Cell-free nucleic acids as non-invasive biomarkers of gynecological disorders, fetal aneuploidy and constitutional maternal chromosomal mosaicism: Figure 1

Sir, We enjoyed reading the article entitled ‘Cell-free nucleic acids as noninvasive biomarkers of gynecological cancers, ovarian, endometrial and obstetric disorders and fetal aneuploidy’ by Traver et al. (2014). It is now widely accepted that massively parallel sequencing of maternal plasma DNA provides high sensitivity and specificity for non-invasive detection of fetal aneuploidy (Wang et al., 2014, 2015). However, recent studies have raised concerns about high false-positive rates of such noninvasive prenatal testing (NIPT) assays reaching up to 7% for trisomy 21 (Wang et al., 2015). One important consideration is the impact of the maternal genome as a possible cause of false-positive results, including gynecological conditions (Lau et al., 2013; Traver et al., 2014). There is also another important factor to consider; the presence of constitutional maternal chromosomal mosaicism. We analyzed a 28-year-old primigravida who was enrolled into our research study under an IRB-approved protocol due to abnormal fetal ultrasound showing cystic hydroma at 10 weeks of gestation. NIPT using plasma DNA sequencing, coupled with the ‘Minimally Invasive Karyotyping’ (MINK) analysis algorithm (Chu et al., 2009) was performed and returned a significant P-value consistent with a gain in copy number of chromosome 21, yet the fetal karyotype was normal as determined by chorionic villus sampling (CVS) followed by classical chromosome and FISH analyses. NIPT was initially interpreted as a false-positive finding. However, we suspected there might be a biological explanation and performed high-resolution copy number variation analyses of maternal genomic DNA, CVS derived DNA and maternal plasma DNA. By comparing the affected maternal plasma and maternal genomic DNA libraries, respectively, against normal plasma and normal genomic DNA libraries, we identified a gain of 26% in maternal plasma and a gain of 28% in maternal genomic DNA involving the 21q11.2–q22.12 chromosome region (chr21:14 350 000–32 650 000) (Fig. 1A and B). Similarly, microarray analysis on a pure maternal DNA sample revealed a gain in the chr21:14 420 615–32 635 501 (hg19) region, which is suggestive of a mosaicism for an extranumerary abnormal chromosome 21 (Fig. 1C). FISH analysis on uncultured interphase cells from maternal peripheral blood (Fig. 1D and E) showed 28% of cells with an additional chromosome 21. Our data demonstrate that an apparent ‘false-positive’ NIPT result for trisomy 21 was due to maternal somatic mosaicism for an extranumerary chromosome 21. Recurrent trisomy 21 in offspring of young, apparently healthy parents has been explained in some families by maternal germline mosaicism, also reviewed by Taylor et al. (2014). We would like to raise an important issue on interpretation of positive NIPT findings and clinical significance of chromosomal mosaicism. Sequencing of the maternal plasma may reveal a maternal genomic abnormality which may elevate the risk of future pregnancies with trisomy 21, trisomy for an abnormal chromosome or 21q deletion. Despite the lack of current apparent health consequences of mosaic trisomy 21 for the mother, she may have elevated risk for hematopoietic malignancy and/or dementia and late onset conditions. Gonadal and somatic mosaicism for point mutations is a well-documented cause for a number of genetic diseases; however low-level mosaicism for aneuploidy or structurally abnormal chromosomes is likely also a common phenomenon, yet underestimated. We propose that all positive NIPT results should be followed by diagnostic fetal testing, parental DNA analysis, and genetic counseling.