Non-invasive Prenatal Genetic Testing Research Articles

Survey of attitudes of Chinese perinatologists and obstetricians toward non-invasive prenatal genetic testing.

The clinical application of non-invasive prenatal testing (NIPT) is still very limited in China. We carried out a survey to assess the willingness of Chinese obstetricians to offer NIPT and to determine how they would implement it and what resources they would need for the testing. Between June 2014 and June 2015, a survey was conducted at a large academic referral center with data obtained from 392 registered perinatologists and obstetricians who completed an entire questionnaire. Most respondents (72.5%) agreed or strongly agreed that the percentage of women patients refusing to accept NIPT would increase if they were charged directly for the test. Most respondents (82.7%) answered affirmatively that the national health administration agencies should formulate a standard charge for NIPT. The most important factors that influence the application of NIPT are the popularity of the test and its cost. The majority of respondents indicated that there are appropriate reasons for NIPT. The importance of NIPT and guidelines for the application of NIPT should be clarified in current clinical practice in China. Extensive education regarding NIPT application is necessary prior to mass implementation.

Expanded indications for noninvasive prenatal genetic testing: Implications for the individual and the public

Summary Objectives Noninvasive prenatal testing (NIPT) has rapidly been integrated into the routine management of women at increased risk for common autosomal aneuploidies. Advances in molecular diagnostics have quickly made it possible to use NIPT to screen for other genetic variants not formerly included in routine prenatal genetic screening or diagnostic testing, including sex chromosome aneuploidies (SCAs) and microdeletion syndromes. Moreover, it is anticipated that NIPT will become available for use by the low-risk obstetric population in the near future as test performance becomes better defined with clinical experience. The pace and trajectory of these advances raise important questions about the clinical, ethical, and public health impact of this new prenatal technology for individual patients, families, and society. Methods A cross-sectional study was conducted of pregnant women who had utilized NIPT to examine key patient-reported aspects of the educational and decision-making process regarding current and forthcoming indications of this screening test. Results A total of 139 participants completed the survey. Two-thirds (n = 90, 65.2%) entered their prenatal care with the intention to undergo prenatal genetic screening or testing primarily because of self-identification as advanced maternal age (AMA); these participants were more likely to have had prenatal genetic assessment in a prior pregnancy (P = 0.016). The remainder proceeded with NIPT following a directed discussion with their clinician about testing options. Leading decision-making factors in favor of NIPT included the absence of iatrogenic risk, the increased accuracy of NIPT as compared to conventional screening to detect trisomy (T) 21, T13, T18, and SCAs, and the ability to determine fetal sex. Participants had greater familiarity with T21, T13, and T18 than with SCAs and overall less familiarity with the microdeletions that NIPT is currently being marketed to detect. While participants recognized the superior accuracy of NIPT over conventional screens, few recognized the different detection and false positive rates within a screening panel for each of the identifiable conditions. Despite their awareness of the increased accuracy of NIPT, 41.3% (n = 57) described the testing process as stressful, 64.7% (n = 90) recommended a formal, written informed consent process in addition to 50.0% (n = 68) genetic counseling prior to screening. Conclusions Pregnant women are less familiar with SCAs, microdeletions, and the capability of NIPT to screen for these variants than they are with T21, T18, and T13. Additionally, despite the increased accuracy and capability of NIPT as compared to conventional screening tests, women reported feeling anxiety during the screening process. The results of this study highlight the need for clinicians and policymakers to provide effective guidance and support for patients’ decision-making about NIPT both at the present time and as the indications expand to include all pregnant populations and detailed fetal genomic analysis.

Online direct-to-consumer messages about non-invasive prenatal genetic testing.

Non-invasive prenatal testing (NIPT) has been integrated into clinical care at a time when patients and healthcare providers increasingly utilize the internet to access health information. This study evaluated online direct-to-consumer information about NIPT produced by commercial laboratories accessible to both patients and healthcare providers. A coding checklist captured areas to describe content and assess concordance with clinical guidelines. We found that the information presented about NIPT is highly variable, both within a single website and broadly across all websites. Variability was noted in how NIPT is characterized, including test characteristics and indications. All laboratories offer NIPT to test for common sex chromosome aneuploidies, although there is a lack of consistency regarding the conditions offered and information provided about each. Although indicated for a subset of women at increased risk of aneuploidy, some laboratories describe the use of NIPT for all pregnant women. A subset of laboratories offers screening for microdeletions, although clinical practice guidelines do not yet recommend for general use for this indication. None of the online materials addressed the ethical issues associated with NIPT. This study highlights the need for clear, consistent, and evidence-based materials to educate patients and healthcare providers about the current and emerging applications of NIPT.

Noninvasive Prenatal Genetic Testing: Current and Emerging Ethical, Legal, and Social Issues.

Noninvasive prenatal genetic testing (NIPT) for chromosomal aneuploidy involving the analysis of cell-free fetal DNA became commercially available in 2011. The low false-positive rate of NIPT, which reduces unnecessary prenatal invasive diagnostic procedures, has led to broad clinician and patient adoption. We discuss the ethical, legal, and social issues raised by rapid and global dissemination of NIPT. The number of women using NIPT is anticipated to expand, and the number of conditions being tested for will continue to increase as well, raising concerns about the routinization of testing and negative impacts on informed decision making. Ensuring that accurate and balanced information is available to all pregnant women and that access to NIPT is equitable will require policy guidance from regulators, professional societies, and payers. Empirical evidence about stakeholders' perspectives and experiences will continue to be essential in guiding policy development so that advances in NIPT can be used effectively and appropriately to improve prenatal care.

Novel options in Prenatal Genetic Diagnostic

Modern molecular approaches to prenatal diagnostic of inherited diseases are briefly reviewed. Advantages and limitations of molecular methods for analysis of chromosomal anomalies (QF-PCR, aCGH, NGS) are considered in line with conventional prenatal karyotyping. The special attention is paid to efficacy, limitations and diagnostic options of noninvasive prenatal genetic testing (NIPT). Some particular problems of its widespread implication into routine clinical practice are discussed. State of art in preimplantation genetic diagnostics (PGD) and obvious great opportunities of preconceptional genetic testing are highlighted.

Nationwide demonstration project of next-generation sequencing of cell-free DNA in maternal plasma in Japan: 1-year experience.

To report the 1-year experience of a nationwide demonstration project to introduce noninvasive prenatal testing of aneuploidy from maternal plasma and discuss how to implement this program in Japan. The test was conducted to detect aneuploidy in high-risk pregnant women with adequate genetic counseling. The clinical data, test results, and pregnancy outcomes were recorded. Of the 7740 women tested, 142 (1.8%) had positive results, 7594 (98.1%) had negative results, and four (0.1%) had results that were not reportable. Of the 142 women who tested positive, 13 cases resulted in intrauterine fetal death, and three cases refused to undergo the invasive procedure. Of the 126 positive cases confirmed on karyotyping, a normal karyotype was observed for trisomy 21 in three cases, trisomy 18 in eight cases, and trisomy 13 in two cases. In the follow-up study of the negative cases (n = 1638), only one false-negative case of trisomy 18 was detected. We described our nationwide 1-year experience with noninvasive prenatal genetic testing. It is expected that the present data will stimulate a debate regarding prenatal genetic testing and hopefully lead to improvements in the perinatal care system with respect to genetic counseling in Japan. © 2014 John Wiley & Sons, Ltd.

Non-invasive prenatal testing: a review of international implementation and challenges.

Noninvasive prenatal genetic testing (NIPT) is an advance in the detection of fetal chromosomal aneuploidies that analyzes cell-free fetal DNA in the blood of a pregnant woman. Since its introduction to clinical practice in Hong Kong in 2011, NIPT has quickly spread across the globe. While many professional societies currently recommend that NIPT be used as a screening method, not a diagnostic test, its high sensitivity (true positive rate) and specificity (true negative rate) make it an attractive alternative to the serum screens and invasive tests currently in use. Professional societies also recommend that NIPT be accompanied by genetic counseling so that families can make informed reproductive choices. If NIPT becomes more widely adopted, States will have to implement regulation and oversight to ensure it fits into existing legal frameworks, with particular attention to returning fetal sex information in areas where sex-based abortions are prevalent. Although there are additional challenges for NIPT uptake in the developing world, including the lack of health care professionals and infrastructure, the use of NIPT in low-resource settings could potentially reduce the need for skilled clinicians who perform invasive testing. Future advances in NIPT technology promise to expand the range of conditions that can be detected, including single gene disorders. With these advances come questions of how to handle incidental findings and variants of unknown significance. Moving forward, it is essential that all stakeholders have a voice in crafting policies to ensure the ethical and equitable use of NIPT across the world.

Next generation sequencing in clinical diagnostics: experiences of early adopters.

Next generation sequencing (NGS)13 technology, also known as massively parallel sequencing (MPS), is being incorporated rapidly to clinical laboratory testing. Current applications include detection of germline variants in inherited diseases, somatic variants in cancers, subpopulations of circulating cell-free DNA, and single viral or microbial genomes in infections or metamicrobial genomes in normal or altered human flora. Each application is unique and has its advantages and disadvantages. The complexity of the NGS processes (specimen preparation and testing and analysis of large amounts of data) leads to challenges in validation, QC, and data interpretation beyond what clinical laboratories have previously encountered. In this QA Richard Press, Carol Beadling, and Shashikant Kulkarni (oncology); Lynn Bry (infectious diseases); and Glenn Palomaki (noninvasive prenatal genetic testing of maternal plasma). The experiences and opinions they share demonstrate the diversity of the challenges and provide examples of how they are being addressed. Validation: Analytical validation measures the performance characteristics of a clinical test for accuracy and reproducibility (precision). In genomic sequencing, analytical sensitivity describes the ability of the assay to detect genetic variants when present, and specificity defines the capacity to correctly identify a “normal” (wild-type) sequence. Although genomic sequencing technology is advancing, regions of the genome remain problematic. 1: What has been the most difficult aspect of validating an assay using MPS? How has the laboratory been able to meet these requirements? Sherri Bale: The major problem in validating an assay using MPS is that it is an iterative process that involves optimizing 3 components at once—the …

The feasibility study of Ion Proton semiconductor sequencing platform in the non-invasive prenatal genetic diagnosis

Objective To evaluate the feasibility of apply Ion Proton semiconductor sequencing platform in non-invasive prenatal genetic diagnosis. Methods Totally 1 000 pregnant women with a singleton pregnancy of 12-32 weeks gestation were selected from the Third affiliated Hospital of Zhengzhou University from Jan to Dec 2013. Using noninvasive prenatal genetic diagnosis based on Ion Proton semiconductor sequencing platform to study their cffDNA. In parallel, 72 pregnant women received invasive prenatal diagnosis by traditional chromosomal analysis with amniocentesis chorionic villus sampling. Results It′s shown that 18 out of 1 000 (1.8%) pregnant women underwent the noninvasive prenatal genetic testing had a high risk for aneuploid chromosomes, including 7 cases of 21-trisomy, 4 cases of 18-trisomy, 2 cases of 13-trisomy, 4 cases of sex chromosomal abnormality, and 1 case of 15-trisomy. It demonstrated that the rate and accuracy of fetal 21-trisomy, 13-trisomy and 18-trisomy by non-invasive prenatal genetic testing were both 100% without misdiagnosis, the rate of detection for sex chromosomal abnormality was 2/2 with a false positive rate of 1/3. However, the 15-trisomy predicted by the non-invasive prenatal diagnosis in a woman was finally proved to be a false positive. Based on the results by karyotyping (55/55) as well as follow-ups (493/493), the specificity of the non-invasive prenatal diagnosis for detection of 21-trisomy, 18-trisomy and 13-trisomy was 100%. One Ion PITM chip could detect 12 to 15 samples in 1.5 h and the whole process of noninvasive detection could be completed in 1 to 1.5 days. Conclusions The non-invasive prenatal diagnosis by Ion Proton semiconductor sequencing platform could provide fast and accurate detection of fetal aneuploidy. The benchtop high-throughput sequencing platform has laid the foundation for the independent application in clinical settings for fetal aneuploidy detection. （Chin J Lab Med，2014,37：861-865） Key words: Prenatal diagnosis; Aneuploidy; Chromosome aberrations; Aneuploidy; Sequence analysis, DNA; High-throughput nucleotide sequencing; Feasibility studies

Use of copy number deletion polymorphisms to assess DNA chimerism.

We describe a novel approach that harnesses the ubiquity of copy number deletion polymorphisms in human genomes to definitively detect and quantify chimeric DNA in clinical samples. Unlike other molecular approaches to chimerism analysis, the copy number deletion (CND) method targets genomic loci (>50 base pairs in length) that are wholly absent from wild-type (i.e., self) background DNA sequences in a sex-independent manner. Bespoke quantitative PCR (qPCR) CND assays were developed and validated using a series of DNA standards and chimeric plasma DNA samples collected from 2 allogeneic kidney transplant recipients and 12 pregnant women. Assay performance and informativeness were assessed using appropriate statistical methods. The CND qPCR assays showed high sensitivity, precision, and reliability for linear quantification of DNA chimerism down to 16 genomic equivalents (i.e., 106 pg). Fetal fraction (%) in 12 singleton male pregnancies was calculated using the CND qPCR approach, which showed closer agreement with single-nucleotide polymorphism-based massively parallel sequencing than the SRY (sex determining region Y) (Y chromosome) qPCR assay. The latter consistently underestimated the fetal fraction relative to the other methods. We also were able to measure biological changes in plasma nonself DNA concentrations in 2 renal transplant recipients. The CND qPCR technique is suitable for measurement of chimerism for monitoring of rejection in allogeneic organ transplantation and quantification of the cell-free fetal DNA fraction in maternal plasma samples used for noninvasive prenatal genetic testing.

Noninvasive Prenatal Testing Goes Global

Noninvasive prenatal genetic testing is becoming available worldwide--particularly in low- and middle-income countries--but practical and ethical challenges must be overcome.

Ethical, Legal, Social, and Policy Implications of Behavioral Genetics

The field of behavioral genetics has engendered a host of moral and social concerns virtually since its inception. The policy implications of a genetic basis for behaviors are widespread and extend beyond the clinic to the socially important realms of education, criminal justice, childbearing, and child rearing. The development of new techniques and analytic approaches, including whole-genome sequencing, noninvasive prenatal genetic testing, and optogenetics, has clearly changed the study of behavioral genetics. However, the social context of biomedical research has also changed profoundly over the past few decades, and in ways that are especially relevant to behavioral genetics. The ever-widening scope of behavioral genetics raises ethical, legal, social, and policy issues in the potential new applications to criminal justice, education, the military, and reproduction. These issues are especially critical to address because of their potentially disproportionate effects on vulnerable populations such as children, the unborn, and the incarcerated.

Introduction

Introduction

Non-Invasive Prenatal Genetic Testing: A Study of Public Attitudes

Background/Objectives: Non-invasive prenatal genetic diagnostic (NIPD) tests are being developed using cell-free fetal DNA in the maternal circulation. NIPD tests avoid or reduce the need for invasive diagnostic procedures for conditions like Down syndrome. Discussion of ethical and social implications of these techniques is increasing. We report findings from a study of public attitudes relevant to the introduction of NIPD. A key aim was to examine the range of attitudes relevant to NIPD within a diverse sample. Methods: Qualitative analysis of written free text ‘first responses’ to a written neutral description of NIPD as part of a Q-methodology study conducted with a purposive sample of the UK population (n = 71). Results: The majority (63%) of respondents described their first response as positive. However, respondents displayed ambivalence, expressing positive views of individual/medical rationale for NIPD and unease concerning public health rationale and societal implications. Unease related to eugenic reasoning underlying existing prenatal testing, ‘too much control’ in reproduction, commercial provision, information and support requirements for expanded testing, and limiting the use of testing. Conclusions: These findings suggest that regulating and monitoring commercial provision of NIPD services, and monitoring introduction and clinical use, are a public preference.

Cell-Free Fetal Nucleic Acid Testing: A Review of the Technology and Its Applications

Cell-free fetal nucleic acids circulating in the blood of pregnant women afford the opportunity for early, noninvasive prenatal genetic testing. The predominance of admixed maternal genetic material in circulation demands innovative means for identification and analysis of cell-free fetal DNA and RNA. Techniques using polymerase chain reaction, mass spectrometry, and sequencing have been developed for the purposes of detecting fetal-specific sequences, such as paternally inherited or de novo mutations, or determining allelic balance or chromosome dosage. Clinical applications of these methods include fetal sex determination and blood group typing, which are currently available commercially although not offered routinely in the United States. Other uses of cell-free fetal DNA and RNA being explored are the detection of single-gene disorders, chromosomal abnormalities, and inheritance of parental polymorphisms across the whole fetal genome. The concentration of cell-free fetal DNA may also provide predictive capabilities for pregnancy-associated complications. The roles that cell-free fetal nucleic acid testing assume in the existing framework of prenatal screening and invasive diagnostic testing will depend on factors such as costs, clinical validity and utility, and perceived benefit-risk ratios for different applications. As cell-free fetal DNA and RNA testing continues to be developed and translated, significant ethical, legal, and social questions will arise that will need to be addressed by those with a stake in the use of this technology. Obstetricians & Gynecologists and Family Physicians Learning Objectives: After participating in this activity, physicians should be better able to evaluate techniques and tools for analyzing cell-free fetal nucleic acids, assess clinical applications of prenatal testing, using cell-free fetal nucleic acids and barriers to implementation, and distinguish between relevant clinical features of cell-free fetal nucleic acid testing and existing prenatal genetic screening and diagnostic procedures.

Fetal cell-free plasma DNA concentrations in maternal blood are stable 24 hours after collection: analysis of first- and third-trimester samples.

The discovery of intact fetal cells in maternal blood has led to the possibility of noninvasive prenatal genetic testing. Although extensive work has been performed on maternal blood in the hope of developing a reliable method for the recovery of intact fetal cells, the total number of recoverable cells is quite small, averaging 19 nucleated fetal cells in 16 mL of blood (1)(2). This is contrasted by a relatively large amount of cell-free fetal DNA detectable in maternal plasma (3). Recent studies suggest that cell-free fetal DNA in maternal plasma can be used as a diagnostic tool for diseases of pregnancy, such as preeclampsia or preterm labor, or for fetal anomalies, such as Down syndrome (4)(5)(6)(7). Because of these potential applications, it is of great importance to determine whether cell-free fetal DNA concentrations remain stable in a test tube after phlebotomy. In this report we evaluate the effects of time after phlebotomy on the quantity of cell-free fetal DNA in blood samples from a pregnant woman and a woman who has just undergone termination of pregnancy. The post-termination samples were used because in the first trimester, there are relatively few fetal cells present in maternal blood. The termination procedure increases fetomaternal transfusion (8). We hypothesized that after phlebotomy, apoptotic fetal cells that die in the venipuncture tube would release their DNA. We also asked whether more fetal cells undergo apoptosis as a result of exposure to the maternal bloodstream and cytokines. In both cases, we hypothesized that cell-free fetal DNA concentrations would increase in the blood samples over time. This study was performed with Institutional Review Board approval and with informed consent from participants at Tufts-New England Medical Center. For early-gestation samples, 29 women who were 6–17 weeks pregnant were recruited from …

Prenatal diagnosis using fetal cells isolated from maternal peripheral blood: a review.

Many questions remain about the feasibility of using fetal cells from maternal blood for prenatal diagnosis. Although recently there has been more focus on clinically relevant methods, many studies have been performed using blood drawn after invasive procedures, and over a wide range of gestational ages. For methods to be applicable to clinical use, more work is needed on isolating cells early in pregnancy, when termination is still an option for parents who are found to have an affected pregnancy. It is generally agreed that fetal nucleated erythrocytes are the most efficacious cell type for prenatal diagnosis, but it has not yet been shown definitively whether there is an ideal gestational age for sampling, whether ABO incompatibility might limit availability of fetal cells, or whether the number of cells present might be different in normal versus abnormal pregnancies. PCR has been shown to be a powerful tool in allowing amplification and identification of very small amounts of fetal DNA. However, this is limited to cases in which a specific and unique gene from the father is sought. This means that there is the potential to diagnose many paternally inherited autosomal dominant diseases and some autosomal recessive diseases, in which the parents have different and identifiable mutations. However, when parents are both carriers of the same autosomal recessive mutations, or when the disease is X linked, PCR will not aid in prenatal diagnosis. Cytogenetic analysis of fetal cells by FISH after cell sorting is another potentially useful method of prenatal diagnosis, but requires relatively pure samples of fetal cells or an independent marker that allows easy microscopic identification. The latter might be accomplished by identifying fetal cells through their expression of embryonic hemoglobins or because they contain HLA-G mRNA. In addition, current techniques of cell sorting must be improved so that a higher percentage of fetal cells can be isolated. Currently, the best cell sorting techniques usually produce a maximum purity of 10% fetal cells. Commonly, in normal pregnancies, fewer than 0.1% of the cells isolated after sorting are fetal in origin. Improving the concentration and quantity of fetal cells will improve the accuracy of FISH. Methods such as immunophenotyping that allow the selective identification of fetal cells by microscopy, and can be used in conjunction with FISH, may be extremely valuable because they may allow the genetic analysis of only the few fetal cells within a background preponderance of maternal cells. Although the retrieval of fetal cells from maternal blood is an attractive concept, it must be clearly stated that presently it is only in the investigational phase because of the low sensitivity and specificity. There is no current application for these methods in clinical practice. It remains to be determined whether testing maternal blood for fetal cells or DNA will be used as a screening tool, similar to the maternal serum screening currently in use, or whether the accuracy can be improved to a level such that the techniques can be used diagnostically. Although there are many questions that remain unanswered at this time, the outlook for noninvasive prenatal genetic testing in the future is optimistic.