High Risk For Fetal Aneuploidy Research Articles

Cell-Free DNA Testing: What Is the Reason Why High-Risk Women Choose It?

Objective: The aim of the study wasto describe the past medical history, sociodemographic, and pregnancy characteristics of women at high risk for aneuploidy and to determine which factors are related to her choice of cell-free DNA (cfDNA) testing instead of invasive diagnostic testing. Methods: We conducted a prospective descriptive study including pregnant women from the Western Barcelona public health area at high risk for fetal aneuploidy, defined as a trisomy 21 or 18 risk between 1/10 and 1/250 at the combined first-trimester or at the second-trimester biochemical screening. During 1 year (December 2018 to November 2019), these women were asked to fill in a confidential questionnaire about her past medical history, demographic and pregnancy characteristics, and her opinion about termination of the pregnancy after a counseling consultation with a maternal-fetal medicine specialist in which advantages and disadvantages of both testing methods, cfDNA or diagnostic testing, were discussed. Logistic regression analysis was used to determine which factors were related with cfDNA uptake. Results: During the study period, 82 pregnant women filled the questionnaire. The median maternal age was 39.6 years (interquartile range [IQR] 37.3–40.9 years), and 73 (89%) of them were 35 years or older. Forty-three (52%) women opted for cfDNA testing, while 39 (48%) chose invasive diagnosis. In a logistic regression analysis, the use of assisted reproductive techniques (OR 13.03; 95% CI: 1.47–115.56; p = 0.021) and Latin American origin (OR 6.66; 95% CI 1.73–25.66; p = 0.006) were shown to be related to a higher cfDNA uptake. In contrast, nonreligious women (OR 0.21; 95% CI: 0.06–0.72; p = 0.013) and a favorable opinion about termination of pregnancy (OR 0.23; 95% CI: 0.06–0.92; p = 0.037) were related with a lower uptake. Conclusion: Half of the pregnant women at high risk for fetal aneuploidy opted for cfDNA testing. The main reason to choose cfDNA was avoiding the risk of pregnancy loss. Women using assisted reproductive techniques and those of Latin American origin preferred cfDNA testing, while nonreligious women and those with a favorable opinion on termination pregnancy preferred invasive testing.

Использование трофобластов цервикального канала для неинвазивной пренатальной детекции частых анеуплоидий

The article presents an experience of isolation of trophoblast cells from cervical samples of pregnant women and demonstrates a possibility of determining aneuploid cells in cervical samples using FISH analysis. Objective. To study methods of effective isolation and reliable detection of trophoblasts for genetic pathology analysis. Patients and methods. The participants of the study were three pregnant women, who according to the findings of the combined first trimester screening test were referred to the group with a high risk for fetal aneuploidy. The terms of gestation varied from 12 to 14 wks. Immunocytochemical detection of trophoblast cells was performed with FITC-labelled HLA-G antibodies. FISH was performed with the use of locus-specific DNA probes on chromosomes 13/21 and a DNA probe on chromosome 18 centromere. The chromosome set of a fetus was verified by the results of standard cytogenetic analysis of semi-direct chromosome preparations from cytotrophoblast cells obtained by chorionic villi biopsy. Results. HLA-G-positive cells were found in all examined cytological specimens. FISH analysis with the use of DNA probes on chronomomes, whose trisomies are compatible with live birth, detected aneuploid cells in all three cases. Conclusion. The use of cervical trophoblasts for noninvasive collection of information about the genetic status of the developing fetus at present needs further study of effective and reliable methods of their isolation, detection and analysis. Key words: aneuploidy, throphoblast cells, NIPT, prenatal diagnosis, HLA-G

Prospective head-to-head comparison of accuracy of two sequencing platforms for screening for fetal aneuploidy by cell-free DNA: the PEGASUS study

We compared clinical validity of two non-invasive prenatal screening (NIPS) methods for fetal trisomies 13, 18, 21, and monosomy X. We recruited prospectively 2203 women at high risk of fetal aneuploidy and 1807 at baseline risk. Three-hundred and twenty-nine euploid samples were randomly removed. The remaining 1933 high risk and 1660 baseline-risk plasma aliquots were assigned randomly between four laboratories and tested with two index NIPS tests, blind to maternal variables and pregnancy outcomes. The two index tests used massively parallel shotgun sequencing (semiconductor-based and optical-based). The reference standard for all fetuses was invasive cytogenetic analysis or clinical examination at birth and postnatal follow-up. For each chromosome of interest, chromosomal ratios were calculated (number of reads for chromosome/total number of reads). Euploid samples’ mean chromosomal ratio coefficients of variation were 0.48 (T21), 0.34 (T18), and 0.31 (T13). According to the reference standard, there were 155 cases of T21, 49 T18, 8 T13 and 22 45,X. Using a fetal fraction ≥4% to call results and a chromosomal ratio z-score of ≥3 to report a positive result, detection rates (DR), and false positive rates (FPR) were not statistically different between platforms: mean DR 99% (T21), 100%(T18, T13); 79%(45,X); FPR < 0.3% for T21, T18, T13, and <0.6% for 45,X. Both methods’ negative predictive values in high-risk pregnancies were >99.8%, except for 45,X(>99.6%). Threshold analysis in high-risk pregnancies with different fetal fractions and z-score cut-offs suggested that a z-score cutoff to 3.5 for positive results improved test accuracy. Both sequencing platforms showed equivalent and excellent clinical validity.

An adaptive detection method for fetal chromosomal aneuploidy using cell-free DNA from 447 Korean women.

BackgroundNoninvasive prenatal testing (NIPT) using massively parallel sequencing of cell-free DNA (cfDNA) is increasingly being used to predict fetal chromosomal abnormalities. However, concerns over erroneous predictions which occur while performing NIPT still exist in pregnant women at high risk for fetal aneuploidy. We performed the largest-scale clinical NIPT study in Korea to date to assess the risk of false negatives and false positives using next-generation sequencing.MethodsA total of 447 pregnant women at high risk for fetal aneuploidy were enrolled at 12 hospitals in Korea. They underwent definitive diagnoses by full karyotyping by blind analysis and received aneuploidy screening at 11–22 weeks of gestation. Three steps were employed for cfDNA analyses. First, cfDNA was sequenced. Second, the effect of GC bias was corrected using normalization of samples as well as LOESS and linear regressions. Finally, statistical analysis was performed after selecting a set of reference samples optimally adapted to a test sample from the whole reference samples. We evaluated our approach by performing cfDNA testing to assess the risk of trisomies 13, 18, and 21 using the sets of extracted reference samples.ResultsThe adaptive selection algorithm presented here was used to choose a more optimized reference sample, which was evaluated by the coefficient of variation (CV), demonstrated a lower CV and higher sensitivity than standard approaches. Our adaptive approach also showed that fetal aneuploidies could be detected correctly by clearly splitting the z scores obtained for positive and negative samples.ConclusionsWe show that our adaptive reference selection algorithm for optimizing trisomy detection showed improved reliability and will further support practitioners in reducing both false negative and positive results.Electronic supplementary materialThe online version of this article (doi:10.1186/s12920-016-0222-5) contains supplementary material, which is available to authorized users.

Noninvasive prenatal testing in routine clinical practice for a high-risk population

This study aimed to summarize the effects of noninvasive prenatal testing (NIPT) on aneuploidy among high-risk participants in Tangshan Maternal and Children Health Hospital.NIPT or invasive prenatal diagnosis was recommended to patients with a high risk of fetal aneuploidy from February 2013 to February 2014. Patients who exhibited eligibility and applied for NIPT from January 2012 to January 2013 were included in a comparison group. The rates of patients who underwent invasive testing, declined to undergo further testing, and manifested trisomies 21, 18, and 13 were compared between two groups. Follow-up data were obtained from the participants who underwent NIPT from 2013 to 2014.A total of 7223 patients (3018 and 4205 individuals before and after NIPT) were eligible for analysis. After NIPT was introduced in 2013 to 2014, 727 patients (17.3%) underwent invasive testing, 2828 preferred NIPT (67.3%), and 650 declined to undergo further testing (15.5%). A total of 34 cases of trisomies 21, 18, and 13 (0.8%) were found. In 2012 to 2013, 565 patients (18.7%) underwent invasive testing and 2453 declined to undergo further testing (81.3%). A total of 7 cases of trisomies 21, 18, and 13 were documented (0.2%). Of these cases, 24 were found from NIPT and 10 cases were found from invasive testing. The number of participants who declined to undergo further testing significantly decreased after NIPT was introduced (81.3% vs. 15.5%, P < 0.001). The sensitivity and specificity of NIPT for trisomies 21, 18, and 13 were 100% and 99.9%, respectively. The detection rates of NIPT for trisomies 21, 18, and 13 also significantly increased (0.2% vs. 0.8%, P < 0.001). By contrast, the overall rates of invasive testing remained unchanged (18.7% vs. 17.3%, P = 0.12). The positive predictive values of NIPT for trisomies 21, 18, and 13 were 100%, 83.3%, and 50.0%, respectively. The false positive rates of NIPT were 0% and 0.04%.With NIPT implementation in clinical practice, the rate of declining a follow-up test among high-risk women was decreased and the detection rate of prenatal chromosomal aneuploidy for trisomies 21, 18, and 13 was increased without requiring numerous invasive procedures.

The Integration of Noninvasive Prenatal Screening into the Existing Prenatal Paradigm: a Survey of Current Genetic Counseling Practice.

Since its introduction four years ago, noninvasive prenatal screening for fetal aneuploidy (NIPS) has been widely adopted as a screening tool for women at a high risk for fetal aneuploidy. As use expands into the general population, questions arise concerning the integration of NIPS into preexisting screening paradigms. This study aims to examine the use of NIPS in current practice among prenatal counselors, predominantly in the United States, in order to inform strategies for the optimal use of both new and existing screening techniques. We electronically surveyed 208 members of the National Society of Genetic Counselors to ascertain how NIPS is currently being used. Genetic counselors were also queried as to the advantages and disadvantages of offering NIPS to all patients regardless of a priori risk. Results indicate substantial variation in practice regarding which patients are offered NIPS and how counselors have incorporated this technology into existing screening routines. The majority of participants report offering NIPS in conjunction with another method of screening for fetal aneuploidy, indicating that NIPS is being used as an addition rather than as a replacement. These screening methods primarily include nuchal translucency (NT) (45.1%, n=78) and first trimester serum screening, with or without an NT (19.7%, n=34). Furthermore, the majority report that they would be concerned about losing the clinical value of an NT in a complete transition to NIPS (85.4%, n=164). Counselors are evenly split on the merits of expanding the use of NIPS to the general population (con: 55.3%, n=105; pro: 44.7%, n=85). The lack of consensus suggests that updated practice guidelines might benefit counselors. In addition, respondents emphasized the need to better educate patients and providers about the risks, benefits, and limitations of the test.

Advantages of the single nucleotide polymorphism-based noninvasive prenatal test

the placenta into the maternal circulation as the cells break down (apoptosis) and the DNA becomes fragmented [4,5]. Circulating fetal cfDNA comprises approximately 3-13% of the total maternal cfDNA. These DNA fragments can be detected as early as 4 weeks of gestation and are generally cleared out within 2 hours after childbirth [6-8]. A noninvasive prenatal test (NIPT) using cfDNA has proven to be highly sensitive and specific for detecting trisomy 21 in both highand low-risk groups [1,2,9,10]. At present, there are about five major companies providing commercial NIPT services globally (Fig. 1). NIPT using cfDNA is now available through these providers in more than 60 countries. According to business reports, over 500,000 NIPT studies on women with a high risk for fetal aneuploidy were performed in the United States in 2013 [11]. These companies have published several articles showing that their own method is very effective for screening for trisomy 21, 18, and 13 Advantages of the single nucleotide polymorphismbased noninvasive prenatal test

Oral Abstracts of the ISPD 19th International Conference on Prenatal Diagnosis and Therapy, Washington, DC, USA, 12-15 July 2015.

OBJECTIVES: The TRIDENT study (Trial by Dutch laboratories for Evaluation of Non-Invasive Prenatal Testing) evaluates the implementation of non-invasive prenatal testing (NIPT) in the Dutch healthcare system. Here we report on the preferences and experiences of pregnant women at high risk for fetal aneuploidy who were offered a choice between NIPT, invasive testing or no follow-up test. METHODS: A nationwide prospective cohort study among pregnant women at high risk for fetal aneuploidy because of first trimester screening results (risk ≥1:200) or medical history. Questionnaires were administered to the women, to be completed after counselling and after receiving the test-results, at seven (of the eight) prenatal diagnostic centers in the Netherlands. The questionnaires addressed: women's preference for follow-up testing, intention to terminate the pregnancy, attitudes, knowledge, decisional conflict, anxiety and satisfaction. RESULTS: Pre- and post-test questionnaires were completed by n = 1,106 (86% response) and n = 686 (67% response) women, respectively. Preliminary analyses show that the majority of respondents (92%) preferred NIPT, 5% invasive testing, 2% were unsure, and 1% declined testing. The main reason to prefer NIPT was safety for the child (92%). Of the 60 women preferring invasive testing, 52% did so because of test-accuracy, 25% desired more rapid test-results, 5% because it provides more information, and 18% reported other reasons. Most women (92%) felt that they made a well-informed decision, and 75% reported that this decision was easy to make. Intention to terminate the pregnancy for Down syndrome was lower among women preferring NIPT (58%) compared to women preferring invasive testing (87%). Women were highly satisfied with NIPT. However, 64% perceived the waiting time for NIPT results (mean: 11 days (range 5-32)) as too long, while 3% in retrospect would have preferred a different follow-up test, mostly to avoid the long waiting time. CONCLUSIONS: Most pregnant women in our study felt they could make an informed decision. The majority of women prefer NIPT, mainly because it has no miscarriage risk. Reducing turnaround time for test-results is our next challenge in meeting women's needs.

Noninvasive prenatal screening for fetal trisomies 21, 18, 13 and the common sex chromosome aneuploidies from maternal blood using massively parallel genomic sequencing of DNA

The objective of this study was to validate the clinical performance of massively parallel genomic sequencing of cell-free deoxyribonucleic acid contained in specimens from pregnant women at high risk for fetal aneuploidy to test fetuses for trisomies 21, 18, and 13; fetal sex; and the common sex chromosome aneuploidies (45, X; 47, XXX; 47, XXY; 47, XYY). This was a prospective multicenter observational study of pregnant women at high risk for fetal aneuploidy who had made the decision to pursue invasive testing for prenatal diagnosis. Massively parallel single-read multiplexed sequencing of cell-free deoxyribonucleic acid was performed in maternal blood for aneuploidy detection. Data analysis was completed using sequence reads unique to the chromosomes of interest. A total of 3430 patients were analyzed for demographic characteristics and medical history. There were 137 fetuses with trisomy 21, 39 with trisomy 18, and 16 with trisomy 13 for a prevalence rate of the common autosomal trisomies of 5.8%. There were no false-negative results for trisomy 21, 3 for trisomy 18, and 2 for trisomy 13; all 3 false-positive results were for trisomy 21. The positive predictive values for trisomies 18 and 13 were 100% and 97.9% for trisomy 21. A total of 8.6% of the pregnancies were 21 weeks or beyond; there were no aneuploid fetuses in this group. All 15 of the common sex chromosome aneuploidies in this population were identified, although there were 11 false-positive results for 45,X. Taken together, the positive predictive value for the sex chromosome aneuploidies was 48.4% and the negative predictive value was 100%. Our prospective study demonstrates that noninvasive prenatal analysis of cell-free deoxyribonucleic acid from maternal plasma is an accurate advanced screening test with extremely high sensitivity and specificity for trisomy 21 (>99%) but with less sensitivity for trisomies 18 and 13. Despite high sensitivity, there was modest positive predictive value for the small number of common sex chromosome aneuploidies because of their very low prevalence rate.

Integration of Noninvasive DNA Testing for Aneuploidy into Prenatal Care: What Has Happened Since the Rubber Met the Road?

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. 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. 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.

Current controversies in prenatal diagnosis 1: should noninvasive DNA testing be the standard screening test for Down syndrome in all pregnant women?

What's already known about this topic? Noninvasive DNA testing (NIDT) is recommended in many developed countries as an option for pregnant women who have already been determined to be at high risk for fetal aneuploidy. Over 2 years of clinical experience has been accumulated with offering NIDT as an advanced screen for fetal autosomal aneuploidy. At the present time, most testing is being performed by commercial organizations. What does this study add? This study provides a written transcript to accompany an oral debate that was presented at the 17th International Conference on Prenatal Diagnosis and Therapy in Lisbon, Portugal, on 3 June 2013. The debaters, who are both experts in maternal–fetal medicine, consider the benefits and limitations of offering NIDT to all pregnant women regardless of their a priori risk of having a fetus with a chromosome abnormality.

Slovenian Five-Year Experiences with Rapid Prenatal Diagnosis of Common Chromosome Aneuploidies Using Quantitative-Fluorescence Polymerase Chain Reaction

Quantitative-fluorescence polymerase chain reaction (QF-PCR) was used to detect common fetal aneuploidies in pregnancies with increased (maternal age) or high risk (increased nuchal translucency, abnormal fetal ultrasonography, positive biochemical hormone test, or positive family history) for fetal aneuploidy. The QF-PCR testing was performed on 642 prenatal samples (73.3% amniotic fluids, 26.7% chorionic villus). DNA from prenatal samples were analyzed using an in-house-developed QF-PCR method with 20 micro-satellite markers located on the chromosomes 13, 18, 21, X and Y. Karyotyping of the 392 samples was done and both results were compared. 634/642 samples were successfully analyzed. In 7.1% of 634 cases numerical chromosome abnormalities were detected. Results of QF-PCR and karyotyping were compared in 392 cases. In the group, with increased risk of fetal trisomy the specificity and sensitivity of QF-PCR method was 100%. Among cases with high risk for fetal aneuploidy, sensitivity was 100% (86.6%-100%); however, the specificity was lower, 91.1% to 100%, depending on the referral reason. In women, at advanced age QF-PCR can be used alone without karyotyping. In cases with higher risk, especially those with abnormal ultrasound findings, analysis performed only with QF-PCR is not a sufficient diagnostic method.

Noninvasive Prenatal Testing for Fetal Aneuploidy

The recent introduction of clinical tests to detect fetal aneuploidy by analysis of cell-free DNA in maternal plasma represents a tremendous advance in prenatal diagnosis and the culmination of many years of effort by researchers in the field. The development of noninvasive prenatal testing for clinical application by commercial industry has allowed much faster introduction into clinical care, yet also presents some challenges regarding education of patients and health care providers struggling to keep up with developments in this rapidly evolving area. It is important that health care providers recognize that the test is not diagnostic; rather, it represents a highly sensitive and specific screening test that should be expected to result in some false-positive and false-negative diagnoses. Although currently being integrated in some settings as a primary screening test for women at high risk of fetal aneuploidy, from a population perspective, a better option for noninvasive prenatal testing may be as a second-tier test for those patients who screen positive by conventional aneuploidy screening. How noninvasive prenatal testing will ultimately fit with the current prenatal testing algorithms remains to be determined. True cost-utility analyses will be needed to determine the actual clinical efficacy of this approach in the general prenatal population.

Non‐invasive prenatal testing for fetal aneuploidy: charting the course from clinical validity to clinical utility

Non‐invasive prenatal testing for fetal aneuploidy: charting the course from clinical validity to clinical utility

Rapid-Fluorescence-in-Situ-Hybridization (FISH) on Uncultured Amniocytes for Avoiding Birth Defects due to Common Chromosomal Aberrations

Background Birth defects are structural or functional abnormalities present at birth that can cause physical or mental disability in new borns. Birth defects can be due to genetic problems caused by mutation in one or more genes, chromosome aneuploidy or environmental factors in women exposed during pregnancy. Objective The aim of the study was to evaluate the effectiveness of the Rapid-Fluorescence-in-Situ-Hybridization (Rapid-FISH) technique in detecting numerical chromosome aberrations of 13, 21, 18, X and Y in amniocyte nuclei from amniotic fluid for avoiding birth defects due to chromosome abnormalities performed on 138 high risk pregnancies. Methods The FISH was performed using AneuVysion kit (Vysis, Inc), according to a standard protocol. Results A total of one hundred thirty eight samples were received for full karyotype and FISH. The average age of mothers and their gestational ages were 33 years and 17.5 weeks respectively. Triple test screening was positive in 52.9% of the women followed by advanced maternal age and ultrasonographic abnormalities. Interphase FISH was performed on seventy eight specimens. All normal and abnormal results were confirmed by classical cytogenetic method (GTG banding and karyotyping). The overall detection rate for aneuploidies through FISH was 100%. Aneuploidy was identified in 5 out of 78 specimens (four cases of trisomy 21 and one case of Turner syndrome) taken for FISH. It was concluded that Rapid-FISH is a reliable and fast method for detecting numerical chromosomal aberrations through prenatal diagnosis and has been implemented as a routine diagnostic procedure in pregnancies with high risk of fetal aneuploidy for avoiding chromosomes 13, 18, 21, X, Y related birth defects.