Fetal DNA Screening May Offer Alternative to Invasive Tests

Norra MacReady

June 11, 2012

June 11, 2012 — A new screening test for fetal trisomy 18 (T18) and 21 (T21) has a high degree of accuracy and a low rate of false-positives and may spare women the need for more invasive and dangerous tests, researchers say.

T21 is the mutation associated with Down syndrome. T18 is associated with Edward syndrome and is characterized by small birth weight, congenital heart defects and other serious medical problems, and developmental delays. About 50% of children with T18 do not survive past the first week of life.

The test "offers high accuracy with a single blood test, as opposed to the complex testing algorithms and multiple blood and ultrasound examinations required with current integrated screening," lead author Mary E. Norton, MD, from the Stanford University/Lucile Packard Children’s Hospital, California, and coauthors write. "As such, this technology is potentially suitable as a replacement for current, relatively inefficient aneuploidy screening." The study was published online June 4 in the American Journal of Obstetrics and Gynecology.

At this time, even the best prenatal screening tests have false-positive rates of 2% to 3% and false-negative rates of 5% or higher, the authors explain. Positive findings on those tests must be confirmed with diagnostic tests such as amniocentesis or chorionic villus sampling (CVS), which are associated with fetal loss rates of approximately 1 in 300 procedures. In addition, there is no standard screening procedure, "with multiple algorithms available for use at various stages of pregnancy [that] can be confusing to incorporate into clinical practice."

The researchers studied chromosomal fragments known as cell-free DNA (cfDNA), which are known to circulate in maternal plasma. Previous investigators have shown cfDNA to be an accurate predictor of fetal T21, the mutation associated with Down syndrome, using massively parallel DNA shotgun sequencing (MPSS), which, although accurate, is considered too complex and expensive for wide clinical practice.

Dr. Norton and colleagues used digital analysis of selected regions (DANSR), which selectively analyzes genomic fragments of cfDNA and is thought to be more efficient and potentially less expensive than MPSS. The researchers then evaluated the results, using a novel analysis algorithm, the fetal-fraction optimized risk for trisomy evaluation (FORTE), which analyzes age-related risks and the percentage of fetal DNA to provide an individualized score of trisomy risk.

The authors conducted a prospective cohort study of women aged 18 years or older who were in their tenth or later week of a singleton pregnancy. The patients were recruited from centers in Sweden, the Netherlands, and the United States. Exclusion criteria included pregnancy with more than 1 fetus, known aneuploidy, active malignancy or a history of metastatic cancer, or prior CVS or amniocentesis during the current pregnancy.

A total of 3228 samples were included in the analysis. The women had a mean age of 34.3 years, with a range of 18 to 50 years, and were ethnically and racially diverse. The mean gestational age was 16.9 weeks, with a range of 10.0 to 38.7 weeks. Of the participants, 2410 (74.7%) underwent amniocentesis, and 818 (25.3%) underwent CVS, in addition to the blood sampling.

The DANSR and FORTE analyses detected all 81 of the cases identified on invasive testing as high-risk for T21, for a sensitivity of 100% (95% confidence interval [CI], 95.5% - 100%). Of the 2888 cases classified as "normal" by invasive testing, the blood analysis identified 2887 as low-risk for T21, for a specificity of 99.97% (95% CI, 99.8% - 99.99%), or a false-positive rate of 0.03% (95% CI, 0.002% - 0.20%).

For T18, DANSR/FORTE identified 37 of 38 cases detected on invasive testing, yielding a sensitivity of 97.4% (95% CI, 86.5% - 99.9%), and classified 2886 of the 2888 normal cases as low-risk for T18, for a specificity of 99.93% (95% CI, 99.75% - 99.98%), or a false-positive rate of 0.07% (95% CI, 0.02% - 0.25%).

"One T18 case by invasive testing was classified as Low Risk with a score of less than 0.01% (1 in 10,000)," the authors add. "Two cases with a normal karyotype by invasive testing were classified as High Risk for T18 with risk scores of 73.8% and greater than 99%."

One limitation of the study is that in 148 (4.6%) of the 3228 of the cases, the assay failed or the fetal DNA fraction was too low for analysis. "Reassuringly, we found that the assay failure rate was comparable in normal versus trisomy cases," the authors write. They also point out that cfDNA screening currently focuses on the most common and clinically significant aneuploidies. Still, "the higher throughput and lower cost make this technique potentially scalable for population screening."

Large-scale Tests in Community Settings Needed

At least 1 independent expert agrees with this assessment. "The studies published to date, including this one, have shown really promising results for this technology," Robert Hopkin, MD, told Medscape Medical News. The reported sensitivity of nearly 100% for Down syndrome is "a lot better than older tests, which missed about 20% of cases."

"The thing we're lacking is large-scale studies showing the viability of this test in community settings," warned Dr. Hopkin, who is associate professor of clinical pediatrics in the Division of Human Genetics, Cincinnati Children's Hospital, Ohio. He explained that the DNA in maternal blood samples is unstable, so 1 big unknown is whether local clinicians will be able to draw and analyze the samples while they are still viable. "I won't be completely convinced until somebody shows that."

Still, he said, "I think it will be a good test. It can be used from early on all the way through to the end of pregnancy. It is an alternative test [to more invasive procedures] that can give you the answer."

This study was sponsored by Ariosa Diagnostics. The study authors are employees of, consultants for, or board members of Ariosa Diagnostics. Dr. Hopkin has disclosed no relevant financial relationships.

Am J Obstet Gynecol. Published online June 4, 2012. Abstract

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