization restores the number of 46 chromosomes needed to create a human being with a complete set of unchangeable DNA for that individual’s entire life. This genetic information is tightly packaged and arranged on chromosomes (numbered 1-22, plus 2 sex chromosomes X and/or Y) and stored in the nucleus of each human cell (FIGURE). The complete human genome (all 46 chromosomes) is present at conception in the earliest single-cell embryo.
DNA consists of four different nucleotides (A, G, C, T) that join together on a single strand and make a sequence. DNA is normally found as a double stranded molecule, in which two separate DNA strands are wound around each other to form a double helix and each nucleotide of one strand forms a base pair with the nucleotide of the opposite strand.
In the same way that 26 letters of the alphabet combine to create a countless number of words and sentences for communicating, so it is with DNA. The four nucleotides combine in different ways to create various sequences, called genes, which are fundamental units of genetic information that provide specific instructions for a particular property or function within the cell. There are approximately three billion base pairs of DNA sequence in each human diploid cell.[2] No two humans ever have been or will be genetically the same.
DNA is the original copy of genetic information, but DNA is not used as the direct source of instructions in the cell. Instead, DNA sends genetic information throughout the cell in the form of messenger ribonucleic acid (mRNA) that is translated to a protein “language” using amino acids. This mechanism of making protein using the information provided by mRNA is called translation. For every three nucleotide bases of mRNA (called a codon) that are translated, one amino acid is positioned in the protein sequence. The translated mRNA sequence will encode for a protein that consists of several amino acids strung together like pearls on a necklace. The final created protein product folds into a three-dimensional structure and carries out its specific function and purpose within the cell (FIGURE).
Sometimes children have a missing or extra chromosome, known as aneuploidy and do not have. Trisomy disorders, such as trisomy 13, 18, and 21 (Down syndrome) are examples of a genetic disorder caused by the presence of an extra copy of chromosome (FIGURE). It is also possible for children to have a small part of a chromosome that is missing some DNA or has extra copies of DNA. The genetic disorder 22q11.2 deletion syndrome is an example of a missing copy of DNA. Other children can have a very small change in their DNA (or mutation) that only affects one nucleotide but can have detrimental effects if the DNA change is located in a critical region of a gene necessary for cell function (FIGURE). Hearing loss and cystic fibrosis are examples of single mutation gene disorders.
Most genetic anomalies occur at conception, when the man’s sperm fuses with a woman’s egg to form a single-cell embryo. Some genetic disorders can be diagnosed before birth, other cannot.
[1] Maureen L. Condic, “When Does Human Life Begin? The Scientific Evidence and Terminology Revisited,” University of St. Thomas Journal of Law and Public Policy 8.1 (2013): 44-81; and idem, “A Scientific View of When Life Begins” (June 11, 2014), Charlotte Lozier Institute, https://lozierinstitute.org/a-scientific-view-of-when-life-begins/ (accessed November 22, 2020).
[2] International Human Genome Sequencing Consortium, “Finishing the Euchromatic Sequence of the Human Genome,” Nature 431 (2004): 931-45, https://doi.org/10.1038/nature0300
[3] Rink, B.D. and M.E. Norton, Screening for fetal aneuploidy. Semin Perinatol, 2016. 40(1): p. 35-43.
[4] Bianchi, D.W. et al., DNA sequencing versus standard prenatal aneuploidy screening. N Engl J Med 370:9, 2014.
[5] Rink, B.D. and M.E. Norton, Screening for fetal aneuploidy. Semin Perinatol 40(1): p. 35-43, 2016
[6] G. S. Dawe et al., Cell migration from baby to mother. Cell Adhesion & Migration 1:19-27, 2007.
[7] ibid
[8] Wapner, R.J and Dugoff, L. Prenatal diagnosis of congenital disorders, in Creasy and Resnik’s Maternal-Fetal Medicine: Principles and Practice 8th Edition, R., Resnik, Lockwood, C.J., Moore, T.R., Greene, M.F., Copel, J.A., and Silver, R.M., Editor. 2019, Elsevier: Philadelphia, PA. p. 506.
[9] H. Ariga et al., Kinetics of fetal cellular and cell‐free DNA in the maternal circulation during and after pregnancy: implications for noninvasive prenatal diagnosis. Transfusion 41:1524-30, 2001
[10] Wapner, R.J and Dugoff, L. Prenatal diagnosis of congenital disorders, in Creasy and Resnik’s Maternal-Fetal Medicine: Principles and Practice 8th Edition, R., Resnik, Lockwood, C.J., Moore, T.R., Greene, M.F., Copel, J.A., and Silver, R.M., Editor. 2019, Elsevier: Philadelphia, PA. p. 510.
[11] ACOG Committee on Genetics, Committee Opinion No. 640: Cell-Free DNA Screening For Fetal Aneuploidy. Obstet Gynecol. 126(3): p. e31-7, 2015
[12] Illumina Verifi Prenatal Test: https://www.illumina.com/clinical/reproductive-genetic-health/nipt/sendout-testing-for-labs.html
[13] A. Kolialexi et al., Rapid Clearance of Fetal Cells from Maternal Circulation After Delivery. Ann N Y Acad Sci 1022, 113-8, 2004
[14] Y. M. D. Lo et al., Rapid Clearance of Fetal DNA from Maternal Plasma. Am. J. Hum. Genet. 64:218–224, 1999
[15] D. W. Bianchi et al., Male progenitor cells persist in maternal blood for as long as 27 years postpartum. Proc Natl Acad Sci USA. 93:705-708, 1996
[16] Invernizzi P. et al., Presence of fetal DNA in maternal plasma decades after pregnancy. Human Genetics, 110(6): 587-591, 2002.
[17] Norton ME et al., Cell-free DNA Analysis for Noninvasive Examination of Trisomy, New England Journal of Medicine 372, 1589, 2015; doi: 10.1056/NEJMoa1407349
[18] National Society of Genetic Counselors, NIPT/Cell free DNA screening predictive value calculator. Available at: https://www.perinatalquality.org/Vendors/NSGC/NIPT/
[19] Bianchi, D.W. et al., DNA sequencing versus standard prenatal aneuploidy screening. N Engl J Med 370:9, 2014.
[20] ibid
Maternal-Fetal Surgery
By Dr. Tara Sander Lee, Ph.D. Charlotte Lozier Institute
A parent may receive news during routine ultrasound scans that their baby or babies have a structural defect. Recent medical and surgical advances have made it possible for some babies to receive life-saving treatment while still inside the womb—long before they are even born!
Fetal surgery has been successful in treating unborn babies for several conditions including the myelomeningocele (spina bifida), twin-to-twin transfusion syndrome (TTTS) sacrococcygeal teratoma (SCT), congenital diaphragmatic hernia (CDH), congenital cystic adenomatoid malformations (CCAM), severe kidney obstruction and oligohydramnios, bladder obstruction, and others.[1] There are also in utero therapies available to treat the unborn, such as maternal steroid therapy to promote lung maturity in premature fetuses and reduce respiratory distress syndrome (RDS).
Amidst the pain and despair of receiving a fetal diagnosis, fetal therapy and surgical interventions for some disorders provide real hope. Diagnoses that were previously thought to be life-limiting or life-threatening are now treated before birth, helping babies survive to birth with increased quality of life and lifespan.
For example, parents may receive a prenatal diagnosis of spina bifida, a severe disorder in which part of the baby’s spinal cord does not close properly, also known as a neural tube defect. Depending on the location of the damage, spina bifida can cause neurologic and intellectual impairment, including paralysis. While surgery in the first few days after birth can help, doctors have discovered that performing revolutionary surgery to repair the baby’s defect before birth, while still in the mother’s womb, leads to better outcomes for the child. In a groundbreaking study published in the New England Journal of Medicine, treatment of babies in the womb was so successful that the trial was stopped early, so that treatment was not withheld from babies randomly selected to receive standard postnatal repair.[2] The study found that when fetal surgery was performed on babies before 26 weeks gestation, there was a decreased risk of death or shunting before postnatal age 12 months, as well as improved mental and motor function (including independent walking) at 30 months of age.
Other parents may receive a prenatal diagnosis of twin-to-twin transfusion syndrome (TTTS), a serious and life-threatening condition for both babies caused by abnormal connections in blood flow between identical twins who share one placenta. This leads to an imbalance in blood flow in which the smaller (donor) twin to pump blood to the other, larger (recipient) twin. If left untreated, advanced forms of the disease can be fatal. This minimally invasive surgery, called fetoscopic laser ablation, uses small, fiber-optic guided instruments called endoscopes to create one small incision to disconnect the shared blood vessels in the placenta between the connecting twins. If performed promptly, fetoscopic laser surgery is the best option for saving both babies, particularly when the disease is identified in its early stages between 16- and 26-weeks’ gestation. Some surgeries have even been performed as early as 15 weeks.[3] High volume fetal therapy centers report a higher than 90% survival rate of at least one baby and a higher than 80% survival rate of both babies after fetal surgery to correct the defect.[4]
[1] Malloy, C., Chireau Wubbenhorst, M., and Sander Lee, T. The Perinatal Revolution. Issues in Law and Medicine. 34(1):15-42, 2019
[2] Adzick, N.S., et al., A randomized trial of prenatal versus postnatal repair of myelomeningocele. N Engl J Med, 2011. 364(11): p. 993-1004
[3] D Magazine. The surgeon who works on babies before they’re born. Available at: https://www.dmagazine.com/publications/d-magazine/2018/october/timothy-crombleholme-works-on-babies-before-theyre-born/; L. Lecointre et al., “Fetoscopic Laser Coagulation for Twin–Twin Transfusion Syndrome before 17 Weeks’ Gestation: Laser Data, Complications and Neonatal Outcome,” Ultrasound in Obstetrics & Gynecology 44, no. 3 (2014): 299–303, https://doi.org/10.1002/uog.13375; Baud et al., “Fetoscopic Laser Therapy for Twin-Twin Transfusion Syndrome before 17 and after 26 Weeks’ Gestation.”
[4] Twin-Twin Transfusion Syndrome (TTTS) | Children's Hospital of Philadelphia (chop.edu)
Medical Team & Medical Support Roles
by Tracy Winsor, Be Not Afraid
Following an initial prenatal diagnosis, your baby will likely require additional medical experts to better understand your baby’s exact condition along with what care and treatments are appropriate. As you carry to term, you have the right to consult with a variety of medical providers. Often a team of medical providers such as a perinatal palliative care team will be available at your hospital to support you. Below is a list of medical providers and the kind of care they provide to mothers carrying to term. Contact us to be matched with a Parent Care Coordinator who can provide you with comprehensive support in seeking appropriate medical consults or working with your palliative care program.
It will be helpful for you to consult with several perinatologists, OB’s to help you in your decision making process. We also suggest you view the PerinatalHospice.org FAQs.
- OB/GYN or Midwives: Provide prenatal care and deliver infants.
- Maternal Fetal Medicine Specialists (MFMs) or Perinatologists: Provide specialized screens, antenatal screening and manage high risk pregnancies.
- Neonatologists: Provide care for newborns in NICU
- Nurse Navigators: May direct appointments within a palliative care program.
- Chaplains: Provide spiritual support in the hospital setting.
- Social Workers: Provide a variety of supportive care to parents. Social workers may work within a Palliative Care Team or on a High Risk Obstetrics Unit, and most NICUs have Social Workers.