False Negative NIPT Results: Risk Figures for Chromosomes 13, 18 and 21 Based on Chorionic Villi Results in 5967 Cases and Literature Review

Abstract

Non-invasive prenatal testing (NIPT) demonstrated a small chance for a false negative result. Since the “fetal” DNA in maternal blood originates from the cytotrophoblast of chorionic villi (CV), some false negative results will have a biological origin. Based on our experience with cytogenetic studies of CV, we tried to estimate this risk. 5967 CV samples of pregnancies at high risk for common aneuplodies were cytogenetically investigated in our centre between January 2000 and December 2011. All cases of fetal trisomy 13, 18 and 21 were retrospectively studied for the presence of a normal karyotype or mosaicism < 30% in short-term cultured (STC-) villi. 404 cases of trisomies 13, 18 and 21 were found amongst 5967 samples (6,8%). Of these 404 cases, 14 (3,7%) had a normal or low mosaic karyotype in STC-villi and therefore would potentially be missed with NIPT. It involved 2% (5/242) of all trisomy 21 cases and 7.3% (9/123) of all trisomy 18 cases. In 1:426 (14/5967) NIPT samples of patients at high risk for common aneuploidies, a trisomy 18 or 21 will potentially be missed due to the biological phenomenon of absence of the chromosome aberration in the cytotrophoblast.

Cytogenetic confirmation of a positive NIPT result: evidence-based choice between chorionic villus sampling and amniocentesis depending on chromosome aberration

“…For trisomy 13,18, and 21 screening, the test performs much better than the traditional first trimester screening (combined test) [1]. Nevertheless, false positive as well as false negative results occur [4,5]. False results may have a biological as well as a technical origin.…”

“…The cytotrophoblast, which is studied in short-term cultured villi (STC-villi) and with NIPT is derived from the trophoblast of the blastocyst, whereas the mesenchymal core, investigated in long-term cultured villi (LTC-villi) originates from the extra-embryonic mesoderm (EEM). Both EEM and fetus are derived from the inner cell mass (ICM) of the blastocyst [5]. respectively, be trisomic or normal as well.…”

Current status of non-invasive prenatal testing (NIPT): genetic counseling, dominant methods and overall performance

“…Instead, long term cultured villi (LTC-villi) originate from the extra-embryonic mesoderm and are closer related to the embryo/fetus than STC-villi. By comparing STC-villi results with LTC-villi, it is possible to identify discordant results from both tissues: for trisomy 21 and 18, around 2% and 7.3% of true trisomic cases, respectively would have resulted in false-negative NIPT reports [6]. In particular, the high number of possible false negative trisomy 18 cases is supported by another study [36].…”

“…For screening methods such as NIPT, this includes the false-positive rate, false-negative rate, positive predictive value (PPV) and the limitations of the investigation. For NIPT, limitations include the origin of fetal DNA from cytotrophoblasts; the informative value can never be better than a CVS direct preparation [6], the fetal fraction of cell-free DNA and its dependence on exogenous factors, limitations in twin pregnancies and in particular the so-called vanishing twin, which can lead to false results. Equally, it should be pointed out that the PPV is dependent on the disease prevalence, therefore, in very young pregnant women with a low aneuploidy risk, a low significance must be assumed.…”

“…This can be explained by the mere laborious and questionable requirement of follow-up testing of every negative NIPT result by pre-or neonatal karyotyping as a means to identify the false negative rate. Nevertheless, one could estimate a false negative frequency by determining the proportion of false negative results after testing short term cultured chorionic villi (STC-villi) [6]. The latter tissue represents the cytotrophoplastic origin of cffDNA in the maternal plasma and is analyzed routinely during CVS.…”