Inactivating TDP2 missense mutation in siblings with congenital abnormalities reminiscent of fanconi anemia.

Mutations in TDP2, encoding tyrosyl-DNA phosphodiesterase 2, have been associated with a syndromal form of autosomal recessive spinocerebellar ataxia, type 23 (SCAR23). This is a very rare and progressive neurodegenerative disorder described in only nine patients to date, and caused by splice site or nonsense mutations that result in greatly reduced or absent TDP2 protein. TDP2 is required for the rapid repair of DNA double-strand breaks induced by abortive DNA topoisomerase II (TOP2) activity, important for genetic stability in post-mitotic cells such as neurons. Here, we describe a sibship that is homozygous for the first TDP2 missense mutation (p.Glu152Lys) and which presents with clinical features overlapping both SCAR23 and Fanconi anemia (FA). We show that in contrast to previously reported SCAR23 patients, fibroblasts derived from the current patient retain significant levels of TDP2 protein. However, this protein is catalytically inactive, resulting in reduced rates of repair of TOP2-induced DNA double-strand breaks and cellular hypersensitivity to the TOP2 poison, etoposide. The TDP2-mutated patient-derived fibroblasts do not display increased chromosome breakage following treatment with DNA crosslinking agents, but both TDP2-mutated and FA cells exhibit increased chromosome breakage in response to etoposide. This suggests that the FA pathway is required in response to TOP2-induced DNA lesions, providing a possible explanation for the clinical overlap between FA and the current TDP2-mutated patients. When reviewing the relatively small number of patients with SCAR23 that have been reported, it is clear that the phenotype of such patients can extend beyond neurological features, indicating that the TDP2 protein influences not only neural homeostasis but also other tissues as well.

What helps define outcomes in persistent uninterpretable non-invasive prenatal testing: Maternal factors, fetal fraction or quality scores?

OBJECTIVES: To assess maternal characteristics and comorbidities in patients with persistent uninterpretable non-invasive prenatal testing (NIPT) and to evaluate the association with adverse pregnancy outcome in a general risk population. METHODS: A retrospective cohort study (July 2017-December 2020) was conducted of patients with persistent uninterpretable NIPT samples. Maternal characteristics and pregnancy outcomes were compared with the general Belgian obstetric population. RESULTS: Of the 148 patients with persistent uninterpretable NIPT, 37 cases were due to a low fetal fraction (LFF) and 111 due to a low quality score (LQS). Both groups (LFF, LQS) showed more obesity (60.6%, 42.4%), multiple pregnancies (18.9%, 4.5%) and more obstetrical complications. In the LQS group, a high rate of maternal auto-immune disorders (30.6%) was seen and hypertensive complications (17.6%), preterm birth (17.6%) and neonatal intensive care unit (NICU) admission (22%) were significantly increased. In the LFF group hypertensive complications (21.6%), gestational diabetes (20.6%), preterm birth (27%), SGA (25.6%), major congenital malformations (11.4%), c-section rate (51.4%) and NICU admission (34.9%) were significantly increased. Chromosomal abnormalities were not increased in both groups. CONCLUSIONS: Patients with persistent uninterpretable NIPT have significantly more maternal obesity, comorbidities and adverse pregnancy outcome than the general population and should receive high-risk pregnancy care. Distinguishing between LFF and LQS optimizes counseling because maternal characteristics and pregnancy outcome differ between these groups.

FOSL2 truncating variants in the last exon cause a neurodevelopmental disorder with scalp and enamel defects.

PURPOSE: We aimed to investigate the molecular basis of a novel recognizable neurodevelopmental syndrome with scalp and enamel anomalies caused by truncating variants in the last exon of the gene FOSL2, encoding a subunit of the AP-1 complex. METHODS: Exome sequencing was used to identify genetic variants in all cases, recruited through Matchmaker exchange. Gene expression in blood was analyzed using reverse transcription polymerase chain reaction. In vitro coimmunoprecipitation and proteasome inhibition assays in transfected HEK293 cells were performed to explore protein and AP-1 complex stability. RESULTS: We identified 11 individuals from 10 families with mostly de novo truncating FOSL2 variants sharing a strikingly similar phenotype characterized by prenatal growth retardation, localized cutis scalp aplasia with or without skull defects, neurodevelopmental delay with autism spectrum disorder, enamel hypoplasia, and congenital cataracts. Mutant FOSL2 messenger RNAs escaped nonsense-mediated messenger RNA decay. Truncated FOSL2 interacts with c-JUN, thus mutated AP-1 complexes could be formed. CONCLUSION: Truncating variants in the last exon of FOSL2 associate a distinct clinical phenotype by altering the regulatory degradation of the AP-1 complex. These findings reveal a new role for FOSL2 in human pathology.

Pan-Cancer Detection and Typing by Mining Patterns in Large Genome-Wide Cell-Free DNA Sequencing Datasets.

BACKGROUND: Cell-free DNA (cfDNA) analysis holds great promise for non-invasive cancer screening, diagnosis, and monitoring. We hypothesized that mining the patterns of cfDNA shallow whole-genome sequencing datasets from patients with cancer could improve cancer detection. METHODS: By applying unsupervised clustering and supervised machine learning on large cfDNA shallow whole-genome sequencing datasets from healthy individuals (n = 367) and patients with different hematological (n = 238) and solid malignancies (n = 320), we identified cfDNA signatures that enabled cancer detection and typing. RESULTS: Unsupervised clustering revealed cancer type-specific sub-grouping. Classification using a supervised machine learning model yielded accuracies of 96% and 65% in discriminating hematological and solid malignancies from healthy controls, respectively. The accuracy of disease type prediction was 85% and 70% for the hematological and solid cancers, respectively. The potential utility of managing a specific cancer was demonstrated by classifying benign from invasive and borderline adnexal masses with an area under the curve of 0.87 and 0.74, respectively. CONCLUSIONS: This approach provides a generic analytical strategy for non-invasive pan-cancer detection and cancer type prediction.

Identifying phenotypic expansions for congenital diaphragmatic hernia plus (CDH+) using DECIPHER data.

Congenital diaphragmatic hernia (CDH) can occur in isolation or in conjunction with other birth defects (CDH+). A molecular etiology can only be identified in a subset of CDH cases. This is due, in part, to an incomplete understanding of the genes that contribute to diaphragm development. Here, we used clinical and molecular data from 36 individuals with CDH+ who are cataloged in the DECIPHER database to identify genes that may play a role in diaphragm development and to discover new phenotypic expansions. Among this group, we identified individuals who carried putatively deleterious sequence or copy number variants affecting CREBBP, SMARCA4, UBA2, and USP9X. The role of these genes in diaphragm development was supported by their expression in the developing mouse diaphragm, their similarity to known CDH genes using data from a previously published and validated machine learning algorithm, and/or the presence of CDH in other individuals with their associated genetic disorders. Our results demonstrate how data from DECIPHER, and other public databases, can be used to identify new phenotypic expansions and suggest that CREBBP, SMARCA4, UBA2, and USP9X play a role in diaphragm development.

Exome sequencing efficacy and phenotypic expansions involving esophageal atresia/tracheoesophageal fistula plus.

Esophageal atresia/tracheoesophageal fistula (EA/TEF) is a life-threatening birth defect that often occurs with other major birth defects (EA/TEF+). Despite advances in genetic testing, a molecular diagnosis can only be made in a minority of EA/TEF+ cases. Here, we analyzed clinical exome sequencing data and data from the DECIPHER database to determine the efficacy of exome sequencing in cases of EA/TEF+ and to identify phenotypic expansions involving EA/TEF. Among 67 individuals with EA/TEF+ referred for clinical exome sequencing, a definitive or probable diagnosis was made in 11 cases for an efficacy rate of 16% (11/67). This efficacy rate is significantly lower than that reported for other major birth defects, suggesting that polygenic, multifactorial, epigenetic, and/or environmental factors may play a particularly important role in EA/TEF pathogenesis. Our cohort included individuals with pathogenic or likely pathogenic variants that affect TCF4 and its downstream target NRXN1, and FANCA, FANCB, and FANCC, which are associated with Fanconi anemia. These cases, previously published case reports, and comparisons to other EA/TEF genes made using a machine learning algorithm, provide evidence in support of a potential pathogenic role for these genes in the development of EA/TEF.

8p21.3 deletions are rare causes of non-syndromic autism spectrum disorder.

A de novo 0.95 Mb 8p21.3 deletion had been identified in an individual with non-syndromic autism spectrum disorder (ASD) through high-resolution copy number variant analysis. Subsequent screening of in-house and publicly available databases resulted in the identification of six additional individuals with 8p21.3 deletions. Through case-based reasoning, we conclude that 8p21.3 deletions are rare causes of non-syndromic neurodevelopmental and neuropsychiatric disorders. Based on literature data, we highlight six genes within the region of minimal overlap as potential ASD genes or genes for neuropsychiatric disorders: DMTN, EGR3, FGF17, LGI3, PHYHIP, and PPP3CC.

Outcome of publicly funded nationwide first-tier noninvasive prenatal screening.

PURPOSE: Noninvasive prenatal screening (NIPS) using cell-free DNA has transformed prenatal care. Belgium was the first country to implement and fully reimburse NIPS as a first-tier screening test offered to all pregnant women. A consortium consisting of all Belgian genetic centers report the outcome of two years genome-wide NIPS implementation. METHODS: The performance for the common trisomies and for secondary findings was evaluated based on 153,575 genome-wide NIP tests. Furthermore, the evolution of the number of invasive tests and the incidence of Down syndrome live births was registered. RESULTS: Trisomies 21, 18, and 13 were detected in respectively 0.32%, 0.07%, and 0.06% of cases, with overall positive predictive values (PPVs) of 92.4%, 84.6%, and 43.9%. Rare autosomal trisomies and fetal segmental imbalances were detected in respectively 0.23% and 0.07% of cases with PPVs of 4.1% and 47%. The number of invasive obstetric procedures decreased by 52%. The number of trisomy 21 live births dropped to 0.04%. CONCLUSION: Expanding the scope of NIPS beyond trisomy 21 fetal screening allows the implementation of personalized genomic medicine for the obstetric population. This genome-wide NIPS approach has been embedded successfully in prenatal genetic care in Belgium and might serve as a framework for other countries offering NIPS.

Missing heritability in Bloom syndrome: First report of a deep intronic variant leading to pseudo-exon activation in the BLM gene.

Pathogenic biallelic variants in the BLM/RECQL3 gene cause a rare autosomal recessive disorder called Bloom syndrome (BS). This syndrome is characterized by severe growth delay, immunodeficiency, dermatological manifestations and a predisposition to a wide variety of cancers, often multiple and very early in life. Literature shows that the main mode of BLM inactivation is protein translation termination. We expanded the molecular spectrum of BS by reporting the first deep intronic variant causing intron exonisation. We describe a patient with a clinical phenotype of BS and a strong increase in sister chromatid exchanges (SCE), who was found to be compound heterozygous for a novel nonsense variant c.3379C>T, p.(Gln1127Ter) in exon 18 and a deep intronic variant c.3020-258A>G in intron 15 of the BLM gene. The deep intronic variant creates a high-quality de novo donor splice site, which leads to retention of two intron segments. Both pseudo-exons introduce a premature stop codon into the reading frame and abolish BLM protein expression, confirmed by Western Blot analysis. These findings illustrate the role of non-coding variation in Mendelian disorders and herewith highlight an unmet need in routine testing of Mendelian disorders, being the added value of RNA-based approaches to provide a complete molecular diagnosis.

Non-invasive prenatal testing suggesting a maternal malignancy: What do we tell the prospective parents in Belgium?

Cancer is diagnosed in one in 1000 to 1500 pregnancies. Most frequently encountered malignancies during pregnancy are breast cancer, hematological cancer, cervical cancer and malignant melanoma. Maternal cancer is associated with an increased risk of IUGR and preterm labor, especially in patients with systemic disease or those receiving chemotherapy during pregnancy, requiring a high-risk obstetrical follow-up. Fetal aneuploidy screening by non-invasive prenatal testing (NIPT) can lead to the incidental identification of copy number alterations derived from non-fetal cell-free DNA (cfDNA), as seen in certain cases of maternal malignancy. The identification of tumor-derived cfDNA requires further clinical, biochemical, radiographic and histological investigations to confirm the diagnosis. In such cases, reliable risk estimation for fetal trisomy 21, 18 and 13 is impossible. Therefore, invasive testing should be offered when ultrasonographic screening reveals an increased risk for chromosomal anomalies, or when a more accurate test is desired. When the fetal karyotype is normal, long term implications for the fetus refer to the consequences of the maternal disease and treatment during pregnancy. This manuscript addresses parental questions when NIPT suggests a maternal malignancy. Based on current evidence and our own experience, a clinical management scheme in a multidisciplinary setting is proposed.

Noninvasive prenatal diagnosis by genome-wide haplotyping of cell-free plasma DNA.

PURPOSE: Whereas noninvasive prenatal screening for aneuploidies is widely implemented, there is an increasing need for universal approaches for noninvasive prenatal screening for monogenic diseases. Here, we present a cost-effective, generic cell-free fetal DNA (cffDNA) haplotyping approach to scan the fetal genome for the presence of inherited monogenic diseases. METHODS: Families participating in the preimplantation genetic testing for monogenic disorders (PGT-M) program were recruited for this study. Two hundred fifty thousand single-nucleotide polymorphisms (SNPs) captured from maternal plasma DNA along with genomic DNA from family members were massively parallel sequenced. Parental genotypes were phased via an available genotype from a close relative, and the fetal genome-wide haplotype and copy number were determined using cffDNA haplotyping analysis based on estimation and segmentation of fetal allele presence in the maternal plasma. RESULTS: In all families tested, mutational profiles from cffDNA haplotyping are consistent with embryo biopsy profiles. Genome-wide fetal haplotypes are on average 97% concordant with the newborn haplotypes and embryo haplotypes. CONCLUSION: We demonstrate that genome-wide targeted capture and sequencing of polymorphic SNPs from maternal plasma cell-free DNA (cfDNA) allows haplotyping and copy-number profiling of the fetal genome during pregnancy. The method enables the accurate reconstruction of the fetal haplotypes and can be easily implemented in clinical practice.

Identity-by-state-based haplotyping expands the application of comprehensive preimplantation genetic testing.

STUDY QUESTION: Is it possible to haplotype parents using parental siblings to leverage preimplantation genetic testing (PGT) for monogenic diseases and aneuploidy (comprehensive PGT) by genome-wide haplotyping? SUMMARY ANSWER: We imputed identity-by-state (IBS) sharing of parental siblings to phase parental genotypes. WHAT IS KNOWN ALREADY: Genome-wide haplotyping of preimplantation embryos is being implemented as a generic approach for genetic diagnosis of inherited single-gene disorders. To enable the phasing of genotypes into haplotypes, genotyping the direct family members of the prospective parent carrying the mutation is required. Current approaches require genotypes of either (i) both or one of the parents of the affected prospective parent or (ii) an affected or an unaffected child of the couple. However, this approach cannot be used when parents or children are not attainable, prompting an investigation into alternative phasing options. STUDY DESIGN, SIZE, DURATION: This is a retrospective validation study, which applied IBS-based phasing of parental haplotypes in 56 embryos derived from 12 PGT families. Genome-wide haplotypes and copy number profiles generated for each embryo using the new phasing approach were compared with the reference PGT method to evaluate the diagnostic concordance. PARTICIPANTS/MATERIALS, SETTING, METHODS: This study included 12 couples with a known hereditary genetic disorder, participating in the comprehensive PGT program and with at least one parental sibling available (e.g. brother and/or sister). Genotyping data from both prospective parents and the parental sibling(s) were used to perform IBS-based phasing and to trace the disease-associated alleles. The outcome of the IBS-based PGT was compared with the results of the clinically implemented reference haplotyping-based PGT method. MAIN RESULTS AND THE ROLE OF CHANCE: IBS-based haplotyping was performed for 12 PGT families. In accordance with the theoretical prediction of allele sharing between sibling pairs, 6 out of 12 (50%) couples or 23 out of 56 embryos could be phased using parental siblings. In families where phasing was possible, haplotype calling in the locus of interest was 100% concordant between the reference PGT method and IBS-based approach using parental siblings. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Phasing of parental haplotypes will only be possible when the disease locus lies in an informative region (categorized as IBS1). Phasing prospective parents using relatives with reduced genetic relatedness as a reference (e.g. siblings) decreases the size and the occurrence of informative IBS1 regions, necessary for haplotype calling. By including more than one extended family member, the chance of obtaining IBS1 coverage in the interrogated locus can be increased. A pre-PGT work-up can define whether the carrier couple could benefit from this approach. WIDER IMPLICATIONS OF THE FINDINGS: Phasing by relatives extends the potential of comprehensive PGT, since it allows the inclusion of couples who do not have access to the standard phasing references, such as parents or offspring. STUDY FUNDING/COMPETING INTEREST(S): The study was funded by the KU Leuven grant (C14/18/092), Research Foundation Flanders (FWO; GA09311N), Horizon 2020 innovation programme (WIDENLIFE, 692065) and Agilent Technologies. J.R.V., T.V. and M.Z.E. are co-inventors of a patent ZL910050-PCT/EP2011/060211-WO/2011/157846 'Methods for haplotyping single-cells' and ZL913096-PCT/EP2014/068315-WO/2015/028576 'Haplotyping and copy number typing using polymorphic variant allelic frequencies' licensed to Agilent Technologies. The other authors have no conflict of interest to declare.

Prenatally detected copy number variants in a national cohort: A postnatal follow-up study.

OBJECTIVE: Belgian genetic centers established a database containing data on all chromosomal microarrays performed in a prenatal context. A study was initiated to evaluate postnatal development in children diagnosed prenatally with a non-benign copy number variant (CNV). METHODS: All children diagnosed with a prenatally detected non-benign CNV in a Belgian genetic center between May 2013 and February 2015 were included in the patient population. The control population consisted of children who had undergone an invasive procedure during pregnancy, with no or only benign CNVs. Child development was evaluated at 36 months using three (3) questionnaires: Ages and Stages Questionnaire Third edition, Ages and Stages Questionnaire Social-Emotional Second Edition and a general questionnaire. RESULTS: A significant difference in communication and personal-social development was detected between children with a reported susceptibility CNV and both children with an unreported susceptibility CNV and the control population. The outcome of children with a particular CNV is discussed in a case-by-case manner. CONCLUSION: Our postnatal follow-up project of children with a prenatally detected non-benign CNV is the first nationwide project of its kind. A higher number of cases for each CNV category is however needed to confirm our findings.

Maternal copy-number variations in the DMD gene as secondary findings in noninvasive prenatal screening.

PURPOSE: Noninvasive prenatal screening (NIPS) using genome sequencing also reveals maternal copy-number variations (CNVs). Those CNVs can be clinically actionable or harmful to the fetus if inherited. CNVs in the DMD gene potentially causing dystrophinopathies are among the most commonly observed maternal CNVs. We present our experience with maternal DMD gene CNVs detected by NIPS. METHODS: We analyzed the data of maternal CNVs detected in the DMD gene revealed by NIPS. RESULTS: Of 26,123 NIPS analyses, 16 maternal CNVs in the DMD gene were detected (1/1632 pregnant women). Variant classification regarding pathogenicity and phenotypic severity was based on public databases, segregation analysis in the family, and prediction of the effect on the reading frame. Ten CNVs were classified as pathogenic, four as benign, and two remained unclassified. CONCLUSION: NIPS leverages CNV screening in the general population of pregnant women. We implemented a strategy for the interpretation and the return of maternal CNVs in the DMD gene detected by NIPS.

Genome-wide haplotyping embryos developing from 0PN and 1PN zygotes increases transferrable embryos in PGT-M.

STUDY QUESTION: Can genome-wide haplotyping increase success following preimplantation genetic testing for a monogenic disorder (PGT-M) by including zygotes with absence of pronuclei (0PN) or the presence of only one pronucleus (1PN)? SUMMARY ANSWER: Genome-wide haplotyping 0PNs and 1PNs increases the number of PGT-M cycles reaching embryo transfer (ET) by 81% and the live-birth rate by 75%. WHAT IS KNOWN ALREADY: Although a significant subset of 0PN and 1PN zygotes can develop into balanced, diploid and developmentally competent embryos, they are usually discarded because parental diploidy detection is not part of the routine work-up of PGT-M. STUDY DESIGN, SIZE, DURATION: This prospective cohort study evaluated the pronuclear number in 2229 zygotes from 2337 injected metaphase II (MII) oocytes in 268 cycles. PGT-M for 0PN and 1PN embryos developing into Day 5/6 blastocysts with adequate quality for vitrification was performed in 42 of the 268 cycles (15.7%). In these 42 cycles, we genome-wide haplotyped 216 good quality embryos corresponding to 49 0PNs, 15 1PNs and 152 2PNs. The reported outcomes include parental contribution to embryonic ploidy, embryonic aneuploidy, genetic diagnosis for the monogenic disorder, cycles reaching ETs, pregnancy and live birth rates (LBR) for unaffected offspring. PARTICIPANTS/MATERIALS, SETTING, METHODS: Blastomere DNA was whole-genome amplified and hybridized on the Illumina Human CytoSNP12V2.1.1 BeadChip arrays. Subsequently, genome-wide haplotyping and copy-number profiling was applied to investigate the embryonic genome architecture. Bi-parental, unaffected embryos were transferred regardless of their initial zygotic PN score. MAIN RESULTS AND THE ROLE OF CHANCE: A staggering 75.51% of 0PN and 42.86% of 1PN blastocysts are diploid bi-parental allowing accurate genetic diagnosis for the monogenic disorder. In total, 31% (13/42) of the PGT-M cycles reached ET or could repeat ET with an unaffected 0PN or 1PN embryo. The LBR per initiated cycle increased from 9.52 to 16.67%. LIMITATIONS, REASONS FOR CAUTION: The clinical efficacy of the routine inclusion of 0PN and 1PN zygotes in PGT-M cycles should be confirmed in larger cohorts from multicenter studies. WIDER IMPLICATIONS OF THE FINDINGS: Genome-wide haplotyping allows the inclusion of 0PN and 1PN embryos and subsequently increases the cycles reaching ET following PGT-M and potentially PGT for aneuploidy (PGT-A) and chromosomal structural rearrangements (PGT-SR). Establishing measures of clinical efficacy could lead to an update of the ESHRE guidelines which advise against the use of these zygotes. STUDY FUNDING/COMPETING INTEREST(S): SymBioSys (PFV/10/016 and C1/018 to J.R.V. and T.V.), the Horizon 2020 WIDENLIFE: 692065 to J.R.V., T.V., E.D., A.D. and M.Z.E. M.Z.E., T.V. and J.R.V. co-invented haplarithmisis ('Haplotyping and copy-number typing using polymorphic variant allelic frequencies'), which has been licensed to Agilent Technologies. H.M. is fully supported by the (FWO) (ZKD1543-ASP/16). The authors have no competing interests to declare.

Maternal liver transplant: Another cause of discordant fetal sex determination using cell-free DNA.

Noninvasive prenatal testing (NIPT) can very accurately determine fetal sex during pregnancy. We present an exceptional case where NIPT contradicts the ultrasound-based sex determination. The pregnant woman was recipient of a liver transplant from a male donor. Graft-derived cell-free DNA released into the maternal circulation clouded the NIPT-based sex determination. Hence, NIPT is not advisable when the pregnant mother underwent an organ transplant.

Predicting fetoplacental chromosomal mosaicism during non-invasive prenatal testing.

OBJECTIVE: Non-invasive prenatal detection of aneuploidies can be achieved with high accuracy through sequencing of cell-free maternal plasma DNA in the maternal blood plasma. However, false positive and negative non-invasive prenatal testing (NIPT) results remain. Fetoplacental mosaicism is the main cause for false positive and false negative NIPT. We set out to develop a method to detect placental chromosomal mosaicism via genome-wide circulating cell-free maternal plasma DNA screening. METHOD: Aneuploidy detection was combined with fetal fraction determination to enable the detection of placental mosaicism. This pipeline was applied to whole genome sequencing data derived from 19 735 plasma samples. Following an abnormal NIPT, test results were validated by conventional invasive prenatal or postnatal genetic testing. RESULTS: Respectively 3.2% (5/154), 12.8% (5/39), and 13.3% (2/15) of trisomies 21, 18, and 13 were predicted and confirmed to be mosaic. The incidence of other, rare autosomal trisomies was ~0.3% (58/19,735), 45 of which were predicted to be mosaic. Twin pregnancies with discordant fetal genotypes were predicted and confirmed. CONCLUSION: This approach permits the non-invasive detection of fetal autosomal aneuploidies and identifies pregnancies with a high risk of fetoplacental mosaicism. Knowledge about the presence of chromosomal mosaicism in the placenta influences risk estimation, genetic counseling, and improves prenatal management.

The Belgian MicroArray Prenatal (BEMAPRE) database: A systematic nationwide repository of fetal genomic aberrations.

OBJECTIVE: With the replacement of karyotyping by chromosomal microarray (CMA) in invasive prenatal diagnosis, new challenges have arisen. By building a national database, we standardize the classification and reporting of prenatally detected copy number variants (CNVs) across Belgian genetic centers. This database, which will link genetic and ultrasound findings with postnatal development, forms a unique resource to investigate the pathogenicity of variants of uncertain significance and to refine the phenotypic spectrum of pathogenic and susceptibility CNVs. METHODS: The Belgian MicroArray Prenatal (BEMAPRE) consortium is a collaboration of all genetic centers in Belgium. We collected data from all invasive prenatal procedures performed between May 2013 and July 2016. RESULTS: In this three-year period, 13 266 prenatal CMAs were performed. By national agreement, a limited number of susceptibility CNVs and no variants of uncertain significance were reported. Added values for using CMA versus conventional karyotyping were 1.8% in the general invasive population and 2.7% in cases with an ultrasound anomaly. Of the reported CNVs, 31.5% would have remained undetected with non-invasive prenatal test as the first-tier test. CONCLUSION: The establishment of a national database for prenatal CNV data allows for a uniform reporting policy and the investigation of the prenatal and postnatal genotype-phenotype correlation.

Accuracy and clinical value of maternal incidental findings during noninvasive prenatal testing for fetal aneuploidies.

PURPOSE: Genome-wide sequencing of cell-free (cf)DNA of pregnant women aims to detect fetal chromosomal imbalances. Because the largest fraction of cfDNA consists of maternal rather than fetal DNA fragments, maternally derived copy-number variants (CNVs) are also measured. Despite their potential clinical relevance, current analyses do not interpret maternal CNVs. Here, we explore the accuracy and clinical value of maternal CNV analysis. METHODS: Noninvasive prenatal testing was performed by whole-genome shotgun sequencing on plasma samples. Following mapping of the sequencing reads, the landscape of maternal CNVs was charted for 9,882 women using SeqCBS analysis. Recurrent CNVs were validated retrospectively by comparing their incidence with published reports. Nonrecurrent CNVs were prospectively confirmed by array comparative genomic hybridization or fluorescent in situ hybridization analysis on maternal lymphocytes. RESULTS: Consistent with population estimates, 10% nonrecurrent and 0.4% susceptibility CNVs for low-penetrant genomic disorders were identified. Five clinically actionable variants were reported to the pregnant women, including haploinsufficiency of RUNX1, a mosaicism for segmental chromosome 13 deletion, an unbalanced translocation, and two interstitial chromosome X deletions. CONCLUSION: Shotgun sequencing of cfDNA not only enables the detection of fetal aneuploidies but also reveals the presence of maternal CNVs. Some of those variants are clinically actionable or could potentially be harmful for the fetus. Interrogating the maternal CNV landscape can improve overall pregnancy management, and we propose reporting those variants if clinically relevant. The identification and reporting of such CNVs pose novel counseling dilemmas that warrant further discussions and development of societal guidelines.Genet Med 19 3, 306-313.

A prospective study of the clinical utility of prenatal chromosomal microarray analysis in fetuses with ultrasound abnormalities and an exploration of a framework for reporting unclassified variants and risk factors.

PURPOSE: To evaluate the clinical utility of chromosomal microarrays for prenatal diagnosis by a prospective study of fetuses with abnormalities detected on ultrasound. METHODS: Patients referred for prenatal diagnosis due to ultrasound anomalies underwent analysis by array comparative genomic hybridization as the first-tier diagnostic test. RESULTS: A total of 383 prenatal samples underwent analysis by array comparative genomic hybridization. Array analysis revealed causal imbalances in a total of 9.6% of patients (n = 37). Submicroscopic copy-number variations were detected in 2.6% of patients (n = 10/37), and arrays added valuable information over conventional karyotyping in 3.9% of patients (n = 15/37). We highlight a novel advantage of arrays; a 500-kb paternal insertional translocation is the likely driver of a de novo unbalanced translocation, thus improving recurrence risk calculation in this family. Variants of uncertain significance were revealed in 1.6% of patients (n = 6/383). CONCLUSION: We demonstrate the added value of chromosomal microarrays for prenatal diagnosis in the presence of ultrasound anomalies. We advocate reporting back only copy-number variations with known pathogenic significance. Although this approach might be considered opposite to the ideal of full reproductive autonomy of the parents, we argue why providing all information to parents may result in a false sense of autonomy.