internationale publicaties

Selectie van internationale publicaties:

Taylor-Phillips S, et al. Accuracy of non-invasive prenatal testing using cell-free DNA for detection of Down, Edwards and Patau syndromes: a systematic review and meta-analysis. BMJ Open, 2016. 6: p. e010002.

Chitty L, et al. Uptake, outcomes, and costs of implementing non-invasive prenatal testing for Down’s syndrome into NHS maternity care: prospective cohort study in eight diverse maternity units. BMJ 2016;354:i3426.

Hill M, J et al. Preferences for prenatal tests for Down syndrome: an international comparison of the views of pregnant women and health professionals. Eur J Hum Genet. 2016; 24:968-75.

Norton, ME, et al. Cell-free DNA Analysis for Noninvasive Examination of Trisomy. New England Journal of Medicine, 2015. 372:1589-1597.

Bianchi DW, et al., Noninvasive Prenatal Testing and Incidental Detection of Occult Maternal Malignancies. JAMA, 2015. 314(2):162-169

Amant et al. Presymptomatic Identification of Cancers in Pregnant Women During Noninvasive Prenatal Testing. JAMA Oncol 2015;1:814-9.

Gil MM, et al., Analysis of cell-free DNA in maternal blood in screening for fetal aneuploidies: updated meta-analysis. Ultrasound in Obstetrics & Gynecology 2015. 45:249-266

Yaron Y, et al. Current Status of Testing for Microdeletion Syndromes and Rare Autosomal Trisomies Using Cell-Free DNA Technology. Obstet Gynecol. 2015;126(5):1095-9.

[toggle Title=”Benn P. et al.
Position statement from the Chromosome Abnormality Screening Committee on behalf of the Board of the International Society for Prenatal Diagnosis.
Prenatal Diagnosis 2015 “]

Benn P, Borrell A, Chiu RW, Cuckle H, Dugoff L, Faas B, Gross S, Huang T, Johnson J, Maymon R, Norton M, Odibo A, Schielen P, Spencer K, Wright D, Yaron Y.

No abstract available.

Prenat Diagn. 2015 Aug;35(8):725-34

[/toggle]

[toggle Title=”Bianchi DW
Pregnancy: Prepare for unexpected prenatal test results

Nature 2015″]

Nature juni 2015
[/toggle]

[toggle Title=”Bianchi DW etal.
DNA sequencing versus standard prenatal aneuploidy screening
NEJM 2014″]

Bianchi DW1, Parker RL, Wentworth J, Madankumar R, Saffer C, Das AF, Craig JA, Chudova DI, Devers PL, Jones KW, Oliver K, Rava RP, Sehnert AJ; CARE Study Group.

Abstract

BACKGROUND:

In high-risk pregnant women, noninvasive prenatal testing with the use of massively parallel sequencing of maternal plasma cell-free DNA (cfDNA testing) accurately detects fetal autosomal aneuploidy. Its performance in low-risk women is unclear.

METHODS:

At 21 centers in the United States, we collected blood samples from women with singleton pregnancies who were undergoing standard aneuploidy screening (serum biochemical assays with or without nuchal translucency measurement). We performed massively parallel sequencing in a blinded fashion to determine the chromosome dosage for each sample. The primary end point was a comparison of the false positive rates of detection of fetal trisomies 21 and 18 with the use of standard screening and cfDNA testing. Birth outcomes or karyotypes were the reference standard.

RESULTS:

The primary series included 1914 women (mean age, 29.6 years) with an eligible sample, a singleton fetus without aneuploidy, results from cfDNA testing, and a risk classification based on standard screening. For trisomies 21 and 18, the false positive rates with cfDNA testing were significantly lower than those with standard screening (0.3% vs. 3.6% for trisomy 21, P<0.001; and 0.2% vs. 0.6% for trisomy 18, P=0.03). The use of cfDNA testing detected all cases of aneuploidy (5 for trisomy 21, 2 for trisomy 18, and 1 for trisomy 13; negative predictive value, 100% [95% confidence interval, 99.8 to 100]). The positive predictive values for cfDNA testing versus standard screening were 45.5% versus 4.2% for trisomy 21 and 40.0% versus 8.3% for trisomy 18.

CONCLUSIONS:

In a general obstetrical population, prenatal testing with the use of cfDNA had significantly lower false positive rates and higher positive predictive values for detection of trisomies 21 and 18 than standard screening. (Funded by Illumina; ClinicalTrials.gov number, NCT01663350.)

NEJM, February 2014
[/toggle]

[toggle Title=”Bianchi DW etal.
Integration of Noninvasive DNA Testing for Aneuploidy into Prenatal Care: What Has Happened Since the Rubber Met the Road
Clinical Chemistry 2013 “]

Bianchi DW, Wilkins-Haug L. 

Mother Infant Research Institute at Tufts Medical Center and Floating Hospital for Children and Departments of Pediatrics and Obstetrics and Gynecology, Tufts University School of Medicine, Boston, MA;

Over the past 2 years, noninvasive prenatal testing (NIPT), which uses massively parallel sequencing to align and count DNA fragments floating in the plasma of pregnant women, has become integrated into prenatal care. Professional societies currently recommend offering NIPT as an advanced screen to pregnant women at high risk for fetal aneuploidy, reserving invasive diagnostic procedures for those at the very highest risk.Content:In this review, we summarize the available information on autosomal and sex chromosome aneuploidy detection. Clinical performance in CLIA-certified, College of American Pathology-accredited laboratories appears to be equivalent to prior clinical validation studies, with high sensitivities and specificities and very high negative predictive values. The main impact on clinical care has been a reduction in invasive procedures. Test accuracy is affected by the fetal fraction, the percentage of fetal DNA in the total amount of circulating cell-free DNA. Fetal fraction is in turn affected by maternal body mass index, gestational age, type of aneuploidy, singleton vs multiples, and mosaicism. Three studies comparing NIPT to serum or combined screening for autosomal aneuploidy all show that NIPT has significantly lower false-positive rates (approximately 0.1%), even in all-risk populations. A significant number of the discordant positive cases have underlying biological reasons, including confined placental mosaicism, maternal mosaicism, cotwin demise, or maternal malignancy.Summary:NIPT performs well as an advanced screen for whole chromosome aneuploidy. Economic considerations will likely dictate whether its use can be expanded to all risk populations and whether it can be applied routinely for the detection of subchromosome abnormalities.

Clinical Chemistry, nov 2013
[/toggle]

[toggle Title=”Ashoor G etal.
Trisomy 13 detection in the first trimester of pregnancy using a chromosome-selective cell-free DNA analysis method.
Ultrasound Obstet Gynaecology 2013 “]

Ashoor G, Syngelaki A, Wang E, Struble C, Oliphant A, Song K, Nicolaides KH.

Harris Birthright Research Centre for Fetal Medicine, King’s College Hospital, London, UK.

OBJECTIVE:
To assess the performance of chromosome-selective sequencing of maternal plasma cell-free DNA (cfDNA) in non-invasive prenatal testing for trisomy 13.

METHODS:
Two-phase case-control study on a single plasma sample per case. The first phase was used to optimize the trisomy 13 algorithm, which was then applied to a second dataset to determine the risk score for trisomy 13 by laboratory personnel who were blinded to the fetal karyotype.

RESULTS:
In the first phase, trisomy 13 risk scores were given for 11 cases of trisomy 13 and 145 euploid cases at 11-13 weeks’ gestation. The test identified seven (63.6%) cases of trisomy 13 with no false positives. The trisomy 13 algorithm was subsequently modified and the trisomy 13 risk score was > 99% in all 11 cases of trisomy 13 and < 0.01% in all 145 euploid cases. In the second phase, the new algorithm was used to generate trisomy 13 risk scores for 10 cases of trisomy 13 and 1939 euploid cases. The trisomy 13 risk scores were > 99% in eight (80.0% (95% confidence interval (CI), 49.0-94.3%)) cases of trisomy 13. In the 1939 euploid cases the risk score for trisomy 13 was < 0.01% in 1937 (99.9%), 0.79% in one, and > 99% in one. Therefore, at the predefined risk cut-off of 1% for classifying a sample as high or low risk, the false-positive rate (FPR) was 0.05% (95% CI, 0.0-0.3%).

CONCLUSIONS:
Chromosome-selective sequencing of cfDNA can detect the majority of cases of trisomy 13 at an FPR of less than 0.1%.

Ultrasound Obstet Gynecol. 2013 Jan
[/toggle]

[toggle Title=”Ashoor G etal.
Fetal fraction in maternal plasma cell-free DNA at 11-13 weeks’ gestation: relation to maternal and fetal characteristics.
Ultrasound Obstet Gynaecology 2013 “]

Ashoor G, Syngelaki A, Poon LC, Rezende JC, Nicolaides KH.

Harris Birthright Research Centre for Fetal Medicine, King’s College Hospital, London, UK.

OBJECTIVE: To examine the possible effects of maternal and fetal characteristics on the fetal fraction in maternal plasma cell-free (cf) DNA at 11-13 weeks’ gestation and estimate the proportion of pregnancies at high risk of non-invasive prenatal testing (NIPT) failure because the fetal fraction is less than 4%.

METHODS:
In 1949 singleton pregnancies at 11-13 weeks’ gestation cf-DNA was extracted from maternal plasma. Chromosome-selective sequencing of non-polymorphic and polymorphic loci, where fetal alleles differ from maternal alleles, was used to determine the proportion of cf-DNA that was of fetal origin. Multivariable regression analysis was used to determine significant predictors of the fetal fraction among maternal and fetal characteristics.

RESULTS:
The fetal fraction decreased with increased maternal weight, it was lower in women of Afro-Caribbean origin than in Caucasians and increased with fetal crown-rump length, serum pregnancy-associated plasma protein-A, serum free β-human chorionic gonadotropin, smoking and trisomy 21 karyotype. The median fetal fraction was 10.0% (interquartile range, 7.8-13.0%) and this decreased with maternal weight from 11.7% at 60 kg to 3.9% at 160 kg. The estimated proportion with fetal fraction below 4% increased with maternal weight from 0.7% at 60 kg to 7.1% at 100 kg and 51.1% at 160 kg.

CONCLUSIONS:
Fetal fraction in maternal plasma cf-DNA is affected by maternal and fetal characteristics.

Ultrasound Obstet Gynecol. 2013 Jan
[/toggle]

[toggle Title=”Nicolaides K.H. etal.
Noninvasive prenatal testing for fetal trisomies in a routinely screened first-trimester population
AJOG 2012 “]

Nicolaides KH, Syngelaki A, Ashoor G, Birdir C, Touzet G.

Harris Birthright Research Centre for Fetal Medicine, King’s College Hospital, London, UK.

OBJECTIVE:
We sought to assess performance of noninvasive prenatal testing for fetal trisomy in a routinely screened first-trimester pregnancy population.

STUDY DESIGN:
This was a cohort study of 2049 pregnant women undergoing routine screening for aneuploidies at 11-13 weeks’ gestation. Plasma cell-free DNA analysis using chromosome-selective sequencing was used. Laboratory testing on a single plasma sample of 2 mL was carried out blindly and results were provided as risk score (%) for trisomies 21 and 18.

RESULTS:
Trisomy risk scores were given for 95.1% (1949 of 2049) of cases including all 8 with trisomy 21 and 2 of the 3 with trisomy 18. The trisomy risk score was >99% in the 8 cases of trisomy 21 and 2 of trisomy 18 and <1% in 99.9% (1937 of 1939) of euploid cases.

CONCLUSION:
Noninvasive prenatal testing using chromosome-selective sequencing in a routinely screened population identified trisomies 21 and 18 with a false-positive rate of 0.1%.

[/toggle]

[toggle Title=”Hill M etal.
Women’s and health professionals’ preferences for prenatal tests for Down syndrome: a discrete choice experiment to contrast noninvasive prenatal diagnosis with current invasive tests.
Genet Med. 2012 “]

Hill M, Fisher J, Chitty LS, Morris S.

1] Clinical and Molecular Genetics, Institute of Child Health and Great Ormond Street Hospital for Children NHS Foundation Trust, London, UK [2] Fetal Medicine Unit, University College London Hospitals NHS Foundation Trust, London, UK.

Abstract

Purpose:To compare the preferences of women and health professionals for key attributes of noninvasive prenatal diagnosis for Down syndrome relative to current invasive tests.Methods:A questionnaire incorporating a discrete choice experiment was used to obtain participants’ stated preference for diagnostic tests that varied according to four attributes: accuracy, time of test, risk of miscarriage, and provision of information about Down syndrome only or Down syndrome and other conditions. Women and health professionals were recruited from five maternity services in England and a patient support group.

Results:Questionnaires from 335 women and 181 health professionals were analyzed. Safe tests, conducted early in pregnancy, with high accuracy and information about Down syndrome and other conditions were preferred. The key attribute affecting women’s preferences for testing was no risk of miscarriage, whereas for health professionals it was accuracy.

Conclusions:Policies for implementing noninvasive prenatal diagnosis must consider the differences between women’s and health professionals’ preferences to ensure the needs of all stakeholders are met. Women’s strong preference for tests with no risk of miscarriage demonstrates that consideration for safety of the fetus is paramount in decision making. Effective pretest counseling is therefore essential to ensure women understand the possible implications of results

Genet Med 2012
[/toggle]

[toggle Title=”Tsaliki E etal.
MeDIP real-time qPCR of maternal peripheral blood reliably identifies trisomy 21.
Prenat Diagn 2012“]

Tsaliki E, Papageorgiou EA, Spyrou C, Koumbaris G, Kypri E, Kyriakou S, Sotiriou C, Touvana E, Keravnou A, Karagrigoriou A, Lamnissou K, Velissariou V, Patsalis PC.

NIPD Genetics Ltd, Nicosia, Cyprus; The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus; Department of Genetics and Biotechnology, Faculty of Biology, School of Science, National and Kapodistrian University of Athens, Panepistimiopolis, Ilissia, Athens, Greece.

Abstract

OBJECTIVE:
To reevaluate the efficiency of the 12 differentially methylated regions (DMRs) used in the methylated DNA immunoprecipitation (MeDIP) real-time quantitative polymerase chain reaction (real-time qPCR) based approach, develop an improved version of the diagnostic formula and perform a larger validation study.

METHODS:
Twelve selected DMRs were checked for copy number variants in the Database of Genomic Variants. The DMRs located within copy number variants were excluded from the analysis. One hundred and seventy-five maternal peripheral blood samples were used to reconstruct and evaluate the new diagnostic formula and for a larger-scale blinded validation study using MeDIP real-time qPCR.

RESULTS:
Seven DMRs entered the final model of the prediction equation and a larger blinded validation study demonstrated 100% sensitivity and 99.2% specificity. No significant evidence for association was observed between cell free fetal DNA concentration and D value.

CONCLUSION:
The MeDIP real-time qPCR method for noninvasive prenatal diagnosis of trisomy 21 was confirmed and revalidated in 175 samples with satisfactory results demonstrating that it is accurate and reproducible. We are currently working towards simplification of the method to make it more robust and therefore easily, accurately, and rapidly reproduced and adopted by other laboratories. Nevertheless, larger scale validation studies are necessary before the MeDIP real-time qPCR-based method could be applied in clinical practice.

Prenat Diagn. 2012 Jul 26:1-6. [Epub ahead of print]

[/toggle]

[toggle Title=”Bianchi DW
From prenatal genomic diagnosis to fetal personalized medicine: progress and challenges.
Nature Med 2012“]

Bianchi DW

The Mother Infant Research Institute at Tufts Medical Center and the Division of Genetics, Department of Pediatrics, Floating Hospital for Children, Boston, Massachusetts, USA.

Abstract
Thus far, the focus of personalized medicine has been the prevention and treatment of conditions that affect adults. Although advances in genetic technology have been applied more frequently to prenatal diagnosis than to fetal treatment, genetic and genomic information is beginning to influence pregnancy management. Recent developments in sequencing the fetal genome combined with progress in understanding fetal physiology using gene expression arrays indicate that we could have the technical capabilities to apply an individualized medicine approach to the fetus. Here I review recent advances in prenatal genetic diagnostics, the challenges associated with these new technologies and how the information derived from them can be used to advance fetal care. Historically, the goal of prenatal diagnosis has been to provide an informed choice to prospective parents. We are now at a point where that goal can and should be expanded to incorporate genetic, genomic and transcriptomic data to develop new approaches to fetal treatment.
AJOG June 2012. [Epub ahead of print]

Nat Med. 2012 Jul 6;18(7):1041-51.

[/toggle]

[toggle Title=” Lo YM
Non-invasive prenatal diagnosis by massively parallel sequencing of maternal plasma DNA.
Open Biology 2012“]

Lo YM

Li Ka Shing Institute of Health Sciences , The Chinese University of Hong Kong , Prince of Wales Hospital, Shatin, New Territories , Hong Kong SAR, People’s Republic of China.

Abstract
The presence of foetal DNA in the plasma of pregnant women has opened up new possibilities for non-invasive prenatal diagnosis. The use of circulating foetal DNA for the non-invasive prenatal detection of foetal chromosomal aneuploidies is challenging as foetal DNA represents a minor fraction of maternal plasma DNA. In 2007, it was shown that single molecule counting methods would allow the detection of the presence of a trisomic foetus, as long as enough molecules were counted. With the advent of massively parallel sequencing, millions or billions of DNA molecules can be readily counted. Using massively parallel sequencing, foetal trisomies 21, 13 and 18 have been detected from maternal plasma. Recently, large-scale clinical studies have validated the robustness of this approach for the prenatal detection of foetal chromosomal aneuploidies. A proof-of-concept study has also shown that a genome-wide genetic and mutational map of a foetus can be constructed from the maternal plasma DNA sequencing data. These developments suggest that the analysis of foetal DNA in maternal plasma would play an increasingly important role in future obstetrics practice. It is thus a priority that the ethical, social and legal issues regarding this technology be systematically studied.

Open Biol. 2012 Jun;2(6):120086.
[/toggle]

[toggle Title=”Norton ME etal.
Non-Invasive Chromosomal Evaluation (NICE) Study: results of a multicenter prospective cohort study for detection of fetal trisomy 21 and trisomy 18.
AJOG 2012“]

Norton ME, Brar H, Weiss J, Karimi A, Laurent LC, Caughey AB, Rodriguez MH, Williams J 3rd, Mitchell ME, Adair CD, Lee H, Jacobsson B, Tomlinson MW, Oepkes D, Hollemon D, Sparks AB, Oliphant A, Song K.

Department of Obstetrics and Gynecology, Stanford University/Lucile Packard Children’s Hospital, Stanford, CA.

Abstract
OBJECTIVE:
We sought to evaluate performance of a noninvasive prenatal test for fetal trisomy 21 (T21) and trisomy 18 (T18).
STUDY DESIGN:
A multicenter cohort study was performed whereby cell-free DNA from maternal plasma was analyzed. Chromosome-selective sequencing on chromosomes 21 and 18 was performed with reporting of an aneuploidy risk (High Risk or Low Risk) for each subject.
RESULTS:
Of the 81 T21 cases, all were classified as High Risk for T21 and there was 1 false-positive result among the 2888 normal cases, for a sensitivity of 100% (95% confidence interval [CI], 95.5-100%) and a false-positive rate of 0.03% (95% CI, 0.002-0.20%). Of the 38 T18 cases, 37 were classified as High Risk and there were 2 false-positive results among the 2888 normal cases, for a sensitivity of 97.4% (95% CI, 86.5-99.9%) and a false-positive rate of 0.07% (95% CI, 0.02-0.25%).
CONCLUSION:
Chromosome-selective sequencing of cell-free DNA and application of an individualized risk algorithm is effective in the detection of fetal T21 and T18.

AJOG June 2012. [Epub ahead of print]

[/toggle]

[toggle Title=”Bianchi DW etal.
Genome-Wide Fetal Aneuploidy Detection by Maternal Plasma DNA Sequencing
Obstet Gynecol 2012“]

Bianchi DW, Platt LD, Goldberg JD, Abuhamad AZ, Sehnert AJ, Rava RP.

From the Mother Infant Research Institute, Tufts Medical Center and Tufts University School of Medicine, Boston Massachusetts; the Center for Fetal Medicine and Women’s Ultrasound and the David Geffen School of Medicine, the University of California, Los Angeles, Los Angeles, California; the Prenatal Diagnosis Center, San Francisco Perinatal Associates, San Francisco, California; the Eastern Virginia Medical School, Norfolk, Virginia; and Verinata Health, Inc., Redwood City, California.

Abstract
OBJECTIVE:
To prospectively determine the diagnostic accuracy of massively parallel sequencing to detect whole chromosome fetal aneuploidy from maternal plasma.

METHODS:
Blood samples were collected in a prospective, blinded study from 2,882 women undergoing prenatal diagnostic procedures at 60 U.S. sites. An independent biostatistician selected all singleton pregnancies with any abnormal karyotype and a balanced number of randomly selected pregnancies with euploid karyotypes. Chromosome classifications were made for each sample by massively parallel sequencing and compared with fetal karyotype.

RESULTS:
Within an analysis cohort of 532 samples, the following were classified correctly: 89 of 89 trisomy 21 cases (sensitivity 100%, 95% [confidence interval] CI 95.9-100), 35 of 36 trisomy 18 cases (sensitivity 97.2%, 95% CI 85.5-99.9), 11 of 14 trisomy 13 cases (sensitivity 78.6%, 95% CI 49.2-99.9), 232 of 233 females (sensitivity 99.6%, 95% CI 97.6 to more than 99.9), 184 of 184 males (sensitivity 100%, 95% CI 98.0-100), and 15 of 16 monosomy X cases (sensitivity 93.8%, 95% CI 69.8-99.8). There were no false-positive results for autosomal aneuploidies (100% specificity, 95% CI more than 98.5 to 100). In addition, fetuses with mosaicism for trisomy 21 (3/3), trisomy 18 (1/1), and monosomy X (2/7), three cases of translocation trisomy, two cases of other autosomal trisomies (20 and 16), and other sex chromosome aneuploidies (XXX, XXY, and XYY) were classified correctly.

CONCLUSION:
This prospective study demonstrates the efficacy of massively parallel sequencing of maternal plasma DNA to detect fetal aneuploidy for multiple chromosomes across the genome. The high sensitivity and specificity for the detection of trisomies 21, 18, 13, and monosomy X suggest that massively parallel sequencing can be incorporated into existing aneuploidy screening algorithms to reduce unnecessary invasive procedures.

Obstet Gynecol. 2012 Feb 22. [Epub ahead of print]

[/toggle]

[toggle Title=”Chiu RW etal.
Noninvasive prenatal diagnosis of fetal chromosomal aneuploidy by massively parallel genomic sequencing of DNA in maternal plasma
PNAS 2008“]

Chiu RW, Chan KC, Gao Y, Lau VY, Zheng W, Leung TY, Foo CH, Xie B, Tsui NB, Lun FM, Zee BC, Lau TK, Cantor CR, Lo YM.

Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, Department of Chemical Pathology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China.

Abstract
Chromosomal aneuploidy is the major reason why couples opt for prenatal diagnosis. Current methods for definitive diagnosis rely on invasive procedures, such as chorionic villus sampling and amniocentesis, and are associated with a risk of fetal miscarriage. Fetal DNA has been found in maternal plasma but exists as a minor fraction among a high background of maternal DNA. Hence, quantitative perturbations caused by an aneuploid chromosome in the fetal genome to the overall representation of sequences from that chromosome in maternal plasma would be small. Even with highly precise single molecule counting methods such as digital PCR, a large number of DNA molecules and hence maternal plasma volume would need to be analyzed to achieve the necessary analytical precision. Here we reasoned that instead of using approaches that target specific gene loci, the use of a locus-independent method would greatly increase the number of target molecules from the aneuploid chromosome that could be analyzed within the same fixed volume of plasma. Hence, we used massively parallel genomic sequencing to quantify maternal plasma DNA sequences for the noninvasive prenatal detection of fetal trisomy 21. Twenty-eight first and second trimester maternal plasma samples were tested. All 14 trisomy 21 fetuses and 14 euploid fetuses were correctly identified. Massively parallel plasma DNA sequencing represents a new approach that is potentially applicable to all pregnancies for the noninvasive prenatal diagnosis of fetal chromosomal aneuploidies.

Proc Natl Acad Sci U S A. 2008 Dec 23;105(51):20458-63.

[/toggle]

[toggle Title=”Chiu RW etal.
Non-invasive prenatal assessment of trisomy 21 by multiplexed maternal plasma DNA sequencing: large scale validity
BMJ 2011“]

Chiu RW, Akolekar R, Zheng YW, Leung TY, Sun H, Chan KC, Lun FM, Go AT, Lau ET, To WW, Leung WC, Tang RY, Au-Yeung SK, Lam H, Kung YY, Zhang X, van Vugt JM, Minekawa R, Tang MH, Wang J, Oudejans CB, Lau TK, Nicolaides KH, Lo YM.

Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, Department of Chemical Pathology, The Chinese University of Hong Kong, Hong Kong SAR, China.

Abstract

OBJECTIVES:
To validate the clinical efficacy and practical feasibility of massively parallel maternal plasma DNA sequencing to screen for fetal trisomy 21 among high risk pregnancies clinically indicated for amniocentesis or chorionic villus sampling.

DESIGN:
Diagnostic accuracy validated against full karyotyping, using prospectively collected or archived maternal plasma samples.

SETTING:
Prenatal diagnostic units in Hong Kong, United Kingdom, and the Netherlands.

PARTICIPANTS:
753 pregnant women at high risk for fetal trisomy 21 who underwent definitive diagnosis by full karyotyping, of whom 86 had a fetus with trisomy 21. Intervention Multiplexed massively parallel sequencing of DNA molecules in maternal plasma according to two protocols with different levels of sample throughput: 2-plex and 8-plex sequencing.

MAIN OUTCOME MEASURES:
Proportion of DNA molecules that originated from chromosome 21. A trisomy 21 fetus was diagnosed when the z score for the proportion of chromosome 21 DNA molecules was >3. Diagnostic sensitivity, specificity, positive predictive value, and negative predictive value were calculated for trisomy 21 detection.

RESULTS:
Results were available from 753 pregnancies with the 8-plex sequencing protocol and from 314 pregnancies with the 2-plex protocol. The performance of the 2-plex protocol was superior to that of the 8-plex protocol. With the 2-plex protocol, trisomy 21 fetuses were detected at 100% sensitivity and 97.9% specificity, which resulted in a positive predictive value of 96.6% and negative predictive value of 100%. The 8-plex protocol detected 79.1% of the trisomy 21 fetuses and 98.9% specificity, giving a positive predictive value of 91.9% and negative predictive value of 96.9%.

CONCLUSION:
Multiplexed maternal plasma DNA sequencing analysis could be used to rule out fetal trisomy 21 among high risk pregnancies. If referrals for amniocentesis or chorionic villus sampling were based on the sequencing test results, about 98% of the invasive diagnostic procedures could be avoided.

BMJ. 2011 Jan 11;342:c7401.

[/toggle]

[toggle Title=”Ehrich M etal.
Noninvasive detection of fetal trisomy 21 by sequencing of DNA in maternal blood: a study in a clinical setting
AJOG 2011“]

Ehrich M, Deciu C, Zwiefelhofer T, Tynan JA, Cagasan L, Tim R, Lu V, McCullough R, McCarthy E, Nygren AO, Dean J, Tang L, Hutchison D, Lu T, Wang H, Angkachatchai V, Oeth P, Cantor CR, Bombard A, van den Boom D.

Sequenom Inc., 3595 John Hopkins Ct., San Diego, CA 92121, USA.

Abstract

OBJECTIVE:
We sought to evaluate a multiplexed massively parallel shotgun sequencing assay for noninvasive trisomy 21 detection using circulating cell-free fetal DNA.

STUDY DESIGN:
Sample multiplexing and cost-optimized reagents were evaluated as improvements to a noninvasive fetal trisomy 21 detection assay. A total of 480 plasma samples from high-risk pregnant women were employed.

RESULTS:
In all, 480 prospectively collected samples were obtained from our third-party storage site; 13 of these were removed due to insufficient quantity or quality. Eighteen samples failed prespecified assay quality control parameters. In all, 449 samples remained: 39 trisomy 21 samples were correctly classified; 1 sample was misclassified as trisomy 21. The overall classification showed 100% sensitivity (95% confidence interval, 89-100%) and 99.7% specificity (95% confidence interval, 98.5-99.9%).

CONCLUSION:
Extending the scope of previous reports, this study demonstrates that plasma DNA sequencing is a viable method for noninvasive detection of fetal trisomy 21 and warrants clinical validation in a larger multicenter study.

Am J Obstet Gynecol. 2011 Mar;204(3):205.e1-11

[/toggle]

[toggle Title=”Ghanta S etal.
Non-Invasive Prenatal Detection of Trisomy 21 Using Tandem Single Nucleotide Polymorphisms
PlosOne 2010“]

Ghanta S, Mitchell ME, Ames M, Hidestrand M, Simpson P, Goetsch M, Thilly WG, Struble CA, Tomita-Mitchell A.

Division of Cardiothoracic Surgery, Department of Surgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.

Abstract

BACKGROUND:
Screening tests for Trisomy 21 (T21), also known as Down syndrome, are routinely performed for the majority of pregnant women. However, current tests rely on either evaluating non-specific markers, which lead to false negative and false positive results, or on invasive tests, which while highly accurate, are expensive and carry a risk of fetal loss. We outline a novel, rapid, highly sensitive, and targeted approach to non-invasively detect fetal T21 using maternal plasma DNA.

METHODS AND FINDINGS:
Highly heterozygous tandem Single Nucleotide Polymorphism (SNP) sequences on chromosome 21 were analyzed using High-Fidelity PCR and Cycling Temperature Capillary Electrophoresis (CTCE). This approach was used to blindly analyze plasma DNA obtained from peripheral blood from 40 high risk pregnant women, in adherence to a Medical College of Wisconsin Institutional Review Board approved protocol. Tandem SNP sequences were informative when the mother was heterozygous and a third paternal haplotype was present, permitting a quantitative comparison between the maternally inherited haplotype and the paternally inherited haplotype to infer fetal chromosomal dosage by calculating a Haplotype Ratio (HR). 27 subjects were assessable; 13 subjects were not informative due to either low DNA yield or were not informative at the tandem SNP sequences examined. All results were confirmed by a procedure (amniocentesis/CVS) or at postnatal follow-up. Twenty subjects were identified as carrying a disomy 21 fetus (with two copies of chromosome 21) and seven subjects were identified as carrying a T21 fetus. The sensitivity and the specificity of the assay was 100% when HR values lying between 3/5 and 5/3 were used as a threshold for normal subjects.

CONCLUSIONS:
In summary, a targeted approach, based on calculation of Haplotype Ratios from tandem SNP sequences combined with a sensitive and quantitative DNA measurement technology can be used to accurately detect fetal T21 in maternal plasma when sufficient fetal DNA is present in maternal plasma.

PLoS One. 2010 Oct 8;5(10):e13184

[/toggle]

[toggle Title=”Palomaki GE etal.
DNA sequencing of maternal plasma to detect Down syndrome: an international clinical validation study
Genet Med. 2011“]

Palomaki GE, Kloza EM, Lambert-Messerlian GM, Haddow JE, Neveux LM, Ehrich M, van den Boom D, Bombard AT, Deciu C, Grody WW, Nelson SF, Canick JA.

Division of Medical Screening and Special Testing, Department of Pathology and Laboratory Medicine, Women & Infants Hospital, Alpert Medical School of Brown University, Providence, Rhode Island 02903, USA. gpalomaki@ipmms.org

Abstract

PURPOSE:
Prenatal screening for Down syndrome has improved, but the number of resulting invasive diagnostic procedures remains problematic. Measurement of circulating cell-free DNA in maternal plasma might offer improvement.

METHODS:
A blinded, nested case-control study was designed within a cohort of 4664 pregnancies at high risk for Down syndrome. Fetal karyotyping was compared with an internally validated, laboratory-developed test based on next-generation sequencing in 212 Down syndrome and 1484 matched euploid pregnancies. None had been previously tested. Primary testing occurred at a CLIA-certified commercial laboratory, with cross validation by a CLIA-certified university laboratory.

RESULTS:
Down syndrome detection rate was 98.6% (209/212), the false-positive rate was 0.20% (3/1471), and the testing failed in 13 pregnancies (0.8%); all were euploid. Before unblinding, the primary testing laboratory also reported multiple alternative interpretations. Adjusting chromosome 21 counts for guanine cytosine base content had the largest impact on improving performance.

CONCLUSION:
When applied to high-risk pregnancies, measuring maternal plasma DNA detects nearly all cases of Down syndrome at a very low false-positive rate. This method can substantially reduce the need for invasive diagnostic procedures and attendant procedure-related fetal losses. Although implementation issues need to be addressed, the evidence supports introducing this testing on a clinical basis.

Genet Med. 2011 Nov;13(11):913-20.
[/toggle]

[toggle Title=”Papageorgiou EA etal.
Fetal-specific DNA methylation ratio permits noninvasive prenatal diagnosis of trisomy 21
Nat Med. 2011“]

Papageorgiou EA, Karagrigoriou A, Tsaliki E, Velissariou V, Carter NP, Patsalis PC.

Cytogenetics and Genomics Department, The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus.

Abstract
The trials performed worldwide toward noninvasive prenatal diagnosis (NIPD) of Down’s syndrome (or trisomy 21) have shown the commercial and medical potential of NIPD compared to the currently used invasive prenatal diagnostic procedures. Extensive investigation of methylation differences between the mother and the fetus has led to the identification of differentially methylated regions (DMRs). In this study, we present a strategy using the methylated DNA immunoprecipitation (MeDiP) methodology in combination with real-time quantitative PCR (qPCR) to achieve fetal chromosome dosage assessment, which can be performed noninvasively through the analysis of fetal-specific DMRs. We achieved noninvasive prenatal detection of trisomy 21 by determining the methylation ratio of normal and trisomy 21 cases for each tested fetal-specific DMR present in maternal peripheral blood, followed by further statistical analysis. The application of this fetal-specific methylation ratio approach provided correct diagnosis of 14 trisomy 21 and 26 normal cases.

Nat Med. 2011 Apr;17(4):510-3
[/toggle]

[toggle Title=”Sparks AB etal.
Selective analysis of cell-free DNA in maternal blood for evaluation of fetal trisomy
Prenat Diagnosis 2012“]

Sparks AB, Wang ET, Struble CA, Barrett W, Stokowski R, McBride C, Zahn J, Lee K, Shen N, Doshi J, Sun M, Garrison J, Sandler J, Hollemon D, Pattee P, Tomita-Mitchell A, Mitchell M, Stuelpnagel J, Song K, Oliphant A.

Aria Diagnostics, Inc., 5945 Optical Court, San Jose, CA, 95138, USA.

Abstract

OBJECTIVE:
To develop a novel prenatal assay based on selective analysis of cell-free DNA in maternal blood for evaluation of fetal Trisomy 21 (T21) and Trisomy 18 (T18).

METHODS:
Two hundred ninety-eight pregnancies, including 39 T21 and seven T18 confirmed fetal aneuploidies, were analyzed using a novel, highly multiplexed assay, termed digital analysis of selected regions (DANSR™). Cell-free DNA from maternal blood samples was analyzed using DANSR assays for loci on chromosomes 21 and 18. Products from 96 separate patients were pooled and sequenced together. A standard Z-test of chromosomal proportions was used to distinguish aneuploid samples from average-risk pregnancy samples. DANSR aneuploidy discrimination was evaluated at various sequence depths.

RESULTS:
At the lowest sequencing depth, corresponding to 204 000 sequencing counts per sample, average-risk cases where distinguished from T21 and T18 cases, with Z statistics for all cases exceeding 3.6. Increasing the sequencing depth to 410 000 counts per sample substantially improved separation of aneuploid and average-risk cases. A further increase to 620 000 counts per sample resulted in only marginal improvement. This depth of sequencing represents less than 5% of that required by massively parallel shotgun sequencing approaches.

CONCLUSION:
Digital analysis of selected regions enables highly accurate, cost efficient, and scalable noninvasive fetal aneuploidy assessment.

Prenat Diagn. 2012 Jan;32(1):3-9
[/toggle]