Noninvasive Prenatal Molecular Karyotyping from Maternal Plasma

Yu, Stephanie C Y; Jiang, Peiyong; Choy, Kwong Wai; Chan, K C Allen; Won, Hye‐Sung; Leung, Wing Cheong; Lau, Esther Yuet Ying; Tang, Mary Hoi Yin; Leung, Tak Yeung; Lo, Y.M. Dennis; Chiu, Rossa W.̉K.

doi:10.1371/journal.pone.0060968

Cited by 77 publications

(62 citation statements)

References 26 publications

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“…The actual diagnostic impact of such algorithms would require future validation using large sample cohorts. At present, we would predict that the AND algorithm and its associated enhancement in specificity would be of value if one wishes to expand the clinical spectrum of noninvasive prenatal testing, e.g., for chromosomal microdeletions or even obtaining a genomewide molecular karyotype (28)(29)(30)(31). It is also possible that an even higher level of diagnostic specificity could be achieved by combining size, count, and DNA methylation information (32).…”

Section: Discussionmentioning

confidence: 99%

Size-based molecular diagnostics using plasma DNA for noninvasive prenatal testing

Chan

Zheng

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Noninvasive prenatal testing using fetal DNA in maternal plasma is an actively researched area. The current generation of tests using massively parallel sequencing is based on counting plasma DNA sequences originating from different genomic regions. In this study, we explored a different approach that is based on the use of DNA fragment size as a diagnostic parameter. This approach is dependent on the fact that circulating fetal DNA molecules are generally shorter than the corresponding maternal DNA molecules. First, we performed plasma DNA size analysis using pairedend massively parallel sequencing and microchip-based capillary electrophoresis. We demonstrated that the fetal DNA fraction in maternal plasma could be deduced from the overall size distribution of maternal plasma DNA. The fetal DNA fraction is a critical parameter affecting the accuracy of noninvasive prenatal testing using maternal plasma DNA. Second, we showed that fetal chromosomal aneuploidy could be detected by observing an aberrant proportion of short fragments from an aneuploid chromosome in the paired-end sequencing data. Using this approach, we detected fetal trisomy 21 and trisomy 18 with 100% sensitivity (T21: 36/36; T18: 27/27) and 100% specificity (non-T21: 88/88; non-T18: 97/97). For trisomy 13, the sensitivity and specificity were 95.2% (20/21) and 99% (102/103), respectively. For monosomy X, the sensitivity and specificity were both 100% (10/10 and 8/8). Thus, this study establishes the principle of size-based molecular diagnostics using plasma DNA. This approach has potential applications beyond noninvasive prenatal testing to areas such as oncology and transplantation monitoring.size profiling | fetal aneuploidy | next-generation sequencing | Down syndrome | Turner syndrome I n the plasma of pregnant women, cell-free fetal DNA is present in a large background of maternally derived DNA (1). Cell-free DNA molecules are mainly short fragments of less than 200 bp (2, 3). Early work based on real-time quantitative PCR has shown that fetal DNA is generally shorter than maternally derived DNA (2). Subsequently, researchers have taken advantage of such a size difference to enrich for fetal DNA in maternal plasma samples for noninvasive prenatal testing (4-6).More recently, the development of paired-end massively parallel sequencing has allowed the size distributions of fetally and maternally derived DNA to be studied at a single-base resolution (7,8). Both the size distributions of fetally and maternally derived DNA exhibit a series of peaks, including a major peak at 166 bp, a smaller peak at 143 bp, and a 10-bp periodicity below 143 bp (8). The most distinctive difference between fetal and maternal DNA in maternal plasma is that fetal DNA shows a reduced proportion of molecules of 166 bp and an increased proportion of molecules of less than 150 bp (8). In this study, we outlined the theoretical basis and explored the implementation of using molecular size analysis of plasma DNA as a diagnostic approach. We demonstrated the feasibility o...

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Section: Discussionmentioning

confidence: 99%

Size-based molecular diagnostics using plasma DNA for noninvasive prenatal testing

Chan

Zheng

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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245

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“…We analysed the z-scores genome wide in 5 Mb intervals, which provides a genome-wide visualization of all potential fetal abnormalities. Similar, although slightly different in approach, genome-wide scanning of binned z-scores has been applied before, to demonstrate the feasibility to noninvasively detect fetal subchromosomal imbalances by deep sequencing, 42,43 as well as by 'shallow-depth' sequencing. 44 Whereas those studies have focused on the ability to detect noninvasively ever smaller fetal copy number variants with the aim to reduce the need for invasive sampling and chromosomal microarray analysis, we have focused on using this additional information to improve the quality of the aneuploidy detection.…”

Section: Discussionmentioning

confidence: 99%

“…A number of case reports, as well as retrospective and prospective validation studies of novel algorithms, have recently been reported on the ability to detect segmental imbalances and even submicroscopic CNVs. 5,13,20,29,[43][44][45][46][47][48] Lau et al 29,47 reported on the use of the FCAPS pipeline detecting a fetal duplication of 18p, which is 14 Mb in size. Here, we show the detection of a segmental trisomy 18, which is 29 Mb in size, demonstrating the power of this genome-wide profiling approach.…”

Section: Discussionmentioning

confidence: 99%

Noninvasive prenatal testing using a novel analysis pipeline to screen for all autosomal fetal aneuploidies improves pregnancy management

et al. 2015

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Noninvasive prenatal testing by massive parallel sequencing of maternal plasma DNA has rapidly been adopted as a mainstream method for detection of fetal trisomy 21, 18 and 13. Despite the relative high accuracy of current NIPT testing, a substantial number of false-positive and false-negative test results remain. Here, we present an analysis pipeline, which addresses some of the technical as well as the biologically derived causes of error. Most importantly, it differentiates high z-scores due to fetal trisomies from those due to local maternal CNVs causing false positives. This pipeline was retrospectively validated for trisomy 18 and 21 detection on 296 samples demonstrating a sensitivity and specificity of 100%, and applied prospectively to 1350 pregnant women in the clinical diagnostic setting with a result reported in 99.9% of cases. In addition, values indicative for trisomy were observed two times for chromosome 7 and once each for chromosomes 15 and 16, and once for a segmental trisomy 18. Two of the trisomies were confirmed to be mosaic, one of which contained a uniparental disomy cell line. As placental trisomies pose a risk for low-grade fetal mosaicism as well as uniparental disomy, genome-wide noninvasive aneuploidy detection is improving prenatal management. INTRODUCTIONThe presence of circulating cell-free fetal DNA in the maternal plasma of the pregnant woman, 1 in combination with recent advances in massively parallel sequencing (MPS) technologies, has made noninvasive prenatal testing (NIPT) of fetal aneuploidy a reality. NIPT reduces the need for invasive sampling and the associated risk of procedure-related pregnancy loss. In 2008, it was demonstrated that noninvasive fetal aneuploidy detection by MPS was feasible. 2,3 Multiple clinical validation studies using either targeted or whole-genome sequencing demonstrated the high sensitivity and specificity of NIPT. [4][5][6][7][8][9][10][11][12][13][14][15] Although most validation studies were predominantly evaluating the clinical validity in pregnancies at increased risk of the most common aneuploidies, it was recently shown that screening all pregnant women has positive predictive values of 45.5% and 40% for detection of trisomies 21 and 18, respectively. 16 MPS for aneuploidy detection applies counting statistics to millions of sequencing reads to identify subtle changes in the small percentage of fetal DNA present in the total cell-free DNA isolated from maternal plasma. 17,18 An increase or decrease in the number of normalized sequencing reads, typically converted to a 'z-score', 18 a 'normalized chromosome value', 13 genome-wide normalized score 19 or by 'withinsample copy number aberration detector' 20 is indicative of aneuploidy for the respective chromosome. Despite the high accuracy of current NIPT testing, a baseline false-positive and false-negative rate remains. Those incorrect results may have both biological and technical causes:

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“…It would be advantageous if one could identify the tissue of origin of the copy number aberrations. For the noninvasive prenatal detection of subchromosomal copy number aberrations (33), it would be useful to identify if the plasma aberrations originated from (i) the placenta alone, (ii) the mother alone, or (iii) both the placenta and the mother. As another example, if the detection of plasma copy number aberrations is eventually used as a cancer screening tool (10,11,32), it would be clinically very informative to be able to identify the tissue of origin of the cancer for subsequent diagnostic or therapeutic procedures.…”

Section: Plasma Dna Methylation Deconvolution In Posttransplantationmentioning

confidence: 99%

Plasma DNA tissue mapping by genome-wide methylation sequencing for noninvasive prenatal, cancer, and transplantation assessments

Sun

Jiang

Chan

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Plasma consists of DNA released from multiple tissues within the body. Using genome-wide bisulfite sequencing of plasma DNA and deconvolution of the sequencing data with reference to methylation profiles of different tissues, we developed a general approach for studying the major tissue contributors to the circulating DNA pool. We tested this method in pregnant women, patients with hepatocellular carcinoma, and subjects following bone marrow and liver transplantation. In most subjects, white blood cells were the predominant contributors to the circulating DNA pool. The placental contributions in the plasma of pregnant women correlated with the proportional contributions as revealed by fetal-specific genetic markers. The graft-derived contributions to the plasma in the transplant recipients correlated with those determined using donor-specific genetic markers. Patients with hepatocellular carcinoma showed elevated plasma DNA contributions from the liver, which correlated with measurements made using tumor-associated copy number aberrations. In hepatocellular carcinoma patients and in pregnant women exhibiting copy number aberrations in plasma, comparison of methylation deconvolution results using genomic regions with different copy number status pinpointed the tissue type responsible for the aberrations. In a pregnant woman diagnosed as having follicular lymphoma during pregnancy, methylation deconvolution indicated a grossly elevated contribution from B cells into the plasma DNA pool and localized B cells as the origin of the copy number aberrations observed in plasma. This method may serve as a powerful tool for assessing a wide range of physiological and pathological conditions based on the identification of perturbed proportional contributions of different tissues into plasma.noninvasive prenatal testing | circulating tumor DNA | liquid biopsy | transplantation monitoring | epigenetics

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Noninvasive Prenatal Molecular Karyotyping from Maternal Plasma

Cited by 77 publications

References 26 publications

Size-based molecular diagnostics using plasma DNA for noninvasive prenatal testing

Size-based molecular diagnostics using plasma DNA for noninvasive prenatal testing

Noninvasive prenatal testing using a novel analysis pipeline to screen for all autosomal fetal aneuploidies improves pregnancy management

Plasma DNA tissue mapping by genome-wide methylation sequencing for noninvasive prenatal, cancer, and transplantation assessments

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