Prenatal Genetic Screening Through Pap Smear?

Ricki Lewis, PhD

November 02, 2016

Targeted DNA sequencing of fetal cells from Papanicolaou (Pap) smears may offer an improved and earlier route to prenatal genetic screening, according to a report published online November 2 in Science Translational Medicine.

The novel strategy, which analyzes DNA from trophoblasts that have been shed into the endocervical canal (ECC), correctly distinguished fetal DNA in 20 consecutive samples. The work paves the way for an improved route to prenatal screening and testing that could begin as early as 5 weeks' gestation.

"This would be very well accepted by physicians and patients, provided the technology is verified and endorsed as safe by a national organization, such as the [American Congress of Obstetricians and Gynecologists]," James Byrne, MD, maternal-fetal medicine specialist and chair of the Department of Obstetrics and Gynecology at the Santa Clara Valley Medical Center and affiliated clinical professor, Stanford University School of Medicine, California, told Medscape Medical News. Dr Byrne was not involved in the study.

The three types of prenatal screening tests currently in widespread use have limitations: invasiveness and use later in pregnancy (8 to 20 or more weeks) for amniocentesis and chorionic villus sampling (CVS) and low fetal fraction for cell-free fetal DNA testing (aka "NIPS" and "NIPT," for noninvasive prenatal screening or testing).

Trophoblasts give rise to the placenta and carry the same genome as the developing embryo and fetus. Thus, probing the DNA in Pap smear trophoblasts could provide the same information as the other prenatal tests but could be performed earlier and less invasively.

In addition, the new procedure overcomes the technical challenge of zeroing in on the fetal contribution among the much more abundant maternal cells and their DNA.

"Straightforward Alternative"

Prior studies have shown that it's possible to genotype cells in chorionic villus fragments shed into the ECC. In the new work, Chandni V. Jain, PhD, a research associate in the laboratory of Sascha Drewlo, PhD, and D. Randall Armant, PhD, and colleagues at Wayne State University School of Medicine, Detroit, Michigan, devised a "nuclear isolation protocol" to exclude most maternal DNA from ECC samples.

The initial separation, called TRIC (trophoblast retrieval isolation from the cervix), uses anti-HLA-G coated to magnetic nanoparticles to collect cells that express HLA-G, a human leukocyte antigen unique to trophoblasts and placental cells. The cells are then mounted on a slide and their nuclei isolated, which eliminates the clinging pieces of maternal DNA. Finally, the denuded trophoblast nuclei are lysed, releasing fetal DNA for analysis.

Teamed with detection of genetic markers that span the genome, the approach is "a straightforward alternative that uses a Pap smear to capture intact fetal trophoblast cells in numbers sufficient for next-generation sequencing as early as 5 weeks of gestation," the investigators write.

The researchers probed 20 consecutive ECC specimens that included pieces of placenta, from gestational ages of 5 to 19 weeks. Beta-human chorionic gonadotropin staining identified the cells as fetal, and fluorescence in situ hybridization distinguished the X and Y chromosomes.

Targeted sequencing genotyped 59 short tandem repeats and 94 single-nucleotide variants that spanned the 24 human chromosome types. Placental DNA served as a control because it should match fetal DNA sequences.

(The paper uses the term "fetal" even for samples from the embryonic period. Reviewers advised this because that is what practitioners use when referring to CVS and amniocentesis, Dr Drewlo told Medscape Medical News.)

All 20 fetal samples matched the placental samples, indicating a high degree of discrimination of fetal genetic markers from those in the maternal genome. The average fetal DNA fraction was 92.2% ± 6.5%, indicating minimal contamination with maternal DNA. In contrast, the fetal fraction in cell-free fetal DNA in the maternal circulation at 10 weeks' gestation is only 4% to 10%.

Noninvasive Testing an Advantage

Pap smear–based prenatal testing offers several potential advantages. A woman's body mass index does not compromise Pap screening results, as it can the other technologies. Dr Byrne lists other advantages. "Reliable testing can be obtained earlier in pregnancy. It is noninvasive, which would provide reassurance to many patients. And it is simpler than NIPT. It has tremendous commercial potential because it would utilize existing pipelines of patients."

One limitation of the study is the small sample size. Another limitation is a natural and not a technological phenomenon: placental mosaicism. A somatic mutation in a placental cell or a cell from the actual embryo or fetus could cause discordance, which would appear as a false negative or positive. "The effect of mosaicism is not well understood. It is a problem for all approaches," said Dr Drewlo.

"Expect some time to allow appropriately large sample sizes to validate the technology. If it is confirmed, and then endorsed by leading organizations, then it's easy to see the potential for rapid market dominance. This is certainly a promising report," concluded Dr Byrne.

Dr Armant and Dr Drewlo have pending patents on the isolation and use of endocervical trophoblast cells for fetal diagnosis and have licensed intellectual property to PerkinElmer Inc. Dr Byrne has disclosed no relevant financial relationships.

Sci Transl Med. Published online November 2, 2016. Abstract

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