FBN2 pathogenic variants in congenital contractural arachnodactyly with severe cardiovascular manifestations.

PURPOSE: Congenital contractural arachnodactyly (CCA) is an extremely rare autosomal dominant connective tissue genetic disorder caused by pathogenic variants in FBN2. CCA is characterized by arachnodactyly, camptodactyly, contracture of major joints, scoliosis, pectus deformities, and crumpled ears, but rarely with lethal cardiovascular manifestations as in Marfan syndrome. It is imperative to conduct a comprehensive analysis and review of the pathogenesis of CCA resulting from pathogenic variants in FBN2 gene. MATERIALS AND METHODS: Using whole-exome sequencing and Sanger sequencing, we identified a novel pathogenic splice-altering variant (c.4472-3C>A) in intron 34 of FBN2 gene in a CCA pedigree. The transcriptional result of the splicing-altering variant was analyzed by RNA sequencing. We systematically analyzed the clinical manifestations of all reported cases of CCA caused by splicing-altering pathogenic variants and focused on all the pathogenic variants in FBN2 gene that are associated with severe cardiovascular manifestations. RESULTS: The splice-altering variant (c.4472-3C>A) in FBN2 was demonstrated to result in the exon 35 skipping and cause an in-frame deletion. Furthermore, we identified exons 31 to 35 may be a hotspot region in FBN2 gene associated with severe cardiovascular phenotype. CONCLUSIONS: This study enriched the pathogenic spectrum of CCA and identified a hotspot region in FBN2 gene associated with severe cardiovascular manifestations. We recommend that patients carrying pathogenic variants in exons 31 to 35 of FBN2 pay more attention to cardiac evaluation.

Loeys-Dietz and Shprintzen-Goldberg syndromes: analysis of TGF-ß-opathies with craniofacial manifestations using an innovative multimodality method.

BACKGROUND: Elevated transforming growth factor-beta (TGF-ß) signalling has been implicated in the pathogenesis of Loeys-Dietz syndrome (LDS) and Shprintzen-Goldberg syndrome (SGS). In this study, we provide a qualitative and quantitative analysis of the craniofacial and functional features among the LDS subtypes and SGS. METHODS: We explore the variability within and across a cohort of 44 patients through deep clinical phenotyping, three-dimensional (3D) facial photo surface analysis, cephalometric and geometric morphometric analyses of cone-beam CT scans. RESULTS: The most common craniofacial features detected in this cohort include mandibular retrognathism (84%), flat midface projection (84%), abnormal eye shape (73%), low-set ears (73%), abnormal nose (66%) and lip shape (64%), hypertelorism (41%) and a relatively high prevalence of nystagmus/strabismus (43%), temporomandibular joint disorders (38%) and obstructive sleep apnoea (23%). 3D cephalometric analysis demonstrated an increased cranial base angle with shortened anterior cranial base and underdevelopment of the maxilla and mandible, with evidence of a reduced pharyngeal airway in 55% of those analysed. Geometric morphometric analysis confirmed that the greatest craniofacial shape variation was among patients with LDS type 2, with distinct clustering of patients with SGS. CONCLUSIONS: This comprehensive phenotypic approach identifies developmental abnormalities that segregate to mutation variants along the TGF-ß signalling pathway, with a particularly severe phenotype associated with TGFBR2 and SKI mutations. Multimodality assessment of craniofacial anomalies objectively reveals the impact of mutations of the TGF-ß pathway with perturbations associated with the cranium and cranial base with severe downstream effects on the orbit, maxilla and mandible with the resultant clinical phenotypes.

Two novel variants in SCARF2 gene underlie van den Ende-Gupta syndrome.

Van den Ende-Gupta syndrome (VDEGS) (MIM#600920) is characterized by skeletal and craniofacial abnormalities that include prominent ears, downslanting palpebral fissures, blepharophimosis, hypoplastic maxilla with or without a cleft palate, a narrow and convex nasal bridge and an everted lower lip, camptodactyly and arachnodactyly. Intelligence is normal. Recent studies have reported that patients with VDEGS have pathogenic variants in the SCARF2 gene on chromosome 22q11.21. Here, we report two Turkish patients with two novel variants [c.2291_2292insC (p.Ser765LeufsTer6) and c.488G>A (p.Cys63Tyr)] in the SCARF2 gene. In silico analysis predicted that both of these novel variants were pathogenic. To the best of our knowledge, this is the first case report of this syndrome in Turkey.

[A novel splicing acceptor variant of the FBN2 gene contributes to a case of congenital contractural arachnodactyly].

OBJECTIVE: To identify the pathogenic variants from a patient with suspected congenital contractural arachnodactyly, and to explore the possible molecular genetic pathogenesis, so as to provide evidence for clinical diagnosis. METHODS: Whole exome sequencing was performed for the patient. The splicing site variation of candidate pathogenic genes was verified by Sanger sequencing, and the new transcript sequence was determined by RT-PCR and TA-cloning sequencing. RESULTS: The patient carried a heterozygous c.533-1G>C variant of FBN2 gene, which was not reported. The sequencing of mRNA showed that the variant leaded to the disappearance of the canonical splice acceptor site of FBN2 gene and the activation of a cryptic splice acceptor site at c.533-71, resulting in the insertion of 70 bp sequence in the new transcript. It was speculated that the polypeptide encoded by the new transcript changed from valine (Val) to serine (Ser) at amino acid 179, and prematurely terminated after 26 aminoacids. According to the guidelines of American College of Medical Genetics and Genomics, the variant of FBN2 gene c. 533-1G>C was determined as pathogenic (PVS1+PM2+PP3 ). CONCLUSION: A novel splicing variant of FBN2 gene (c.533-1G>C) was identified, which can lead to congenital contractural arachnodactyly.

[Analysis of phenotype and genetic variant in a family with Shprintzen-Goldberg syndrome].

OBJECTIVE: To explore the genetic basis for a proband with Shprintzen-Goldberg syndrome (SGS). METHODS: Whole exome sequencing was carried out to detect potential variants associated with the relevant phenotypes. Candidate variants were verified by Sanger sequencing of the patient and her family. RESULTS: DNA sequencing revealed that that the proband has carried a de novo heterozygous missense c.94C>G (p.Leu32Val) variant in exon 1 of the SKI gene (NM_003036), which has been reported previously. The same variant was not detected in either parent. Based on the guidelines of the American College of Medical Genetics and Genomics, the variant was predicted to be pathogenic (PS1+PS2+PM1+PM2+PP2+PP3). CONCLUSION: The SKI c.94C>G (p. Leu32Val) variant probably underlay the autosomal dominant SGS in this patient.

Arachnodactyly represented in art.

Arachnodactyly, a term used since 1902 to describe abnormally long (spider-like) fingers, is a pathologic feature of several heritable conditions, notably the Marfan syndrome and congenital contractural arachnodactyly. A number of prominent artists, dating from the 16th to the 20th centuries, have depicted subjects with unusually long fingers, sometime associated with elongation of the body, neck and head. El Greco incorporated this style in many paintings. Little evidence supports any subject in any of these paintings as having a congenital deformity.

Further delineation of van den Ende-Gupta syndrome: Genetic heterogeneity and overlap with congenital heart defects and skeletal malformations syndrome.

Van den Ende-Gupta syndrome (VDEGS) is a rare autosomal recessive condition characterized by distinctive facial and skeletal features, and in most affected persons, by biallelic pathogenic variants in SCARF2. We review the type and frequency of the clinical features in 36 reported individuals with features of VDEGS, 15 (42%) of whom had known pathogenic variants in SCARF2, 6 (16%) with negative SCARF2 testing, and 15 (42%) not tested. We also report three new individuals with pathogenic variants in SCARF2 and clinical features of VDEGS. Of the six persons without known pathogenic variants in SCARF2, three remain unsolved despite extensive genetic testing. Three were found to have pathogenic ABL1 variants using whole exome sequencing (WES) or whole genome sequencing (WGS). Their phenotype was consistent with the congenital heart disease and skeletal malformations syndrome (CHDSKM), which has been associated with ABL1 variants. Of the three unsolved cases, two were brothers who underwent WGS and targeted long-range sequencing of both SCARF2 and ABL1, and the third person who underwent WES and RNA sequencing for SCARF2. Because these affected individuals with classical features of VDEGS lacked a detectable pathogenic SCARF2 variant, genetic heterogeneity is likely. Our study shows the importance of performing genetic testing on individuals with the VDEGS "phenotype," either as a targeted gene analysis (SCARF2, ABL1) or WES/WGS. Additionally, individuals with the combination of arachnodactyly and blepharophimosis should undergo echocardiography while awaiting results of molecular testing due to the overlapping physical features of VDEGS and CHDSKM.

Expanding the phenotypic spectrum of Mendelian connective tissue disorders to include prominent kidney phenotypes.

Heritable connective tissue disorders are a group of diseases, each rare, characterized by various combinations of skin, joint, musculoskeletal, organ, and vascular involvement. Although kidney abnormalities have been reported in some connective tissue disorders, they are rarely a presenting feature. Here we present three patients with prominent kidney phenotypes who were found by whole exome sequencing to have variants in established connective tissue genes associated with Loeys-Dietz syndrome and congenital contractural arachnodactyly. These cases highlight the importance of considering connective tissue disease in children presenting with structural kidney disease and also serves to expand the phenotype of Loeys-Dietz syndrome and possibly congenital contractural arachnodactyly to include cystic kidney disease and cystic kidney dysplasia, respectively.

Confirming the involvement of PIEZO2 in the etiology of Marden-Walker syndrome.

Pathogenic heterozygous variants in PIEZO2 typically cause distal arthrogryposis type 5 (DA5) and the closely related Gordon syndrome (GS). Only one case of PIEZO2-related Marden-Walker syndrome (MWS) has been reported to date. We report the phenotypic features of a Saudi female patient with features consistent with MWS in whom we identified a novel de novo likely pathogenic variant in PIEZO2. Our case lends support to the link between PIEZO2 and MWS.

22q11.2 deletion syndrome and congenital heart disease.

The 22q11.2 deletion syndrome has an estimated prevalence of 1 in 4-6,000 livebirths. The phenotype varies widely; the most common features include: facial dysmorphia, hypocalcemia, palate and speech disorders, feeding and gastrointestinal disorders, immunodeficiency, recurrent infections, neurodevelopmental and psychiatric disorders, and congenital heart disease. Approximately 60-80% of patients have a cardiac malformation most commonly including a subset of conotruncal defects (tetralogy of Fallot, truncus arteriosus, interrupted aortic arch type B), conoventricular and/or atrial septal defects, and aortic arch anomalies. Cardiac patients with a 22q11.2 deletion do not generally experience higher mortality upon surgical intervention but suffer more peri-operative complications than their non-syndromic counterparts. New guidelines suggest screening for a 22q11.2 deletion in the patient with tetralogy of Fallot, truncus arteriosus, interrupted aortic arch type B, conoventricular septal defects as well as those with an isolated aortic arch anomaly. Early identification of a 22q11.2 deletion in the neonate or infant when other syndromic features may not be apparent allows for timely parental screening for reproductive counseling and anticipatory evaluation of cardiac and noncardiac features. Screening the at-risk child or adult allows for important age-specific clinical, neurodevelopmental, psychiatric, and reproductive issues to be addressed.

A new mutational hotspot in the SKI gene in the context of MFS/TAA molecular diagnosis.

SKI pathogenic variations are associated with Shprintzen-Goldberg Syndrome (SGS), a rare systemic connective tissue disorder characterized by craniofacial, skeletal and cardiovascular features. So far, the clinical description, including intellectual disability, has been relatively homogeneous, and the known pathogenic variations were located in two different hotspots of the SKI gene. In the course of diagnosing Marfan syndrome and related disorders, we identified nine sporadic probands (aged 2-47 years) carrying three different likely pathogenic or pathogenic variants in the SKI gene affecting the same amino acid (Thr180). Seven of these molecular events were confirmed de novo. All probands displayed a milder morphological phenotype with a marfanoid habitus that did not initially lead to a clinical diagnosis of SGS. Only three of them had learning disorders, and none had intellectual disability. Six out of nine presented thoracic aortic aneurysm, which led to preventive surgery in the oldest case. This report extends the phenotypic spectrum of variants identified in the SKI gene. We describe a new mutational hotspot associated with a marfanoid syndrome with no intellectual disability. Cardiovascular involvement was confirmed in a significant number of cases, highlighting the importance of accurately diagnosing SGS and ensuring appropriate medical treatment and follow-up.

A clinical scoring system for congenital contractural arachnodactyly.

PURPOSE: Congenital contractural arachnodactyly (CCA) is an autosomal dominant connective tissue disorder manifesting joint contractures, arachnodactyly, crumpled ears, and kyphoscoliosis as main features. Due to its rarity, rather aspecific clinical presentation, and overlap with other conditions including Marfan syndrome, the diagnosis is challenging, but important for prognosis and clinical management. CCA is caused by pathogenic variants in FBN2, encoding fibrillin-2, but locus heterogeneity has been suggested. We designed a clinical scoring system and diagnostic criteria to support the diagnostic process and guide molecular genetic testing. METHODS: In this retrospective study, we assessed 167 probands referred for FBN2 analysis and classified them into a FBN2-positive (n = 44) and FBN2-negative group (n = 123) following molecular analysis. We developed a 20-point weighted clinical scoring system based on the prevalence of ten main clinical characteristics of CCA in both groups. RESULTS: The total score was significantly different between the groups (P < 0.001) and was indicative for classifying patients into unlikely CCA (total score <7) and likely CCA (total score ≥7) groups. CONCLUSIONS: Our clinical score is helpful for clinical guidance for patients suspected to have CCA, and provides a quantitative tool for phenotyping in research settings.

The Genetic Epidemiology of Pediatric Pulmonary Arterial Hypertension.

OBJECTIVE: To describe the prevalence of pulmonary arterial hypertension (PAH)-associated gene mutations, and other genetic characteristics in a national cohort of children with PAH from the Dutch National registry and to explore genotype-phenotype associations and outcomes. STUDY DESIGN: Children (n = 70) diagnosed with idiopathic PAH, heritable PAH, PAH associated with congenital heart disease with coincidental shunt (PAH-congenital heart disease group 3), PAH after closure of a cardiac shunt (PAH-congenital heart disease group 4), or PAH associated with other noncardiac conditions were enrolled. Targeted next-generation sequencing was performed on PAH-associated genes (BMPR2, ACVRL1, EIF2AK4, CAV1, ENG, KCNK3, SMAD9, and TBX4). Also, children were tested for specific genetic disorders in case of clinical suspicion. Additionally, children were tested for copy number variations. RESULTS: Nineteen children (27%) had a PAH-associated gene mutation/variant: BMPR2 n = 7, TBX4 n = 8, ACVRL1 n = 1, KCNK3 n = 1, and EIF2AK4 n = 2. Twelve children (17%) had a genetic disorder with an established association with PAH (including trisomy 21 and cobalamin C deficiency). In another 16 children (23%), genetic disorders without an established association with PAH were identified (including Noonan syndrome, Beals syndrome, and various copy number variations). Survival rates differed between groups and was most favorable in TBX4 variant carriers. CONCLUSIONS: Children with PAH show a high prevalence of genetic disorders, not restricted to established PAH-associated genes. Genetic architecture could play a role in risk-stratified care management in pediatric PAH.

Phenotypic spectrum of TGFB3 disease-causing variants in a Dutch-French cohort and first report of a homozygous patient.

Disease-causing variants in TGFB3 cause an autosomal dominant connective tissue disorder which is hard to phenotypically delineate because of the small number of identified cases. The purpose of this retrospective cross-sectional multicenter study is to elucidate the genotype and phenotype in an international cohort of TGFB3 patients. Eleven (eight novel) TGFB3 disease-causing variants were identified in 32 patients (17 families). Aortic root dilatation and mitral valve disease represented the most common cardiovascular findings, reported in 29% and 32% of patients, respectively. Dissection involving distal aortic segments occurred in two patients at age 50 and 52 years. A high frequency of systemic features (65% high-arched palate, 63% arachnodactyly, 57% pectus deformity, 52% joint hypermobility) was observed. In familial cases, incomplete penetrance and variable clinical expressivity were noted. Our cohort included the first described homozygous patient, who presented with a more severe phenotype compared to her heterozygous relatives. In conclusion, TGFB3 variants were associated with a high percentage of systemic features and aortic disease (dilatation/dissection) in 35% of patients. No deaths occurred from cardiovascular events or pregnancy-related complications. Nevertheless, homozygosity may be driving a more severe phenotype.

Variant filtering, digenic variants, and other challenges in clinical sequencing: a lesson from fibrillinopathies.

Genome-scale high-throughput sequencing enables the detection of unprecedented numbers of sequence variants. Variant filtering and interpretation are facilitated by mutation databases, in silico tools, and population-based reference datasets such as ExAC/gnomAD, while variants are classified using the ACMG/AMP guidelines. These methods, however, pose clinically relevant challenges. We queried the gnomAD dataset for (likely) pathogenic variants in genes causing autosomal-dominant disorders. Furthermore, focusing on the fibrillinopathies Marfan syndrome (MFS) and congenital contractural arachnodactyly (CCA), we screened 500 genomes of our patients for co-occurring variants in FBN1 and FBN2. In gnomAD, we detected 2653 (likely) pathogenic variants in 253 genes associated with autosomal-dominant disorders, enabling the estimation of variant-filtering thresholds and disease predisposition/prevalence rates. In our database, we discovered two families with hitherto unreported co-occurrence of FBN1/FBN2 variants causing phenotypes with mixed or modified MFS/CCA clinical features. We show that (likely) pathogenic gnomAD variants may be more frequent than expected and are challenging to classify according to the ACMG/AMP guidelines as well as that fibrillinopathies are likely underdiagnosed and may co-occur. Consequently, selection of appropriate frequency cutoffs, recognition of digenic variants, and variant classification represent considerable challenges in variant interpretation. Neglecting these challenges may lead to incomplete or missed diagnoses.

Exploring the potential association among sleep disturbances, cognitive impairments, and immune activation in 22q11.2 deletion syndrome.

22q11.2 deletion syndrome (22q11.DS) is a neurogenetic disorder caused by a microdeletion in chromosome 22. Its phenotype includes high rates of psychiatric disorders, immune system abnormalities, and cognitive impairments. We assessed the quality of sleep in 22q11.2DS and its potential link to inflammatory markers and cognitive deficits. Thirty-three 22q11.2DS individuals and 24 healthy controls were studied. Sleep parameters were assessed by the Pittsburgh sleep quality index (PSQI) questionnaire and correlated with serum cytokine levels and cognitive functioning, measured using the Penn computerized neurocognitive battery (CNB). The 22q11.2DS individuals had significantly worse sleep quality scores than the controls, unrelated to the psychiatric or physical comorbidities common to 22q11.2DS. Interleukin 6 levels were correlated with the overall score of the PSQI questionnaire for nonpsychotic 22q11.2DS participants only. Several domains of the CNB were associated with poorer sleep quality, suggesting that cognitive impairments in 22q11.2DS may be at least partially explained by poor sleep quality. Our findings confirm sleep impairments in individuals with 22q11.2DS, which might negatively affect their cognitive functioning, and corroborate a potential role of immunological pathways in the 22q11.2DS neuro-phenotype.

[Pathological variant of FBN2 gene identified in a pedigree affected with congenital contracture arachnodactyly].

OBJECTIVE: To detect pathological variant in a Chinese pedigree affected with congenital contractural arachnodactyly (CCA). METHODS: Next generation sequencing (NGS) was used to scan the whole exome of the proband. Potential variant of the FBN2 gene was also detected in all members of the pedigree and 100 healthy controls by Sanger sequencing. With the determination of the genotype, prenatal diagnosis was carried out by amniotic fluid sampling. RESULTS: A c.3528C>A (p.Asn1176Lys) variant was identified in the FBN2 gene of the proband, other patients from this pedigree, as well as the fetus. The same variant was not found among healthy members from this pedigree and the 100 healthy controls. CONCLUSION: The c.3528C>A (p.Asn1176Lys) variant of the FBN2 gene probably underlies the pathogenesis of CCA in our case. The new variant has enriched pathological spectrum of the FBN2 gene.

Biallelic Loss of Proprioception-Related PIEZO2 Causes Muscular Atrophy with Perinatal Respiratory Distress, Arthrogryposis, and Scoliosis.

We report ten individuals of four independent consanguineous families from Turkey, India, Libya, and Pakistan with a variable clinical phenotype that comprises arthrogryposis, spontaneously resolving respiratory insufficiency at birth, muscular atrophy predominantly of the distal lower limbs, scoliosis, and mild distal sensory involvement. Using whole-exome sequencing, SNPchip-based linkage analysis, DNA microarray, and Sanger sequencing, we identified three independent homozygous frameshift mutations and a homozygous deletion of two exons in PIEZO2 that segregated in all affected individuals of the respective family. The mutations are localized in the N-terminal and central region of the gene, leading to nonsense-mediated transcript decay and consequently to lack of PIEZO2 protein. In contrast, heterozygous gain-of-function missense mutations, mainly localized at the C terminus, cause dominant distal arthrogryposis 3 (DA3), distal arthrogryposis 5 (DA5), or Marden-Walker syndrome (MWKS), which encompass contractures of hands and feet, scoliosis, ophthalmoplegia, and ptosis. PIEZO2 encodes a mechanosensitive ion channel that plays a major role in light-touch mechanosensation and has recently been identified as the principal mechanotransduction channel for proprioception. Mice ubiquitously depleted of PIEZO2 are postnatally lethal. However, individuals lacking PIEZO2 develop a not life-threatening, slowly progressive disorder, which is likely due to loss of PIEZO2 protein in afferent neurons leading to disturbed proprioception causing aberrant muscle development and function. Here we report a recessively inherited PIEZO2-related disease and demonstrate that depending on the type of mutation and the mode of inheritance, PIEZO2 causes clinically distinguishable phenotypes.

A de novo mutation in DHD domain of SKI causing spina bifida with no craniofacial malformation or intellectual disability.

Shprintzen-Goldberg syndrome (SGS) is a rare systemic connective tissue disorder characterized by craniofacial, skeletal, and cardiovascular manifestations. It is associated with a significant risk of intellectual disability, a feature which distinguishes it from Marfan and Loeys-Dietz syndromes. SGS is mainly caused by mutations in the SKI gene, a repressor of TGF-ß activity. Most SKI mutations are found in exon 1 of the gene and are located in the R-SMAD domain, a proposed hotspot for de novo mutations. Here, we report on a de novo SKI mutation located in the DHD domain of SKI. By adding our finding to previously reported de novo SKI mutations, a new mutational hotspot in the DHD domain is proposed. Our patient presented with a lipomeningomyelocele, tethered cord, and spina bifida but with no SGS-related clinical findings apart from a marfanoid habitus and long slender fingers. Specifically, she did not have an intellectual disability, craniofacial, or cardiovascular abnormalities. By comparing the clinical findings on patients with mutations in the R-SMAD and DHD domains of SKI, we propose that mutations in those domains have different effects on TGF-ß activity during embryonic development with resulting phenotypic differences.

Molecular Characterization of Three Canine Models of Human Rare Bone Diseases: Caffey, van den Ende-Gupta, and Raine Syndromes.

One to two percent of all children are born with a developmental disorder requiring pediatric hospital admissions. For many such syndromes, the molecular pathogenesis remains poorly characterized. Parallel developmental disorders in other species could provide complementary models for human rare diseases by uncovering new candidate genes, improving the understanding of the molecular mechanisms and opening possibilities for therapeutic trials. We performed various experiments, e.g. combined genome-wide association and next generation sequencing, to investigate the clinico-pathological features and genetic causes of three developmental syndromes in dogs, including craniomandibular osteopathy (CMO), a previously undescribed skeletal syndrome, and dental hypomineralization, for which we identified pathogenic variants in the canine SLC37A2 (truncating splicing enhancer variant), SCARF2 (truncating 2-bp deletion) and FAM20C (missense variant) genes, respectively. CMO is a clinical equivalent to an infantile cortical hyperostosis (Caffey disease), for which SLC37A2 is a new candidate gene. SLC37A2 is a poorly characterized member of a glucose-phosphate transporter family without previous disease associations. It is expressed in many tissues, including cells of the macrophage lineage, e.g. osteoclasts, and suggests a disease mechanism, in which an impaired glucose homeostasis in osteoclasts compromises their function in the developing bone, leading to hyperostosis. Mutations in SCARF2 and FAM20C have been associated with the human van den Ende-Gupta and Raine syndromes that include numerous features similar to the affected dogs. Given the growing interest in the molecular characterization and treatment of human rare diseases, our study presents three novel physiologically relevant models for further research and therapy approaches, while providing the molecular identity for the canine conditions.