A group of Stanford-affiliated researchers, including the lead investigator of the March of Dimes Prematurity Research Center (PRC) at the university, have published a major review paper on the state of Noninvasive Prenatal Testing (NIPT), the now routine end-of-first-trimester screening test used by about half of all pregnant people in the U.S.
Published in August in The Annual Review of Biomedical Data Science, the paper chronicles the progression of NIPT since it became clinically available in 2011, charting the scientific advances, unresolved challenges, future possibilities, and ongoing ethical considerations of the technology.
The paper was penned by a group of global leaders in the field of prenatal diagnostics, including Stanford PRC Lead Investigator David Stevenson, PRC Investigator Gary Shaw, former Stanford PRC Collaborator and current Stanford Science Fellow Mira Moufarrej, Stanford Physicist Stephen Quake, and the Director of the Eunice Kennedy Shriver National Institute of Child Health and Human Development Diana Bianchi.
It details how the test has evolved from analyzing cell-free DNA (cfDNA) to include analyzing cell-free RNA (cfRNA) and expanded from offering insights on the health of the fetus to offering them on the health of the mom, too.
“When this technology was first introduced, it wasn’t fully understood or appreciated,” said Dr. Moufarrej, the paper’s corresponding author. “There was a focus on DNA testing to screen for conditions in the fetus, but not maternal health.”
“Now we know that with one sample of maternal blood, we can get information on chromosomal and single gene disorders in the fetus, cancers, fibroids, autoimmune conditions, and infections in the pregnant individual and more recently a glimpse into due date and prenatal risks, too.”
“These advances are remarkable, but with them, and future advances, come questions about who gets to make decisions with this powerful information, and in whose benefit are those decisions being made.”
At clinical introduction, NIPT for pregnant people was comprised of a blood draw that tested cfDNA, or the DNA present in plasma, which is the liquid portion of blood that does not contain red or white blood cells.
This testing was done primarily to screen for problems with the fetus and was looking at bits of fetal cfDNA in the maternal bloodstream (a pregnant woman’s blood contains DNA from her and her baby, via the placenta).
Specifically, NIPT was originally screening only for chromosomal imbalances, or aneuploidies that are life threatening for babies or result in developmental delays. These include the most common imbalance, Down syndrome (trisomy 21), as well as Edward’s syndrome (trisomy 18) and Patau syndrome (trisomy 13).
In addition to the above trisomies, the NIPT tests of today can also screen for much rarer imbalances, chromosomal micro-deletions and single gene disorders like cystic fibrosis or sickle cell disease, among other rare genetic occurrences in the fetus. The tests can also screen for sex chromosome disorders and is popular for its ability to determine fetal sex.
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At the same time as science propelled the NIPT test forward in terms of how many conditions cfDNA could screen for, another peculiar—and critical—NIPT development was taking place.
About a decade ago, clinicians and scientists like study author Dr. Bianchi started wondering whether the NIPT test held promise to screen for maternal conditions, too. The realization came after a phenomenon where babies whose NIPT screening results were positive for aneuploidy actually ended up having normal chromosomes following later diagnostic testing. In some of these cases, the mom was subsequently diagnosed with cancer. It appeared the chromosomal aneuploidy was coming from the sample’s maternal DNA—and was indicative of cancer. In fact, the problematic DNA belonged to a tumor inside the mom and was picked up by the test.
Along with a colleague from the National Cancer Institute, Dr. Bianchi leads the IDENTIFY study, which brings pregnant and postpartum women to the NIH to learn more about the relationship between cancer and NIPT test results in clinically asymptomatic patients.
Specifically, the study aims to understand how accurate these false-positives, or “nonreportable” test results (the latter consists of an erratic and difficult to interpret result) are in identifying cancer risk.
So far, Dr. Bianchi’s study has diagnosed about 60% of study participants with cancer—a striking figure. This means that of all those with false positive or nonreportable results who were referred for the study, more than half had cancer at the time of the NIPT test without knowing it. It also means that a genomic sequencing platform designed for fetal anomalies can detect cancer in the pregnant person. However, the rest of the women enrolled in the study—the other 40%—did not develop cancer. Some had uterine fibroids that were shedding DNA picked up by the test.
The study’s other aim is to develop a guidance for how doctors should handle these results ethically in the future.
Should a doctor tell an otherwise healthy woman during pregnancy, one of the potentially most exciting times of her life, that she may have cancer? How sure is the doctor with the information at hand?
Should the patient be treated immediately, or can treatment wait until the baby is born? Should the baby be delivered early?
Many cancers can be treated safely in the late second or third trimester. Dr. Bianchi anticipates that women with these confusing results will be flagged for further testing, and if they have cancer, be treated during later pregnancy rather than waiting months until delivery, a decision that could save their lives.
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Another open question regarding NIPT concerns recessive single gene disorders like cystic fibrosis or sickle cell anemia.
The concerns are two-fold: first, because most women don’t receive single gene testing prior to conception, a positive NIPT result for a recessive single gene disorder could indicate the altered gene in the mom, instead of meaning the baby has the condition. But because doctors have no way to know with the NIPT screen alone whether the signal is coming from the mom or the baby, they suggest invasive diagnostic testing (either amniocentesis or chorionic villus sampling) that carries a small miscarriage risk.
The other issue is that these recessive single gene disorders are so rare, the NIPT predictive value (how often the test is accurate) is low (30-40%), meaning there tend to be quite a few false positives. False positives can create unnecessary angst.
With these open questions, the study authors ask: should doctors still recommend invasive diagnostic testing that carries a miscarriage risk? And should companies who provide NIPT testing continue to expand the number of diseases a woman can be tested for, even if the conditions are so rare that they could lead to a frequent false positive result?
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While the cfDNA tests are continuing to excite researchers and clinicians in the NIPT space, a close relative, cfRNA, is giving DNA a run for its money.
After researchers began uncovering cfRNA’s talents in the latter half of the 2010’s, they haven’t stopped.
While cfDNA is like the “blueprint for the house” of our bodies, cfRNA is like the “orders for that blueprint, the directions for that blueprint,” Dr. Moufarrej said.
It’s a molecule that gives instructions, and with this role, gives researchers a wealth of scientific information about the dynamics and direction of a pregnancy—even before that pregnancy goes off track.
Also analyzed through a simple blood test “liquid biopsy” (but not part of standard, doctor’s office NIPT testing), cfRNA reveals information about the fetus, the placenta, and the mom.
In the handful of years since it has been understood as a powerful information source in pregnancy, it has already been used to predict gestational age, risk of preterm birth, and preeclampsia—the latter two of which, especially, are remarkable breakthroughs.
All three came out of the Stanford PRC.
While the cfRNA preterm birth risk test does not indicate when a women will go into preterm labor, it has been shown to be effective up to two months before the due date. The cfRNA gestational age predictor test is considered as accurate as today’s gold standard, ultrasound. And the preeclampsia test is reported to be 75% accurate in predicting a woman’s risk of developing the condition later in pregnancy as early as the fifth gestational week—an incredible improvement over existing screening methods that may, once the test is clinically available in a few years, allow doctors to provide early monitoring and preventive treatment to women.
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The study authors also tackled the biggest unknown in NIPT testing: the future of innovation and possibility in the field—and the ethical questions they carry.
For example, newborn gene editing using CRISPR technology, something that was in the realm of science fiction a decade ago, is already occurring successfully around the world. One single gene disorder being reversed with CRISPR is spinal muscular atrophy (SMA), the most common genetic disease that causes infant mortality. After a baby has the procedure, SMA symptoms are mostly resolved. In a not-so-distant future, Dr. Moufarrej said, in utero gene editing could be a reality.
“In the case this type of procedure creeps into pregnancy, decisions will need to be made about which conditions should be ‘fixed,’ and those decisions can be controversial,” she said.
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Although unanswered questions remain about the ethics and rules surrounding NIPT-related decision-making, and work remains to increase the predictive value of various NIPT tests, especially ones dealing with recessive single gene disorders, according to Dr. Moufarrej, the overall progress of NIPT testing over the last two decades is breathtaking.
Most importantly, the progress, especially with cfRNA, brings the medical community one step closer to moving away from one size fits all solutions, treating each pregnancy as unique and offering precision medicine specific to each patient.
“A good example of this shift,” Dr. Moufarrej said, “is the very definition of preterm birth as babies born before 37 weeks’ gestation. Recent science has proven this cutoff to be arbitrary, as each baby develops differently, with some babies born ‘early’ being healthy and others born ‘full term’ needing medical support.”
“When we look at the insights gleaned from liquid biopsies about the biological trajectories of moms and babies during pregnancy, we realize that the taxonomy in this field may be out of date. And these advances are adding to that evolving taxonomy.”
“We’re moving into a new medical era—one that is individualized, patient-focused, proactive and very exciting.”