Comprehensive Recommendations for the Clinical Management of Pregnant Women With Noninvasive Prenatal Test Results Suspicious of a Maternal Malignancy.

In this article, we defined comprehensive recommendations for the clinical follow-up of pregnant women with a malignancy-suspicious NIPT result, on the basis of the vast experience with population-based NIPT screening programs in two European countries complemented with published large data sets. These recommendations provide a tool for classifying NIPT results as malignancy-suspicious, and guide health care professionals in structured clinical decision making for the diagnostic process of pregnant women who receive such a malignancy-suspicious NIPT result.

Clinical Guideline for Preimplantation Genetic Testing in Inherited Cardiac Diseases.

BACKGROUND: Preimplantation genetic testing (PGT) is a reproductive technology that selects embryos without (familial) genetic variants. PGT has been applied in inherited cardiac disease and is included in the latest American Heart Association/American College of Cardiology guidelines. However, guidelines selecting eligible couples who will have the strongest risk reduction most from PGT are lacking. We developed an objective decision model to select eligibility for PGT and compared its results with those from a multidisciplinary team. METHODS: All couples with an inherited cardiac disease referred to the national PGT center were included. A multidisciplinary team approved or rejected the indication based on clinical and genetic information. We developed a decision model based on published risk prediction models and literature, to evaluate the severity of the cardiac phenotype and the penetrance of the familial variant in referred patients. The outcomes of the model and the multidisciplinary team were compared in a blinded fashion. RESULTS: Eighty-three couples were referred for PGT (1997-2022), comprising 19 different genes for 8 different inherited cardiac diseases (cardiomyopathies and arrhythmias). Using our model and proposed cutoff values, a definitive decision was reached for 76 (92%) couples, aligning with 95% of the multidisciplinary team decisions. In a prospective cohort of 11 couples, we showed the clinical applicability of the model to select couples most eligible for PGT. CONCLUSIONS: The number of PGT requests for inherited cardiac diseases increases rapidly, without the availability of specific guidelines. We propose a 2-step decision model that helps select couples with the highest risk reduction for cardiac disease in their offspring after PGT.

Considerations of breast cancer survivors to return for embryo transfer after fertility preservation: A qualitative study.

OBJECTIVE: To gain insight into the considerations of breast cancer survivors to return or not for embryo transfer after the use of fertility preservation. STUDY DESIGN: This is a qualitative study with semi-structured interviews. The interviews were planned until saturation of themes had been achieved. Content analysis was used to analyze the data. Sixteen out of 35 approached women took part in this study. Interviews were conducted with women who had oocytes or embryos cryopreserved prior to breast cancer treatment at the Maastricht University Medical Center between 2008 and 2016. All women who had cryopreservation more than two years ago were invited for the interviews. Women who had recurrence of disease were excluded. In the interviews we hypothesized the situation 'suppose the menses would have been recovered completely' for women who still had chemotherapy-induced menopause or used an GnRH (Gonadotropin-releasing hormone) analogue. RESULTS: Most women had a strong intrinsic motivation to pursue natural conception over the use of earlier cryopreserved oocytes or embryos. Time pressure was the most mentioned consideration to use cryopreserved oocytes or embryos. The wish to use pre-implantation genetic testing (PGT) in the presence of a germline BRCA1/2 mutation was another consideration to opt for embryo transfer. Furthermore, the physician's advice was an important motivation to choose for either natural conception or the use of cryopreserved oocytes or embryos. CONCLUSION: Multiple considerations influence women's decision making on the mode of conception after breast cancer. Although it concerned a single-center study in a highly-selected population, insight into these considerations can help physicians to address these important topics in counseling these women.

Preimplantation genetic testing for Neurofibromatosis type 1: more than 20 years of clinical experience.

Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder that affects the skin and the nervous system. The condition is completely penetrant with extreme clinical variability, resulting in unpredictable manifestations in affected offspring, complicating reproductive decision-making. One of the reproductive options to prevent the birth of affected offspring is preimplantation genetic testing (PGT). We performed a retrospective review of the medical files of all couples (n = 140) referred to the Dutch PGT expert center with the indication NF1 between January 1997 and January 2020. Of the couples considering PGT, 43 opted out and 15 were not eligible because of failure to identify the underlying genetic defect or unmet criteria for in vitro fertilization (IVF) treatment. The remaining 82 couples proceeded with PGT. Fertility assessment prior to IVF treatment showed a higher percentage of male infertility in males affected with NF1 compared to the partners of affected females. Cardiac evaluations in women with NF1 showed no contraindications for IVF treatment or pregnancy. For 67 couples, 143 PGT cycles were performed. Complications of IVF treatment were not more prevalent in affected females compared to partners of affected males. The transfer of 174 (out of 295) unaffected embryos led to 42 ongoing pregnancies with a pregnancy rate of 24.1% per embryo transfer. There are no documented cases of misdiagnosis following PGT in this cohort. With these results, we aim to provide an overview of PGT for NF1 with regard to success rate and safety, to optimize reproductive counseling and PGT treatment for NF1 patients.

Consanguineous couples' experiences and views regarding expanded carrier screening: Barriers and facilitators in the decision-making process.

Expanded carrier screening (ECS) entails a screening offer for multiple recessive disorders at the same time, and allows testing of individuals or couples regardless of ancestry or geographic origin. Children of consanguineous couples have a higher-than-average risk of manifesting autosomal recessive disorders. This study aims to contribute to the responsible implementation of ECS for consanguineous couples. Seven semi-structured interviews were conducted with consanguineous couples who had recently participated in Whole Exome Sequencing (WES)-based ECS at Maastricht University Medical Center (MUMC+), the Netherlands. The test offered at MUMC+ covers a large number of disease-related genes (~2000), including severe, relatively mild, early- and late-onset disorders. Respondents were interviewed about their views on, and experiences with participation in WES-based ECS. Overall, participation was experienced as worthwhile: it enabled respondents to make informed choices with regard to family planning as well as to take on the presumed parental responsibility to deliver their children as healthy as possible. Furthermore, our findings suggest that (1) true consent for having this test requires timely information about the possible implications of a positive test result for specific categories of findings, as well as about the success rates of the available reproductive options; (2) the clinical geneticist can play a pivotal part in informing participants as well as providing clear information about autosomal recessive inheritance; (3) more research is needed to explore what type of genetic risk information is considered 'meaningful' by participants and actually contributes to reproductive decision-making.

[Couples with risk of a child with a mitochondrial disease: wat are the reproductive options?] / Paren met risico op een kind met een mitochondriële ziekte.

Mitochondrial diseases are the most common inborn errors of metabolism. These severe multisystem disorders cause serious morbidity and mortality. Generally no treatment is available. This underlines the importance of counseling about the reproductive options to prevent the transmission of mitochondrial disorders. The majority of mitochondrial disorders is caused by a defect in a nuclear gene, in which cases the standard reproductive options can be applied, such as prenatal diagnosis (PND) and preimplantation genetic testing (PGT). For mitochondrial disorders caused by a mitochondrial DNA (mtDNA) mutation, reproductive options are determined by the recurrence risk, requiring specific reproductive counseling. For de novomtDNA mutations and inherited mtDNA mutations with a low recurrence risk, PND is possible. In case of a moderate or higher recurrence risk, PGT is the best option. In case the risk of a healthy embryo is (very) low, mitochondrial replacement therapy (MRT) may be a possibility in the future.

The effects of an online decision aid to support the reproductive decision-making process of genetically at risk couples-A pilot study.

Couples at risk of transmitting a genetic disease to their offspring may experience doubts about their reproductive options. This study examines the effects of an online decision aid (DA) on the (joint) reproductive decision-making process of couples (not pregnant at time of inclusion) at risk of transmitting a genetic disease to their offspring. The primary outcome is decisional conflict, and secondary outcomes are knowledge, realistic expectations, deliberation, joint informed decision-making, and decisional self-efficacy. These outcomes were measured with a pretest-posttest design: before use (T0), after use (T1), and 2 weeks after use (T2) of the decision aid (DA). Usability of the DA was assessed at T1. Paired sample t-tests were used to compute differences between baseline and subsequent measurements. The comparisons of T0-T1 and T0-T2 indicate a significant reduction in mean decisional conflict scores with stronger effects for participants with high baseline decisional conflict scores. Furthermore, use of the DA led to increased knowledge, improved realistic expectations, and increased levels of deliberation, with higher increase in participants with low baseline scores. Decision self-efficacy only improved for participants with lower baseline scores. Participants indicated that the information in the DA was comprehensible and clearly organized. These first results indicate that this online DA is an appropriate tool to support couples at risk of transmitting a genetic disease and a desire to have (a) child(ren) in their reproductive decision-making process.

Embryo tracking system for high-throughput sequencing-based preimplantation genetic testing.

STUDY QUESTION: Can the embryo tracking system (ETS) increase safety, efficacy and scalability of massively parallel sequencing-based preimplantation genetic testing (PGT)? SUMMARY ANSWER: Applying ETS-PGT, the chance of sample switching is decreased, while scalability and efficacy could easily be increased substantially. WHAT IS KNOWN ALREADY: Although state-of-the-art sequencing-based PGT methods made a paradigm shift in PGT, they still require labor intensive library preparation steps that makes PGT cost prohibitive and poses risks of human errors. To increase the quality assurance, efficiency, robustness and throughput of the sequencing-based assays, barcoded DNA fragments have been used in several aspects of next-generation sequencing (NGS) approach. STUDY DESIGN, SIZE, DURATION: We developed an ETS that substantially alleviates the complexity of the current sequencing-based PGT. With (n = 693) and without (n = 192) ETS, the downstream PGT procedure was performed on both bulk DNA samples (n = 563) and whole-genome amplified (WGAed) few-cell DNA samples (n = 322). Subsequently, we compared full genome haplotype landscapes of both WGAed and bulk DNA samples containing ETS or no ETS. PARTICIPANTS/MATERIALS, SETTING, METHODS: We have devised an ETS to track embryos right after whole-genome amplification (WGA) to full genome haplotype profiles. In this study, we recruited 322 WGAed DNA samples derived from IVF embryos as well as 563 bulk DNA isolated from peripheral blood of prospective parents. To determine possible interference of the ETS in the NGS-based PGT workflow, barcoded DNA fragments were added to DNA samples prior to library preparation and compared to samples without ETS. Coverages and variants were determined. MAIN RESULTS AND THE ROLE OF CHANCE: Current PGT protocols are quality sensitive and prone to sample switching. To avoid sample switching and increase throughput of PGT by sequencing-based haplotyping, six control steps should be carried out manually and checked by a second person in a clinical setting. Here, we developed an ETS approach in which one step only in the entire PGT procedure needs the four-eyes principal. We demonstrate that ETS not only precludes error-prone manual checks but also has no effect on the genomic landscape of preimplantation embryos. Importantly, our approach increases efficacy and throughput of the state-of-the-art PGT methods. LIMITATIONS, REASONS FOR CAUTION: Even though the ETS simplified sequencing-based PGT by avoiding potential errors in six steps in the protocol, if the initial assignment is not performed correctly, it could lead to cross-contamination. However, this can be detected in silico following downstream ETS analysis. Although we demonstrated an approach to evaluate purity of the ETS fragment, it is recommended to perform a pre-PGT quality control assay of the ETS amplicons with non-human DNA, such that the purity of each ETS molecule can be determined prior to ETS-PGT. WIDER IMPLICATIONS OF THE FINDINGS: The ETS-PGT approach notably increases efficacy and scalability of PGT. ETS-PGT has broad applicative value, as it can be tailored to any single- and few-cell sequencing approach where the starting specimen is scarce, as opposed to other methods that require a large number of cells as the input. Moreover, ETS-PGT could easily be adapted to any sequencing-based diagnostic method, including PGT for structural rearrangements and aneuploidies by low-pass sequencing as well as non-invasive prenatal testing. STUDY FUNDING/COMPETING INTEREST(S): M.Z.E. is supported by the EVA (Erfelijkheid Voortplanting & Aanleg) specialty program (grant no. KP111513) of Maastricht University Medical Centre (MUMC+), and the Horizon 2020 innovation (ERIN) (grant no. EU952516) of the European Commission. TRIAL REGISTRATION NUMBER: N/A.

Noninvasive Prenatal Test Results Indicative of Maternal Malignancies: A Nationwide Genetic and Clinical Follow-Up Study.

PURPOSE: Noninvasive prenatal testing (NIPT) for fetal aneuploidy screening using cell-free DNA derived from maternal plasma can incidentally raise suspicion for cancer. Diagnostic routing after malignancy suspicious-NIPT faces many challenges. Here, we detail malignancy suspicious-NIPT cases, and describe the clinical characteristics, chromosomal aberrations, and diagnostic routing of the patients with a confirmed malignancy. Clinical lessons can be learned from our experience. METHODS: Patients with NIPT results indicative of a malignancy referred for tumor screening between April 2017 and April 2020 were retrospectively included from a Dutch nationwide NIPT implementation study, TRIDENT-2. NIPT profiles from patients with confirmed malignancies were reviewed, and the pattern of chromosomal aberrations related to tumor type was analyzed. We evaluated the diagnostic contribution of clinical and genetic examinations. RESULTS: Malignancy suspicious-NIPT results were reported in 0.03% after genome-wide NIPT, and malignancies confirmed in 16 patients (16/48, 33.3%). Multiple chromosomal aberrations were seen in 23 of 48 patients with genome-wide NIPT, and a malignancy was confirmed in 16 patients (16/23, 69.6%). After targeted NIPT, 0.005% malignancy suspicious-NIPT results were reported, in 2/3 patients a malignancy was confirmed. Different tumor types and stages were diagnosed, predominantly hematologic malignancies (12/18). NIPT data showed recurrent gains and losses in primary mediastinal B-cell lymphomas and classic Hodgkin lymphomas. Magnetic resonance imaging and computed tomography were most informative in diagnosing the malignancy. CONCLUSION: In 231,896 pregnant women, a low percentage (0.02%) of NIPT results were assessed as indicative of a maternal malignancy. However, when multiple chromosomal aberrations were found, the risk of a confirmed malignancy was considerably high. Referral for extensive oncologic examination is recommended, and may be guided by tumor-specific hallmarks in the NIPT profile.

Facioscapulohumeral muscular dystrophy-Reproductive counseling, pregnancy, and delivery in a complex multigenetic disease.

Reproductive counseling in facioscapulohumeral muscular dystrophy (FSHD) can be challenging due to the complexity of its underlying genetic mechanisms and due to incomplete penetrance of the disease. Full understanding of the genetic causes and potential inheritance patterns of both distinct FSHD types is essential: FSHD1 is an autosomal dominantly inherited repeat disorder, whereas FSHD2 is a digenic disorder. This has become even more relevant now that prenatal diagnosis and preimplantation genetic diagnosis options are available for FSHD1. Pregnancy and delivery outcomes in FSHD are usually favorable, but clinicians should be aware of the risks. We aim to provide clinicians with case-based strategies for reproductive counseling in FSHD, as well as recommendations for pregnancy and delivery.

Liquid biopsy: state of reproductive medicine and beyond.

Liquid biopsy is the process of sampling and analyzing body fluids, which enables non-invasive monitoring of complex biological systems in vivo. Liquid biopsy has myriad applications in health and disease as a wide variety of components, ranging from circulating cells to cell-free nucleic acid molecules, can be analyzed. Here, we review different components of liquid biopsy, survey state-of-the-art, non-invasive methods for detecting those components, demonstrate their clinical applications and discuss ethical considerations. Furthermore, we emphasize the importance of artificial intelligence in analyzing liquid biopsy data with the aim of developing ethically-responsible non-invasive technologies that can enhance individualized healthcare. While previous reviews have mainly focused on cancer, this review primarily highlights applications of liquid biopsy in reproductive medicine.

Disease Onset in Huntington's Disease: When Is the Conversion?

BACKGROUND: Determination of disease onset in Huntington's disease is made by clinical experience. The diagnostic confidence level is an assessment regarding the certainty about the clinical diagnosis based on motor signs. A level of 4 means the rater has ≥99% confidence motor abnormalities are unequivocal signs of disease. However, it does not state which motor abnormalities are signs of disease and how many must be present. OBJECTIVE: Our aim is to explore how accurate the diagnostic confidence level is in estimating disease onset using the Enroll-HD data set. For clinical disease onset we use a cut-off total motor score >5 of the Unified Huntington's Disease Rating Scale. This score is used in the TRACK-HD study, with ≤5 indicating no substantial motor signs in premanifests. METHODS: At baseline premanifest participants who converted to manifest (converters) and non-converters were compared for clinical symptoms and diagnostic confidence level. Clinical symptoms and diagnostic confidence levels were longitudinally displayed in converters. RESULTS: Of 3731 eligible participants, 455 were converters and 3276 non-converters. Baseline diagnostic confidence levels were significantly higher in converters compared to non-converters (P < 0.001). 232 (51%) converters displayed a baseline motor score >5 (mean = 6.7). Converters had significantly more baseline clinical symptoms, and higher disease burden compared to non-converters (P < 0.001). Diagnostic confidence level before disease onset ranged between 1 and 3 in converters. CONCLUSIONS: According to this data the diagnostic confidence level is not an accurate instrument to determine phenoconversion. With trials evaluating disease modifying therapies it is important to develop more reliable diagnostic criteria.

Diagnostic exome-based preconception carrier testing in consanguineous couples: results from the first 100 couples in clinical practice.

PURPOSE: Consanguineous couples are at increased risk of being heterozygous for the same autosomal recessive (AR) disorder(s), with a 25% risk of affected offspring as a consequence. Until recently, comprehensive preconception carrier testing (PCT) for AR disorders was unavailable in routine diagnostics. Here we developed and implemented such a test in routine clinical care. METHODS: We performed exome sequencing (ES) for 100 consanguineous couples. For each couple, rare variants that could give rise to biallelic variants in offspring were selected. These variants were subsequently filtered against a gene panel consisting of ~2,000 genes associated with known AR disorders (OMIM-based). Remaining variants were classified according to American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines, after which only likely pathogenic and pathogenic (class IV/V) variants, present in both partners, were reported. RESULTS: In 28 of 100 tested consanguineous couples (28%), likely pathogenic and pathogenic variants not previously known in the couple or their family were reported conferring 25% risk of affected offspring. CONCLUSION: ES-based PCT provides a powerful diagnostic tool to identify AR disease carrier status in consanguineous couples. Outcomes provided significant reproductive choices for a higher proportion of these couples than previous tests.

Reproductive decision-making in the context of hereditary cancer: the effects of an online decision aid on informed decision-making.

Individuals having a genetic predisposition to cancer and their partners face challenging decisions regarding their wish to have children. This study aimed to determine the effects of an online decision aid to support couples in making an informed decision regarding their reproductive options. A nationwide pretest-posttest study was conducted in the Netherlands among 131 participants between November 2016 and May 2018. Couples were eligible for participation if one partner had a pathogenic variant predisposing for an autosomal dominant hereditary cancer syndrome. Participants completed a questionnaire before use (T0), and at 3 months (T3) after use of the decision aid to assess the primary outcome measure informed decision-making, and the secondary outcome measures decisional conflict, knowledge, realistic expectations, level of deliberation, and decision self-efficacy. T0-T3 comparisons show an overall positive effect for all outcome measures (all ps < 0.05; knowledge (ES = - 1.05), decisional conflict (ES = 0.99), participants' decision self-efficacy (ES = -0.55), level of deliberation (ES = - 0.50), and realistic expectations (ES = - 0.44). Informed decision-making increased over time and 58.0% of the participants made an informed reproductive decision at T3. The online decision aid seems to be an appropriate tool to complement standard reproductive counseling to support our target group in making an informed reproductive decision. Use of the decision aid may lessen the negative psychological impact of decision-making on couples' daily life and wellbeing.

Preimplantation Genetic Testing for Monogenic Kidney Disease.

BACKGROUND AND OBJECTIVES: A genetic cause can be identified for an increasing number of pediatric and adult-onset kidney diseases. Preimplantation genetic testing (formerly known as preimplantation genetic diagnostics) is a reproductive technology that helps prospective parents to prevent passing on (a) disease-causing mutation(s) to their offspring. Here, we provide a clinical overview of 25 years of preimplantation genetic testing for monogenic kidney disease in The Netherlands. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: This is a retrospective cohort study of couples counseled on preimplantation genetic testing for monogenic kidney disease in the national preimplantation genetic testing expert center (Maastricht University Medical Center+) from January 1995 to June 2019. Statistical analysis was performed through chi-squared tests. RESULTS: In total, 98 couples were counseled regarding preimplantation genetic testing, of whom 53% opted for preimplantation genetic testing. The most frequent indications for referral were autosomal dominant polycystic kidney disease (38%), Alport syndrome (26%), and autosomal recessive polycystic kidney disease (9%). Of couples with at least one preimplantation genetic testing cycle with oocyte retrieval, 65% experienced one or more live births of an unaffected child. Of couples counseled, 38% declined preimplantation genetic testing for various personal and technical reasons. CONCLUSIONS: Referrals, including for adult-onset disease, have increased steadily over the past decade. Though some couples decline preimplantation genetic testing, in the couples who proceed with at least one preimplantation genetic testing cycle, almost two thirds experienced at least one live birth rate.

Predictive genetic testing in Huntington's disease: should a neurologist be involved?

International guidelines on Huntington's Disease recommend neurological examination in the predictive testing trajectory. Experiences and personal wishes of persons at risk of Huntington's Disease regarding this topic have never been evaluated. The objective was to provide an overview of the experiences of Dutch at-risk persons, opting for predictive testing, in consulting a neurologist before and after DNA analysis. Persons who were counseled in four Dutch clinics between 2017 and 2019 were retrospectively or prospectively approached for a questionnaire which listed topics as experiences with consultation and personal wishes. From 71 participants, 44 participants visited a neurologist. 41 participants indicated their visit to a neurologist as positive (93.2%). The majority of participants (n = 59) desired consulting a neurologist. Thirty-two participants indicated consultation shortly after (Desired After Group) and twenty-seven before DNA analysis (Desired Before Group) as personal wish. The Desired Before Group consisted of a significantly higher number of participants who actually consulted a neurologist before predictive testing (n = 26) compared with the number of participants who actually consulted a neurologist after DNA analysis in the Desired After Group (n = 11) (p < 0.001). The Desired After Group (n = 19) had a significantly higher number of Huntington's disease gene expansion carriers compared with the Desired Before Group (n = 5) (p 0.003). Participants are content with consultation. However, persons without the gene expansion still feel the need to get in touch with a neurologist. Therefore, offering a consultation with a neurologist before DNA analysis might be beneficial for all.

Preserving fertility in young women undergoing chemotherapy for early breast cancer; the Maastricht experience.

PURPOSE: We assessed the uptake of fertility preservation (FP), recovery of ovarian function (OFR) after chemotherapy, live birth after breast cancer, and breast cancer outcomes in women with early-stage breast cancer. METHODS: Women aged below 41 years and referred to our center for FP counseling between 2008 and 2015 were included. Data on patient and tumor characteristics, ovarian function, cryopreservation (embryo/oocyte) and transfer, live birth, and disease-free survival were collected. Kaplan-Meier analyses were performed for time-to-event analyses including competing risk analyses, and patients with versus without FP were compared using the logrank test. RESULTS: Of 118 counseled women with a median age of 31 years (range 19-40), 34 (29%) chose FP. Women who chose FP had less often children, more often a male partner and more often favorable tumor characteristics. The 5-year OFR rate was 92% for the total group of counseled patients. In total, 26 women gave birth. The 5-year live birth rate was 27% for the total group of counseled patients. Only three women applied for transfer of their cryopreserved embryo(s), in two combined with preimplantation genetic diagnosis (PGD) because of BRCA1-mutation carrier ship. The 5-year disease-free survival rate was 91% versus 88%, for patients with versus without FP (P = 0.42). CONCLUSIONS: Remarkably, most women achieved OFR, probably related to the young age at diagnosis. Most pregnancies occurred spontaneously, two of three women applied for embryo transfer because of the opportunity to apply for PGD.

Parental repeat length instability in myotonic dystrophy type 1 pre- and protomutations.

Myotonic dystrophy type 1 (DM1) is caused by a CTG trinucleotide repeat expansion on chromosome 19q13.3. While DM1 premutation (36-50 repeats) and protomutation (51-80 repeats) allele carriers are mostly asymptomatic, offspring is at risk of inheriting expanded, symptom-associated, (CTG)n repeats of n > 80. In this study we aimed to evaluate the intergenerational instability of DM1 pre- and protomutation alleles, focussing on the influence of parental gender. One hundred and forty-six parent-child pairs (34 parental premutations, 112 protomutations) were retrospectively selected from the DM1 patient cohort of the Maastricht University Medical Center+. CTG repeat size of parents and children was determined by (triplet-primed) PCR followed by fragment length analysis and Southern blot analysis. Fifty-eight out of eighty-one (71.6%) paternal transmissions led to a (CTG)n repeat of n > 80 in offspring, compared with 15 out of 65 (23.1%) maternal transmissions (p < 0.001). Repeat length instability occurred for paternal (CTG)n repeats of n ≥ 45, while maternal instability did not occur until (CTG)n repeats reached a length of n ≥ 71. Transmission of premutations caused (CTG)n repeats of n > 80 in offspring only when paternally transmitted (two cases), while protomutations caused (CTG)n repeats of n > 80 in offspring in 71 cases, of which 56 (78.9%) were paternally transmitted. In conclusion, our data show that paternally transmitted pre- and protomutations were more unstable than maternally transmitted pre- and protomutations. For genetic counseling, this implies that males with a small DMPK mutation have a higher risk of symptomatic offspring compared with females. Consequently, we suggest addressing sex-dependent factors in genetic counseling of small-sized CTG repeat carriers.

TRIDENT-2: National Implementation of Genome-wide Non-invasive Prenatal Testing as a First-Tier Screening Test in the Netherlands.

The Netherlands launched a nationwide implementation study on non-invasive prenatal testing (NIPT) as a first-tier test offered to all pregnant women. This started on April 1, 2017 as the TRIDENT-2 study, licensed by the Dutch Ministry of Health. In the first year, NIPT was performed in 73,239 pregnancies (42% of all pregnancies), 7,239 (4%) chose first-trimester combined testing, and 54% did not participate. The number of trisomies 21 (239, 0.33%), 18 (49, 0.07%), and 13 (55, 0.08%) found in this study is comparable to earlier studies, but the Positive Predictive Values (PPV)-96% for trisomy 21, 98% for trisomy 18, and 53% for trisomy 13-were higher than expected. Findings other than trisomy 21, 18, or 13 were reported on request of the pregnant women; 78% of women chose to have these reported. The number of additional findings was 207 (0.36%); these included other trisomies (101, 0.18%, PPV 6%, many of the remaining 94% of cases are likely confined placental mosaics and possibly clinically significant), structural chromosomal aberrations (95, 0.16%, PPV 32%,) and complex abnormal profiles indicative of maternal malignancies (11, 0.02%, PPV 64%). The implementation of genome-wide NIPT is under debate because the benefits of detecting other fetal chromosomal aberrations must be balanced against the risks of discordant positives, parental anxiety, and a potential increase in (invasive) diagnostic procedures. Our first-year data, including clinical data and laboratory follow-up data, will fuel this debate. Furthermore, we describe how NIPT can successfully be embedded into a national screening program with a single chain for prenatal care including counseling, testing, and follow-up.