Rare variants in NR2F2 cause congenital heart defects in humans.

Congenital heart defects (CHDs) are the most common birth defect worldwide and are a leading cause of neonatal mortality. Nonsyndromic atrioventricular septal defects (AVSDs) are an important subtype of CHDs for which the genetic architecture is poorly understood. We performed exome sequencing in 13 parent-offspring trios and 112 unrelated individuals with nonsyndromic AVSDs and identified five rare missense variants (two of which arose de novo) in the highly conserved gene NR2F2, a very significant enrichment (p = 7.7 × 10(-7)) compared to 5,194 control subjects. We identified three additional CHD-affected families with other variants in NR2F2 including a de novo balanced chromosomal translocation, a de novo substitution disrupting a splice donor site, and a 3 bp duplication that cosegregated in a multiplex family. NR2F2 encodes a pleiotropic developmental transcription factor, and decreased dosage of NR2F2 in mice has been shown to result in abnormal development of atrioventricular septa. Via luciferase assays, we showed that all six coding sequence variants observed in individuals significantly alter the activity of NR2F2 on target promoters.

Outcome after transcatheter occlusion of patent ductus arteriosus in infants less than 6 kg: A national study from United Kingdom and Ireland.

OBJECTIVES: This study aimed to report our national experience with transcatheter patent ductus arteriosus (PDA) occlusion in infants weighing <6 kg. BACKGROUND: The technique of transcatheter PDA closure has evolved in the past two decades and is increasingly used in smaller patients but data on safety and efficacy are limited. METHODS: Patients weighing < 6 kg in whom transcatheter PDA occlusion was attempted in 13 tertiary paediatric cardiology units in the United Kingdom and Ireland were retrospectively analyzed to review the outcome and complications. RESULTS: A total of 408 patients underwent attempted transcatheter PDA closure between January 2004 and December 2014. The mean weight at catheterization was 4.9 ± 1.0 kg and mean age was 5.7 ± 3.0 months. Successful device implantation was achieved in 374 (92%) patients without major complication and of these, complete occlusion was achieved in 356 (95%) patients at last available follow-up. Device embolization occurred in 20 cases (5%). The incidence of device related obstruction to the left pulmonary artery or aorta and access related peripheral vascular injury were low. There were no deaths related to the procedure. CONCLUSIONS: Transcatheter closure of PDA can be accomplished in selected infants weighing <6 kg despite the manufacturer's recommended weight limit of 6 kg for most ductal occluders. The embolization rate is higher than previously reported in larger patients. Retrievability of the occluder and duct morphology needs careful consideration before deciding whether surgical ligation or transcatheter therapy is the better treatment option.

Exome sequencing identifies rare variants in multiple genes in atrioventricular septal defect.

PURPOSE: The genetic etiology of atrioventricular septal defect (AVSD) is unknown in 40% cases. Conventional sequencing and arrays have identified the etiology in only a minority of nonsyndromic individuals with AVSD. METHODS: Whole-exome sequencing was performed in 81 unrelated probands with AVSD to identify potentially causal variants in a comprehensive set of 112 genes with strong biological relevance to AVSD. RESULTS: A significant enrichment of rare and rare damaging variants was identified in the gene set, compared with controls (odds ratio (OR): 1.52; 95% confidence interval (CI): 1.35-1.71; P = 4.8 × 10(-11)). The enrichment was specific to AVSD probands, compared with a cohort without AVSD with tetralogy of Fallot (OR: 2.25; 95% CI: 1.84-2.76; P = 2.2 × 10(-16)). Six genes (NIPBL, CHD7, CEP152, BMPR1a, ZFPM2, and MDM4) were enriched for rare variants in AVSD compared with controls, including three syndrome-associated genes (NIPBL, CHD7, and CEP152). The findings were confirmed in a replication cohort of 81 AVSD probands. CONCLUSION: Mutations in genes with strong biological relevance to AVSD, including syndrome-associated genes, can contribute to AVSD, even in those with isolated heart disease. The identification of a gene set associated with AVSD will facilitate targeted genetic screening in this cohort.

Targeted Next-Generation Sequencing Analysis of 1,000 Individuals with Intellectual Disability.

To identify genetic causes of intellectual disability (ID), we screened a cohort of 986 individuals with moderate to severe ID for variants in 565 known or candidate ID-associated genes using targeted next-generation sequencing. Likely pathogenic rare variants were found in ∼11% of the cases (113 variants in 107/986 individuals: ∼8% of the individuals had a likely pathogenic loss-of-function [LoF] variant, whereas ∼3% had a known pathogenic missense variant). Variants in SETD5, ATRX, CUL4B, MECP2, and ARID1B were the most common causes of ID. This study assessed the value of sequencing a cohort of probands to provide a molecular diagnosis of ID, without the availability of DNA from both parents for de novo sequence analysis. This modeling is clinically relevant as 28% of all UK families with dependent children are single parent households. In conclusion, to diagnose patients with ID in the absence of parental DNA, we recommend investigation of all LoF variants in known genes that cause ID and assessment of a limited list of proven pathogenic missense variants in these genes. This will provide 11% additional diagnostic yield beyond the 10%-15% yield from array CGH alone.

Genome-wide association study identifies loci on 12q24 and 13q32 associated with tetralogy of Fallot.

We conducted a genome-wide association study to search for risk alleles associated with Tetralogy of Fallot (TOF), using a northern European discovery set of 835 cases and 5159 controls. A region on chromosome 12q24 was associated (P = 1.4 × 10(-7)) and replicated convincingly (P = 3.9 × 10(-5)) in 798 cases and 2931 controls [per allele odds ratio (OR) = 1.27 in replication cohort, P = 7.7 × 10(-11) in combined populations]. Single nucleotide polymorphisms in the glypican 5 gene on chromosome 13q32 were also associated (P = 1.7 × 10(-7)) and replicated convincingly (P = 1.2 × 10(-5)) in 789 cases and 2927 controls (per allele OR = 1.31 in replication cohort, P = 3.03 × 10(-11) in combined populations). Four additional regions on chromosomes 10, 15 and 16 showed suggestive association accompanied by nominal replication. This study, the first genome-wide association study of a congenital heart malformation phenotype, provides evidence that common genetic variation influences the risk of TOF.

Contribution of global rare copy-number variants to the risk of sporadic congenital heart disease.

Previous studies have shown that copy-number variants (CNVs) contribute to the risk of complex developmental phenotypes. However, the contribution of global CNV burden to the risk of sporadic congenital heart disease (CHD) remains incompletely defined. We generated genome-wide CNV data by using Illumina 660W-Quad SNP arrays in 2,256 individuals with CHD, 283 trio CHD-affected families, and 1,538 controls. We found association of rare genic deletions with CHD risk (odds ratio [OR] = 1.8, p = 0.0008). Rare deletions in study participants with CHD had higher gene content (p = 0.001) with higher haploinsufficiency scores (p = 0.03) than they did in controls, and they were enriched with Wnt-signaling genes (p = 1 × 10(-5)). Recurrent 15q11.2 deletions were associated with CHD risk (OR = 8.2, p = 0.02). Rare de novo CNVs were observed in ~5% of CHD trios; 10 out of 11 occurred on the paternally transmitted chromosome (p = 0.01). Some of the rare de novo CNVs spanned genes known to be involved in heart development (e.g., HAND2 and GJA5). Rare genic deletions contribute ~4% of the population-attributable risk of sporadic CHD. Second to previously described CNVs at 1q21.1, deletions at 15q11.2 and those implicating Wnt signaling are the most significant contributors to the risk of sporadic CHD. Rare de novo CNVs identified in CHD trios exhibit paternal origin bias.

Phenotype-specific effect of chromosome 1q21.1 rearrangements and GJA5 duplications in 2436 congenital heart disease patients and 6760 controls.

Recurrent rearrangements of chromosome 1q21.1 that occur via non-allelic homologous recombination have been associated with variable phenotypes exhibiting incomplete penetrance, including congenital heart disease (CHD). However, the gene or genes within the ~1 Mb critical region responsible for each of the associated phenotypes remains unknown. We examined the 1q21.1 locus in 948 patients with tetralogy of Fallot (TOF), 1488 patients with other forms of CHD and 6760 ethnically matched controls using single nucleotide polymorphism genotyping arrays (Illumina 660W and Affymetrix 6.0) and multiplex ligation-dependent probe amplification. We found that duplication of 1q21.1 was more common in cases of TOF than in controls [odds ratio (OR) 30.9, 95% confidence interval (CI) 8.9-107.6); P = 2.2 × 10(-7)], but deletion was not. In contrast, deletion of 1q21.1 was more common in cases of non-TOF CHD than in controls [OR 5.5 (95% CI 1.4-22.0); P = 0.04] while duplication was not. We also detected rare (n = 3) 100-200 kb duplications within the critical region of 1q21.1 in cases of TOF. These small duplications encompassed a single gene in common, GJA5, and were enriched in cases of TOF in comparison to controls [OR = 10.7 (95% CI 1.8-64.3), P = 0.01]. These findings show that duplication and deletion at chromosome 1q21.1 exhibit a degree of phenotypic specificity in CHD, and implicate GJA5 as the gene responsible for the CHD phenotypes observed with copy number imbalances at this locus.

Low-frequency intermediate penetrance variants in the ROCK1 gene predispose to Tetralogy of Fallot.

BACKGROUND: Epidemiological studies indicate a substantial excess familial recurrence of non-syndromic Tetralogy of Fallot (TOF), implicating genetic factors that remain largely unknown. The Rho induced kinase 1 gene (ROCK1) is a key component of the planar cell polarity signalling pathway, which plays an important role in normal cardiac development. The aim of this study was to investigate the role of genetic variation in ROCK1 on the risk of TOF. RESULTS: ROCK1 was sequenced in a discovery cohort of 93 non-syndromic TOF probands to identify rare variants. TagSNPs were selected to capture commoner variation in ROCK1. Novel variants and TagSNPs were genotyped in a discovery cohort of 458 TOF cases and 1331 healthy controls, and positive findings were replicated in a further 209 TOF cases and 1290 healthy controls. Association between genotypes and TOF was assessed using LAMP.A rare SNP (c.807C > T; rs56085230) discovered by sequencing was associated with TOF risk (p = 0.006) in the discovery cohort. The variant was also significantly associated with the risk of TOF in the replication cohort (p = 0.018). In the combined cohorts the odds ratio for TOF was 2.61 (95% CI 1.58-4.30); p < 0.0001. The minor allele frequency of rs56085230 in the cases was 0.02, and in the controls it was 0.007. The variant accounted for 1% of the population attributable risk (PAR) of TOF. We also found significant association with TOF for an uncommon TagSNP in ROCK1, rs288979 (OR 1.64 [95% CI 1.15-2.30]; p = 1.5x10⁻5). The minor allele frequency of rs288979 in the controls was 0.043, and the variant accounted for 11% of the PAR of TOF. These association signals were independent of each other, providing additional internal validation of our result. CONCLUSIONS: Low frequency intermediate penetrance (LFIP) variants in the ROCK1 gene predispose to the risk of TOF.

Outcomes of manually modified microvascular plugs to pulmonary flow restrictors in various congenital heart lesions.

Background: The development of microvascular plugs (MVPs) has enabled novel transcatheter deliverable endoluminal pulmonary flow restrictors (PFRs) with the potential to treat newborns and infants with life-threatening congenital heart diseases (CHDs) in a minimally invasive manner. We present our experience to evaluate the efficacy of this concept in controlling pulmonary blood flow in various CHDs. Methods: Retrospective clinical data review of patients with CHD and pulmonary over-circulation who received bilateral PFRs percutaneously. Results: Twenty-eight PFRs (7 MVP-5Q, 12 MVP-7Q, and 9 MVP-9Q) were finally implanted in 14 patients with a median age of 1.6 months (IQR, 0.9-2.3) and a median weight of 3.1 Kg (IQR, 2.7-3.6). Nine patients had large intra-cardiac left-to-right shunts (including 3 with fatal trisomy and palliative programs), 2 had borderline left ventricles, 2 had Taussig-Bing anomaly, and one had a hypoplastic left heart. Four patients had concomitant ductal stenting. Two MVP-5Qs were snare-removed and upsized to MVP-7Q. Patients experienced a significant drop in oxygen saturation and Qp/Qs. All patients were discharged from the ICU after a median of 3.5 days (IQR, 2-5.8) postoperative. Five patients had routine inter-stage catheterization and no device embolization or pulmonary branch distortion was seen. Fourteen (50%) PFRs were surgically explanted uneventfully on a median of 4.3 months (IQR, 1.2-6) post-implantation during biventricular repair in 6 patients and stage-2 palliation in one patient. The latter died 1 month post-operative from severe sepsis. Four patients are scheduled for surgical PFR removal and biventricular repair. Two patients with trisomy 18 died at 1 and 6.8 months post-procedure from non-cardiac causes. One patient with trisomy 13 is alive at 2.7 months post-procedure. Conclusion: It is feasible to bespoke MVPs and implant them as effective PFRs in various CHDs. This approach enables staged left ventricular recruitment, comprehensive stage-2 or biventricular repair with lower risk by postponing surgeries to later infancy. Device explantation is uneventful, and the outcomes afterward are promising.

Maternal high-fat diet interacts with embryonic Cited2 genotype to reduce Pitx2c expression and enhance penetrance of left-right patterning defects.

Deficiency of the transcription factor Cited2 in mice results in cardiac malformation, adrenal agenesis, neural tube, placental defects and partially penetrant cardiopulmonary laterality defects resulting from an abnormal Nodal->Pitx2c pathway. Here we show that a maternal high-fat diet more than doubles the penetrance of laterality defects and, surprisingly, induces palatal clefting in Cited2-deficient embryos. Both maternal diet and Cited2 deletion reduce embryo weight and kidney and thymus volume. Expression profiling identified 40 embryonic transcripts including Pitx2 that were significantly affected by embryonic genotype-maternal diet interaction. We show that a high-fat diet reduces Pitx2c levels >2-fold in Cited2-deficient embryos. Taken together, these results define a novel interaction between maternal high-fat diet and embryonic Cited2 deficiency that affects Pitx2c expression and results in abnormal laterality. They suggest that appropriate modifications of maternal diet may prevent such defects in humans.

Echocardiographically guided catheter closure of arterial ducts in small preterm infants on the neonatal intensive care unit.

OBJECTIVE: To describe closure of haemodynamically significant arterial ducts in preterm infants using an echocardiographically guided cardiac catheter technique in selected infants in the neonatal nursery and in preference to cardiac surgery. BACKGROUND: Persistently patent arterial ducts are common in preterm infants and are associated with significant morbidity and mortality. Cardiac catheter techniques continue to improve and occlusion of arterial ducts in preterm infants is becoming technically feasible. Closure of arterial ducts by cardiac catheter techniques would enable selected infants to avoid surgery and a lateral thoracotomy, as well as potentially obviating the need for transfer of sick preterm infants between units for duct closure. METHODS AND RESULTS: This brief report describes placement of coils or Amplatzer duct devices to occlude arterial ducts in small premature infants exclusively under echocardiographic guidance in the Neonatal Intensive Care Unit. CONCLUSIONS: Closing arterial ducts in the neonatal nursery by an echocardiographically guided cardiac catheter technique with minimal morbidity is becoming achievable and is a significant advance in neonatal care.

The Ross-Konno operation for neonates and infants with severe aortic incompetence following treatment for critical aortic stenosis.

OBJECTIVES: Aortic valve stenosis in neonates and infants is associated with congestive cardiac failure, and balloon or surgical valvuloplasty provides relief of stenosis. Occasionally severe aortic insufficiency necessitates urgent aortic valve replacement. We reviewed our experience with the Ross-Konno procedure in patients <1 year. METHODS: Between October 2013 and May 2020, 36 patients underwent balloon (34) or surgical (2) aortic valvuloplasty for aortic stenosis. Six patients subsequently underwent a Ross-Konno procedure. The median age at operation was 55 (27-116) days and weight was 4.25 (2.5-12) kg. All patients were in severe cardiac failure and had a small aortic annulus with Z-score -3.1 (-1 to -4.4). RESULTS: There were no early or late deaths. At the latest follow-up at 39 (13-60) months, ventricular function had improved in all patients and no patient was on anti-failure medication. On echocardiography, there wasno more than trivial aortic regurgitation and no left ventricular outflow tract obstruction. One patient required right ventricle to pulmonary artery conduit replacement and one patient had homograft stenting. CONCLUSIONS: Despite the severe preoperative haemodynamic compromise, the urgent Ross-Konno procedure was associated with excellent operative survival and recovery of ventricular function. The need for reintervention to the pulmonary conduit remains a cause for concern.

Endovascular stent placement is an acceptable alternative to reoperation in selected infants with residual or recurrent aortic arch obstruction.

OBJECTIVE: To describe endovascular stent placement in infants as a technically feasible option in circumstances where surgery is considered less favorable. BACKGROUND: Endovascular stent placement has become established as a first line therapy for native coarctation of the aorta or recoarctation in older children where stents capable of expansion to adult size can be placed safely. Surgery remains the therapy of choice in infants and young children. The management of aortic arch obstruction in infants is, however, frequently complicated by complex anatomy or clinical condition that may make surgery or further surgery an unattractive option. There is little reported data and the implications thereof of transcatheter stent placement in aortic arch obstruction in infants. METHODS: Between August 2004 and November 2009, 11 patients had aortic arch obstruction treated with endovascular stent placement. The median age and weight at first stent placement was 46 days (range 3-399 days) and 4 kg (range 1.4-8 kg), respectively. In 10 patients, surgical intervention preceded transcatheter stent placement. Four had complex aortic arch obstruction and seven had recoarctation. RESULTS: Reduction in peak systolic gradient to <10 mm Hg was achieved in seven of 10 patients with an improvement in aortic artery diameter to >90% of adjacent aorta in all. The diameter of the arch obstruction increased from a median of 1.60 to 4.90 mm (P = 0.001) and the peak systolic gradient from 45 mm Hg to 8 mm Hg (P < 0.0001). Adverse events occurred in two patients one who required further surgical revision and a second who required placement of a second stent. The median follow up was 3.60 years (range 0.4-5.5 years) with two patients having died at 1.34 and 1.42 years poststent placement. Of the nine patients alive, six have since undergone further angioplasty at a median time interval of 0.77 years (range 0.17-2.76 years). Long-term complications occurred in none. CONCLUSIONS: Endovascular stent placement in infants is technically feasible with good results achievable even in small babies. It should be considered as a therapeutic option in complex cases when surgical alternatives are less favorable.

Occlusion of anomalous systemic arterial supply in Scimitar syndrome using the new Amplatz vascular plug IV.

Scimitar syndrome is a rare condition often with a separate systemic arterial supply from the abdominal aorta. Occlusion of this systemic arterial supply is frequently performed, though it can be difficult in small patients or in those with tortuous vessels. This case documents use of the new Amplatz vascular plug IV for arterial occlusion. It has major advantages in being able to deliver the device through a 4F catheter without the need to upsize to a dedicated delivery sheath. This is particularly appealing to paediatric practice or to those with difficult anatomy in older patients.

Early outcomes of percutaneous pulmonary valve implantation using the Edwards SAPIEN 3 transcatheter heart valve system.

AIMS: Multiple surgical revisions are often necessary in individuals with congenital heart defects affecting the RVOT or pulmonary valve. There are no multicentre data on the feasibility and safety of percutaneous pulmonary valve implantation (PPVI) using the SAPIEN 3 (S3) transcatheter heart valve. The aim of this study was to explore the short-term safety, feasibility, and haemodynamic outcomes of PPVI using the S3 transcatheter heart valve. METHODS AND RESULTS: Pulmonic S3 is an observational registry of patients undergoing PPVI with the S3 valve across centres in Europe and Canada. Data for 82 patients (mean age 27.3 years) were obtained. The most common underlying diagnosis was tetralogy of Fallot (ToF) (58.5%), with 16.0% of patients having native RVOT anatomy; 90.2% received pre-stenting. Prosthesis dislodgement occurred in one patient and conduit perforation in another. Both were successfully resolved without the need for open surgery. Peak systolic gradient over the RVOT fell from 46.3 mmHg to 17.2 mmHg, moderate/severe pulmonary regurgitation from 86.3% to 0.0%, and NYHA ≥II from 86.0% to 15.2%. During follow-up, valve thrombosis was observed in two patients which resolved with adequate anticoagulation. No other procedural complications, endocarditis, stent fracture or death were reported within two years. CONCLUSIONS: PPVI with the S3 valve appears feasible and safe in a wide range of patients with congenital heart defects, with good short-term haemodynamic and functional outcomes.

Genetic mechanisms controlling cardiovascular development.

Congenital heart disease (CHD) is a major cause of childhood morbidity and death in the West; the incidence is approximately 1 in 145 live births. Mendelian and chromosomal syndromes account for approximately 20% of CHD. The genetic mechanisms underlying non-chromosomal or non-Mendelian "sporadic" CHD, which account for the remaining 80%, are poorly understood. The genetic architecture of sporadic CHD likely includes accumulation of rare nonsynonymous variants in cardiac developmental genes leading to mutational loading of cardiac developmental networks, copy number variation in cardiac developmental genes, and common variants that may not be obviously linked to cardiac development but may alter genetic buffering pathways (e.g., folate metabolism). The rare mutations typically associated with sporadic CHD likely arise from the severe decrease in reproductive fitness selecting against any CHD-causing gene variant. The resulting allelic heterogeneity reduces the power of genome-wide association studies for CHD. A complementary approach to the genetic analysis of CHD is to resequence candidate genes that have been shown to be necessary for mouse heart development. The number of such genes likely exceeds 1700. To identify these genes, we have developed an enabling technology (high-throughput magnetic resonance imaging of mouse embryos), which is used in combination with N-ethyl-N-nitrosourea/transposon mutagenesis and knockout techniques. Key future challenges now involve translating discoveries made in mouse models to human CHD genetics and understanding the mechanisms that create and disrupt genetic buffering. A long-term goal in CHD is to manipulate these pathways to enhance buffering and prevent disease in a manner analogous to the use of folate in preventing neural tube defects.

Distinct genetic architectures for syndromic and nonsyndromic congenital heart defects identified by exome sequencing.

Congenital heart defects (CHDs) have a neonatal incidence of 0.8-1% (refs. 1,2). Despite abundant examples of monogenic CHD in humans and mice, CHD has a low absolute sibling recurrence risk (∼2.7%), suggesting a considerable role for de novo mutations (DNMs) and/or incomplete penetrance. De novo protein-truncating variants (PTVs) have been shown to be enriched among the 10% of 'syndromic' patients with extra-cardiac manifestations. We exome sequenced 1,891 probands, including both syndromic CHD (S-CHD, n = 610) and nonsyndromic CHD (NS-CHD, n = 1,281). In S-CHD, we confirmed a significant enrichment of de novo PTVs but not inherited PTVs in known CHD-associated genes, consistent with recent findings. Conversely, in NS-CHD we observed significant enrichment of PTVs inherited from unaffected parents in CHD-associated genes. We identified three genome-wide significant S-CHD disorders caused by DNMs in CHD4, CDK13 and PRKD1. Our study finds evidence for distinct genetic architectures underlying the low sibling recurrence risk in S-CHD and NS-CHD.

Cerebral infarct and meningitis secondary to Lemierre's syndrome.

The case is reported of a young man with Fusobacterium necrophorum septicemia who developed cavernous sinus thrombosis, meningitis, carotid artery stenosis and stroke. This article presents the only reported case of arterial stroke in Lemierre's syndrome. Clinical presentation, diagnostic difficulty and treatment are discussed.