Systematic evaluation of genome sequencing for the diagnostic assessment of autism spectrum disorder and fetal structural anomalies.

Short-read genome sequencing (GS) holds the promise of becoming the primary diagnostic approach for the assessment of autism spectrum disorder (ASD) and fetal structural anomalies (FSAs). However, few studies have comprehensively evaluated its performance against current standard-of-care diagnostic tests: karyotype, chromosomal microarray (CMA), and exome sequencing (ES). To assess the clinical utility of GS, we compared its diagnostic yield against these three tests in 1,612 quartet families including an individual with ASD and in 295 prenatal families. Our GS analytic framework identified a diagnostic variant in 7.8% of ASD probands, almost 2-fold more than CMA (4.3%) and 3-fold more than ES (2.7%). However, when we systematically captured copy-number variants (CNVs) from the exome data, the diagnostic yield of ES (7.4%) was brought much closer to, but did not surpass, GS. Similarly, we estimated that GS could achieve an overall diagnostic yield of 46.1% in unselected FSAs, representing a 17.2% increased yield over karyotype, 14.1% over CMA, and 4.1% over ES with CNV calling or 36.1% increase without CNV discovery. Overall, GS provided an added diagnostic yield of 0.4% and 0.8% beyond the combination of all three standard-of-care tests in ASD and FSAs, respectively. This corresponded to nine GS unique diagnostic variants, including sequence variants in exons not captured by ES, structural variants (SVs) inaccessible to existing standard-of-care tests, and SVs where the resolution of GS changed variant classification. Overall, this large-scale evaluation demonstrated that GS significantly outperforms each individual standard-of-care test while also outperforming the combination of all three tests, thus warranting consideration as the first-tier diagnostic approach for the assessment of ASD and FSAs.

Amniocentesis in pregnancies at or beyond 24 weeks: an international multicenter study.

BACKGROUND: Amniocentesis for genetic diagnosis is most commonly done between 15 and 22 weeks of gestation but can be performed at later gestational ages. The safety and genetic diagnostic accuracy of amniocentesis have been well-established through numerous large-scale multicenter studies for procedures before 24 weeks, but comprehensive data on late amniocentesis remain sparse. OBJECTIVE: To evaluate the indications, diagnostic yield, safety, and maternal and fetal outcomes associated with amniocentesis performed at or beyond 24 weeks of gestation. STUDY DESIGN: We conducted an international multicenter retrospective cohort study examining pregnant individuals who underwent amniocentesis for prenatal diagnostic testing at gestational ages between 24w0d and 36w6d. The study, spanning from 2011 to 2022, involved 9 referral centers. We included singleton or twin pregnancies with documented outcomes, excluding cases where other invasive procedures were performed during pregnancy or if amniocentesis was conducted for obstetric indications. We analyzed indications for late amniocentesis, types of genetic tests performed, their results, and the diagnostic yield, along with pregnancy outcomes and postprocedure complications. RESULTS: Of the 752 pregnant individuals included in our study, late amniocentesis was primarily performed for the prenatal diagnosis of structural anomalies (91.6%), followed by suspected fetal infection (2.3%) and high-risk findings from cell-free DNA screening (1.9%). The median gestational age at the time of the procedure was 28w5d, and 98.3% of pregnant individuals received results of genetic testing before birth or pregnancy termination. The diagnostic yield was 22.9%, and a diagnosis was made 2.4 times more often for fetuses with anomalies in multiple organ systems (36.4%) compared to those with anomalies in a single organ system (15.3%). Additionally, the diagnostic yield varied depending on the specific organ system involved, with the highest yield for musculoskeletal anomalies (36.7%) and hydrops fetalis (36.4%) when a single organ system or entity was affected. The most prevalent genetic diagnoses were aneuploidies (46.8%), followed by copy number variants (26.3%) and monogenic disorders (22.2%). The median gestational age at delivery was 38w3d, with an average of 59 days between the procedure and delivery date. The overall complication rate within 2 weeks postprocedure was 1.2%. We found no significant difference in the rate of preterm delivery between pregnant individuals undergoing amniocentesis between 24 and 28 weeks and those between 28 and 32 weeks, reinforcing the procedure's safety across these gestational periods. CONCLUSION: Late amniocentesis, at or after 24 weeks of gestation, especially for pregnancies complicated by multiple congenital anomalies, has a high diagnostic yield and a low complication rate, underscoring its clinical utility. It provides pregnant individuals and their providers with a comprehensive diagnostic evaluation and results before delivery, enabling informed counseling and optimized perinatal and neonatal care planning.

The expanded spectrum of human disease associated with GREB1L likely includes complex congenital heart disease.

OBJECTIVE: GREB1L has been linked prenatally to Potter's sequence, as well as less severe anomalies of the kidney, uterus, inner ear, and heart. The full phenotypic spectrum is unknown. The purpose of this study was to characterize known and novel pre- and postnatal phenotypes associated with GREB1L. METHODS: We solicited cases from the Fetal Sequencing Consortium, screened a population-based genomic database, and conducted a comprehensive literature search to identify disease cases associated with GREB1L. We present a detailed phenotypic spectrum and molecular changes. RESULTS: One hundred twenty-seven individuals with 51 unique pathogenic or likely pathogenic GREB1L variants were identified. 24 (47%) variants were associated with isolated kidney anomalies, 19 (37%) with anomalies of multiple systems, including one case of hypoplastic left heart syndrome, five (10%) with isolated sensorineural hearing loss, two (4%) with isolated uterine agenesis; and one (2%) with isolated tetralogy of Fallot. CONCLUSION: GREB1L may cause complex congenital heart disease (CHD) in humans. Clinicians should consider GREB1L testing in the setting of CHD, and cardiac screening in the setting of GREB1L variants.

Processing the process: Reflections on genetic counselor-led student supervision groups and practical tips for future facilitators.

Genetic counseling graduate students face growth and challenges across a variety of axes both personally and professionally throughout their training. The formation of leader-led supervision groups for second-year genetic counseling students has created a safe space for students to give and receive feedback, process their positionality in complex clinical scenarios and the medical system at large, dissect psychosocial counseling theory, and share personal and professional experiences with the overall aim of supporting their growth. This work requires faculty facilitators who are invested in student growth and operate from a framework of empathy, humanism, curiosity, and vulnerability. The authors share their reflections on stepping into the facilitator role with no prior experience other than their work in clinical genetic counseling in varied practice settings. Common themes across four cohorts of students are presented along with reflections on facilitator growth, consideration of student developmental stages, and the parallel process between providing clinical services to clients and educating students. The authors hope to highlight the value of processing cases, inspire other genetic counselors to engage in this work, and normalize the experiences of those already running genetic counseling student process groups.

Mouse and human studies support DSTYK loss of function as a low-penetrance and variable expressivity risk factor for congenital urinary tract anomalies.

PURPOSE: Previous work identified rare variants in DSTYK associated with human congenital anomalies of the kidney and urinary tract (CAKUT). Here, we present a series of mouse and human studies to clarify the association, penetrance, and expressivity of DSTYK variants. METHODS: We phenotypically characterized Dstyk knockout mice of 3 separate inbred backgrounds and re-analyzed the original family segregating the DSTYK c.654+1G>A splice-site variant (referred to as "SSV" below). DSTYK loss of function (LOF) and SSVs were annotated in individuals with CAKUT, epilepsy, or amyotrophic lateral sclerosis vs controls. A phenome-wide association study analysis was also performed using United Kingdom Biobank (UKBB) data. RESULTS: Results demonstrate ∼20% to 25% penetrance of obstructive uropathy, at least, in C57BL/6J and FVB/NJ Dstyk-/- mice. Phenotypic penetrance increased to ∼40% in C3H/HeJ mutants, with mild-to-moderate severity. Re-analysis of the original family segregating the rare SSV showed low penetrance (43.8%) and no alternative genetic causes for CAKUT. LOF DSTYK variants burden showed significant excess for CAKUT and epilepsy vs controls and an exploratory phenome-wide association study supported association with neurological disorders. CONCLUSION: These data support causality for DSTYK LOF variants and highlights the need for large-scale sequencing studies (here >200,000 cases) to accurately assess causality for genes and variants to lowly penetrant traits with common population prevalence.

Prenatal phenotyping: A community effort to enhance the Human Phenotype Ontology.

Technological advances in both genome sequencing and prenatal imaging are increasing our ability to accurately recognize and diagnose Mendelian conditions prenatally. Phenotype-driven early genetic diagnosis of fetal genetic disease can help to strategize treatment options and clinical preventive measures during the perinatal period, to plan in utero therapies, and to inform parental decision-making. Fetal phenotypes of genetic diseases are often unique and at present are not well understood; more comprehensive knowledge about prenatal phenotypes and computational resources have an enormous potential to improve diagnostics and translational research. The Human Phenotype Ontology (HPO) has been widely used to support diagnostics and translational research in human genetics. To better support prenatal usage, the HPO consortium conducted a series of workshops with a group of domain experts in a variety of medical specialties, diagnostic techniques, as well as diseases and phenotypes related to prenatal medicine, including perinatal pathology, musculoskeletal anomalies, neurology, medical genetics, hydrops fetalis, craniofacial malformations, cardiology, neonatal-perinatal medicine, fetal medicine, placental pathology, prenatal imaging, and bioinformatics. We expanded the representation of prenatal phenotypes in HPO by adding 95 new phenotype terms under the Abnormality of prenatal development or birth (HP:0001197) grouping term, and revised definitions, synonyms, and disease annotations for most of the 152 terms that existed before the beginning of this effort. The expansion of prenatal phenotypes in HPO will support phenotype-driven prenatal exome and genome sequencing for precision genetic diagnostics of rare diseases to support prenatal care.

Diagnostic sequencing to support genetically stratified medicine in a tertiary care setting.

PURPOSE: The goal of stratified medicine is to identify subgroups of patients with similar disease mechanisms and specific responses to treatments. To prepare for stratified clinical trials, genome-wide genetic analysis should occur across clinical areas to identify undiagnosed genetic diseases and new genetic causes of disease. METHODS: To advance genetically stratified medicine, we have developed and implemented broad exome sequencing infrastructure and research protocols at Columbia University Irving Medical Center/NewYork-Presbyterian Hospital. RESULTS: We enrolled 4889 adult and pediatric probands and identified a primary result in 572 probands. The cohort was phenotypically and demographically heterogeneous because enrollment occurred across multiple specialty clinics (eg, epilepsy, nephrology, fetal anomaly). New gene-disease associations and phenotypic expansions were discovered across clinical specialties. CONCLUSION: Our study processes have enabled the enrollment and exome sequencing/analysis of a phenotypically and demographically diverse cohort of patients within 1 tertiary care medical center. Because all genomic data are stored centrally with permission for longitudinal access to the electronic medical record, subjects can be recontacted with updated genetic diagnoses or for participation in future genotype-based clinical trials. This infrastructure has allowed for the promotion of genetically stratified clinical trial readiness within the Columbia University Irving Medical Center/NewYork-Presbyterian Hospital health care system.

Clinical genetic counselor experience in the adoption of telehealth in the United States and Canada during the COVID-19 pandemic.

The COVID-19 pandemic has significantly impacted the service delivery model (SDM) of clinical genetic counseling across the United States and Canada. A cross-sectional survey was distributed to 4,956 genetic counselors (GCs) from the American Board of Genetic Counselors and Canadian Association of Genetic Counselors mailing lists in August 2020 to assess the change in utilization of telehealth for clinical genetic counseling during the COVID-19 pandemic compared with prior to the pandemic. Data from 411 eligible clinical genetic counselors on GC attitudes and their experiences prior to and during the pandemic were collected and analyzed to explore the change in SDM, change in appointment characteristics, change in billing practices, GC perceived benefits and limitations of telehealth, and prediction of future trends in SDM in the post-pandemic era. The study showed the overall utilization of audiovisual and telephone encounters increased by 43.4% and 26.2%, respectively. The majority of respondents who provided audiovisual and telephone encounters reported increased patient volume compared with prior to the pandemic, with an average increase of 79.4% and 42.8%, respectively. There was an increase of 69.4% of GCs rendering genetic services from home offices. The percentage of participants who billed for telehealth services increased from 45.7% before the pandemic to 80.3% during the pandemic. The top GC perceived benefits of telehealth included safety for high-risk COVID patients (95.2%) and saved commute time for patients (94.7%). The top GC perceived limitations of telehealth included difficulty to conduct physician evaluation/coordinating with healthcare providers (HCP) (73.7%) and difficulty addressing non-English speaking patients (68.5%). Overall, 89.6% of GCs were satisfied with telehealth; however, 55.3% reported uncertainty whether the newly adopted SDM would continue after the pandemic subsides. Results from this study demonstrate the rapid adoption of telehealth for clinical genetic counseling services as a result of the COVID-19 pandemic, an increase in billing for these services, and support the feasibility of telehealth for genetic counseling as a longer term solution to reach patients who are geographically distant.

Whole-exome sequencing in the evaluation of fetal structural anomalies: a prospective cohort study.

BACKGROUND: Identification of chromosomal aneuploidies and copy number variants that are associated with fetal structural anomalies has substantial value. Although whole-exome sequencing (WES) has been applied to case series of a few selected prenatal cases, its value in routine clinical settings has not been prospectively assessed in a large unselected cohort of fetuses with structural anomalies. We therefore aimed to determine the incremental diagnostic yield (ie, the added value) of WES following uninformative results of standard investigations with karyotype testing and chromosomal microarray in an unselected cohort of sequential pregnancies showing fetal structural anomalies. METHODS: In this prospective cohort study, the parents of fetuses who were found to have a structural anomaly in a prenatal ultrasound were screened for possible participation in the study. These participants were predominantly identified in or were referred to the Columbia University Carmen and John Thain Center for Prenatal Pediatrics (New York, NY, USA). Fetuses with confirmed aneuploidy or a causal pathogenic copy number variant were excluded from WES analyses. By use of WES of the fetuses and parents (parent-fetus trios), we identified genetic variants that indicated an underlying cause (diagnostic genetic variants) and genetic variants that met the criteria of bioinformatic signatures that had previously been described to be significantly enriched among diagnostic genetic variants. FINDINGS: Between April 24, 2015, and April 19, 2017, 517 sequentially identified pregnant women found to have fetuses with a structural anomaly were screened for their eligibility for inclusion in our study. 71 (14%) couples declined testing, 87 (17%) trios were missing at least one DNA sample (from either parent or the fetus), 69 (13%) trios had a clinically relevant abnormal karyotype or chromosomal microarray finding, 51 (10%) couples did not consent to WES or withdrew consent, and five (1%) samples were not of good enough quality for analysis. DNA samples from 234 (45%) eligible trios were therefore used for analysis of the primary outcome. By use of trio sequence data, we identified diagnostic genetic variants in 24 (10%) families. Mutations with bioinformatic signatures that were indicative of pathogenicity but with insufficient evidence to be considered diagnostic were also evaluated; 46 (20%) of the 234 fetuses assessed were found to have such signatures. INTERPRETATION: Our analysis of WES data in a prospective cohort of unselected fetuses with structural anomalies shows the value added by WES following the use of routine genetic tests. Our findings suggest that, in cases of fetal anomalies in which assessment with karyotype testing and chromosomal microarray fail to determine the underlying cause of a structural anomaly, WES can add clinically relevant information that could assist current management of a pregnancy. The unique challenges of WES-based prenatal diagnostics require analysis by a multidisciplinary team of perinatal practitioners and laboratory specialists. FUNDING: Institute for Genomic Medicine (Columbia University Irving Medical Center).

Genomics of stillbirth.

Stillbirth, defined as fetal death at 20 weeks gestation or later, is a devastating pregnancy outcome affecting 1 in 175 pregnancies in the United States. Although efforts to understand the etiology of stillbirth have expanded, 25 % of cases remain unexplained and some cases previously thought to be explained may have additional unknown causative factors. Determining an etiology for stillbirth is important for clinical management and for grieving families to obtain closure, to find meaning, and to understand recurrence risks. However, the evaluation of stillbirth is not completed uniformly despite American College of Obstetrics and Gynecology (ACOG) guidelines and stillbirth data is frequently incomplete due to lack of genomic analysis, fetal autopsy, and placental pathology. Karyotype and chromosomal microarray have been the gold standard in genetic analysis in perinatal medicine for many years, but next generation sequencing holds promise towards improving diagnostic yields and providing clarity for both clinicians and patients.

TMEM218 dysfunction causes ciliopathies, including Joubert and Meckel syndromes.

The Joubert-Meckel syndrome spectrum is a continuum of recessive ciliopathy conditions caused by primary cilium dysfunction. The primary cilium is a microtubule-based, antenna-like organelle that projects from the surface of most human cell types, allowing them to respond to extracellular signals. The cilium is partitioned from the cell body by the transition zone, a known hotspot for ciliopathy-related proteins. Despite years of Joubert syndrome (JBTS) gene discovery, the genetic cause cannot be identified in up to 30% of individuals with JBTS, depending on the cohort, sequencing method, and criteria for pathogenic variants. Using exome and targeted sequencing of 655 families with JBTS, we identified three individuals from two families harboring biallelic, rare, predicted-deleterious missense TMEM218 variants. Via MatchMaker Exchange, we identified biallelic TMEM218 variants in four additional families with ciliopathy phenotypes. Of note, four of the six families carry missense variants affecting the same highly conserved amino acid position 115. Clinical features included the molar tooth sign (N = 2), occipital encephalocele (N = 5, all fetuses), retinal dystrophy (N = 4, all living individuals), polycystic kidneys (N = 2), and polydactyly (N = 2), without liver involvement. Combined with existing functional data linking TMEM218 to ciliary transition zone function, our human genetic data make a strong case for TMEM218 dysfunction as a cause of ciliopathy phenotypes including JBTS with retinal dystrophy and Meckel syndrome. Identifying all genetic causes of the Joubert-Meckel spectrum enables diagnostic testing, prognostic and recurrence risk counseling, and medical monitoring, as well as work to delineate the underlying biological mechanisms and identify targets for future therapies.

Clinical whole exome sequencing from dried blood spot identifies novel genetic defect underlying asparagine synthetase deficiency.

We add two novel variants to the existing mutation spectrum of ASNS gene. Loss of ASNS function should be suspected in newborns presenting with congenital microcephaly, intellectual disability, progressive cerebral atrophy, and intractable seizures. Acquisition and sequencing of stored newborn blood spot can be a valuable option when no biological samples are available from a deceased child.