Isochromosome 21q is overrepresented among false-negative cell-free DNA prenatal screening results involving Down syndrome

Abstract

False-negative cell-free DNA (cfDNA) screening results involving Down syndrome are rare, but have high clinical impact on patients and their healthcare providers. Understanding the biology behind these results may allow for improved diagnostic follow-up and counseling. In 5 different centers offering cfDNA prenatal screening, 9 false-negative results were documented in 646 confirmed cases of trisomy 21; a false-negative rate of 1.4% (95% CI, 0.7-2.6). False-negative results included 4 cases of classical trisomy 21 and 5 cases with a de novo 21q;21q rearrangement. Two out of five rearrangements had molecular studies and were confirmed as isochromosomes. When combined with reports from the cfDNA screening literature, 8 out of 29 (28%) Down syndrome cases with a false-negative "non-invasive prenatal test" (NIPT) were associated with a 21q;21q rearrangement, compared with 2% reported in live born children with Down syndrome. In our laboratory series, evidence for placental or fetal mosaicism was present in 3 out of 3 true-positive cases involving a 21q;21q rearrangement and was confirmed in one false-negative case where placental material was available for study. Isochromosome 21q rearrangements are thus overrepresented among false-negative cfDNA screening results involving Down syndrome. Postzygotic isochromosome formation leading to placental mosaicism provides a biological cause for the increased prevalence of these rearrangements among false-negative cases. For clinical practice, a low trisomic fraction (z-score or equivalent measure) relative to the fetal fraction suggests placental mosaicism. Care should be taken as these cases may not reflect confined placental mosaicism, but rather full trisomy in the presence of a placenta containing normal cells.

Fig. 1

Singleton pregnancies (n = 132) at increased risk for trisomy 21 after cfDNA screening and their cytogenetic outcomes. For each sample, the x-axis indicates the fetal fraction (FF) and the y-axis shows the trisomic fraction. Non-mosaic true positive samples with trisomy 21 (black circles) lie close to the unity line (y = x). Samples that plot below the unity line have a trisomic fraction that is substantially lower than the FF. These samples were associated with true fetal mosaicism (TFM) for trisomy 21 (yellow circles) or mosaicism confined to the placenta (CPM) (blue circles). Trisomy 21 samples with TFM and CPM may occasionally plot on the unity line. Three true positive trisomy 21 samples with 21q;21q rearrangements are represented by red circles. Two plot well below the unity line, indicating placental mosaicism, but were associated with non-mosaic trisomy 21 in the fetus (cases 7 and 8 in main text). The third case plots on the unity line and was associated with TFM for trisomy 21 after amniocentesis, with a low proportion of trisomic cells (3/85) confirmed in the newborn (case 6 in main text)