Whole-exome-based single nucleotide variants and copy number analysis for prenatal diagnosis of compound heterozygosity of SMPD4.

BACKGROUND: Biallelic loss-of-function variants in SMPD4 cause a rare and severe neurodevelopmental disorder. These variants have been identified in a group of children with neurodevelopmental disorders with microcephaly, arthrogryposis, and structural brain anomalies. SMPD4 encodes a sphingomyelinase that hydrolyzes sphingomyelin into ceramide at neutral pH and can thereby affect membrane lipid homeostasis. SMPD4 localizes to the membranes of the endoplasmic reticulum and nuclear envelope and interacts with nuclear pore complexes. MATERIALS AND METHODS: For the efficient prenatal diagnosis of rare and undiagnosed diseases, the parallel detection of copy number variants (CNVs) and single nucleotide variants using whole-exome analysis is required. A physical examination of the parents was performed. Karyotype and whole-exome analysis were performed for the fetus and the parents. RESULTS: A fetus with microcephaly and arthrogryposis; biallelic null variants (c.387-1G>A; Chr2[GRCh38]: g.130142742_130202459del) were detected by whole-exome sequencing (WES). We have reported for the first time the biallelic loss-of-function mutations in SMPD4 in patients born to unrelated parents in China. CONCLUSION: WES could replace chromosomal microarray analysis and copy number variation sequencing as a more cost-effective genetic test for detecting CNVs and diagnosing highly heterogeneous conditions.

Compound heterozygous variants in MYBPC1 lead to severe distal arthrogryposis type-1 manifestations.

Dominant missense variants in MYBPC1 encoding slow Myosin Binding Protein-C (sMyBP-C) have been increasingly linked to arthrogryposis syndromes and congenital myopathy with tremor. Herein, we describe novel compound heterozygous variants - NM_002465.4:[c.2486_2492del];[c.2663A > G] - present in fibronectin-III (Fn-III) C7 and immunoglobulin (Ig) C8 domains, respectively, manifesting as severe, early-onset distal arthrogryposis type-1, with the carrier requiring intensive care and several surgical interventions at an early age. Computational modeling predicts that the c.2486_2492del p.(Lys829IlefsTer7) variant destabilizes the structure of the Fn-III C7 domain, while the c.2663A > G p.(Asp888Gly) variant causes minimal structural alterations in the Ig C8 domain. Although the parents of the proband are heterozygous carriers for a single variant, they exhibit no musculoskeletal defects, suggesting a complex interplay between the two mutant alleles underlying this disorder. As emerging novel variants in MYBPC1 are shown to be causatively associated with musculoskeletal disease, it becomes clear that MYBPC1 should be included in relevant genetic screenings.

Functional assessment of a novel biallelic MYH3 variation causing CPSKF1B (contractures, pterygia, and spondylocarpotarsal fusion syndrome1B).

BACKGROUND: The MYH3-associated myosinopathies comprise a spectrum of rare neuromuscular disorders mainly characterized by distal arthrogryposis with or without other features like pterygia and vertebrae fusion. CPSKF1B (contractures, pterygia, and spondylocarpotarsal fusion syndrome1B) is the only known autosomal recessiveMYH3-associated myosinopathy so far, with no more than two dozen cases being reported. MATERIALS AND METHODS: A boy with CPSKF1B was recruited and subjected to a comprehensive clinical and imaging evaluation. Genetic detection with whole-exome sequencing (WES) was performed on the patient and extended family members to identify the causative variation. A series of in silico and in vitro investigations were carried out to verify the pathogenicity of the two variants of the identified compound heterozygous variation. RESULTS: The patient exhibited moderate CPSKF1B symptoms including multiarticular contractures, webbed neck, and spondylocarpotarsal fusion. WES detected a compound heterozygous MYH3 variation consisting of two variants, namely NM_002470.4: c.3377A>G; p. (E1126G) and NM_002470.4: c.5161-2A>C. It was indicated that the NM_002470.4: c.3377A>G; p. (E1126G) variant mainly impaired the local hydrogen bond formation and impacted the TGF-B pathway, while the NM_002470.4: c.5161-2A>C variant could affect the normal splicing of pre-mRNA, resulting in the appearance of multiple abnormal transcripts. CONCLUSIONS: The findings of this study expanded the mutation spectrum of CPSKF1B, provided an important basis for the counseling of the affected family, and also laid a foundation for the functional study of MYH3 mutations.

A mutation in F-actin polymerization factor suppresses the distal arthrogryposis type 5 PIEZO2 pathogenic variant in Caenorhabditis elegans.

The mechanosensitive PIEZO channel family has been linked to over 26 disorders and diseases. Although progress has been made in understanding these channels at the structural and functional levels, the underlying mechanisms of PIEZO-associated diseases remain elusive. In this study, we engineered four PIEZO-based disease models using CRISPR/Cas9 gene editing. We performed an unbiased chemical mutagen-based genetic suppressor screen to identify putative suppressors of a conserved gain-of-function variant pezo-1[R2405P] that in human PIEZO2 causes distal arthrogryposis type 5 (DA5; p. R2718P). Electrophysiological analyses indicate that pezo-1(R2405P) is a gain-of-function allele. Using genomic mapping and whole-genome sequencing approaches, we identified a candidate suppressor allele in the C. elegans gene gex-3. This gene is an ortholog of human NCKAP1 (NCK-associated protein 1), a subunit of the Wiskott-Aldrich syndrome protein (WASP)-verprolin homologous protein (WAVE/SCAR) complex, which regulates F-actin polymerization. Depletion of gex-3 by RNAi, or with the suppressor allele gex-3(av259[L353F]), significantly increased brood size and ovulation rate, as well as alleviating the crushed oocyte phenotype of the pezo-1(R2405P) mutant. Expression of GEX-3 in the soma is required to rescue the brood size defects in pezo-1(R2405P) animals. Actin organization and orientation were disrupted and distorted in the pezo-1 mutants. Mutation of gex-3(L353F) partially alleviated these defects. The identification of gex-3 as a suppressor of the pathogenic variant pezo-1(R2405P) suggests that the PIEZO coordinates with the cytoskeleton regulator to maintain the F-actin network and provides insight into the molecular mechanisms of DA5 and other PIEZO-associated diseases.

Clinical and molecular characteristics of 26 fetuses with lethal multiple congenital contractures.

Multiple congenital contractures (MCC) due to fetal akinesia manifest across a broad spectrum of diseases, ranging from mild distal arthrogryposis to lethal fetal akinesia deformation sequence. We hereby present a series of 26 fetuses displaying severe MCC phenotypes from 18 families and describe detailed prenatal ultrasound findings, postmortem clinical evaluations, and genetic investigations. Most common prenatal findings were abnormal facial profile (65%), central nervous system abnormalities (62%), polyhydramnios (50%), increased nuchal translucency (50%), and fetal hydrops (35%). Postmortem examinations unveiled additional anomalies including facial dysmorphisms, dysplastic skeletal changes, ichthyosis, multiple pterygia, and myopathy, allowing preliminary diagnosis of particular Mendelian disorders in multiple patients. Evaluation of the parents revealed maternal grip myotonia in one family. By exome sequencing and targeted testing, we identified causative variants in ACTC1, CHST14, COG6, DMPK, DOK7, HSPG2, KLHL7, KLHL40, KIAA1109, NEB, PSAT1, RAPSN, USP14, and WASHC5 in 15 families, and one patient with a plausible diagnosis associated with biallelic NEB variants. Three patients received a dual diagnosis. Pathogenic alterations in newly discovered genes or in previously known genes recently linked to new MCC phenotypes were observed in 44% of the cohort. Our results provide new insights into the clinical and molecular landscape of lethal MCC phenotypes.

Arthrogryposis multiplex congenita: dental and maxillofacial phenotype - A scoping review.

INTRODUCTION: Arthrogryposis multiplex congenita (AMC) is a heterogeneous group of disorders associated with decreased fetal movement, with a prevalence between 1/3000 and 1/5200 live births. Typical features of AMC include multiple joint contractures present at birth, and can affect all joints of the body, from the jaw, and involving the upper limbs, lower limbs and spine. The jaws may be affected in 25 % of individuals with AMC, with limited jaw movement and mouth opening. Other oral and maxillofacial deformities may be present in AMC, including cleft palate, micrognathia, periodontitis and delayed teething. To our knowledge, oral and maxillofacial abnormalities have not been systematically assessed in individuals with AMC. Therefore, this scoping review was conducted to identify, collect, and describe a comprehensive map of the existing knowledge on dental and maxillofacial involvement in individuals with AMC. METHODOLOGY: A scoping review was conducted in accordance with the Preferred Reporting Items for Systematic reviews and Meta-Analyses extension for Scoping Reviews guidelines. The PRISMA guidelines for scoping reviews were followed and databases were searched for empirical articles in English and French published until October 2022. We searched MEDLINE, Embase, Web of Science and ERIC databases. Two authors independently reviewed the articles and extracted the data. RESULTS: Of a total of 997 studies that were identified, 96 met the inclusion criteria and were subsequently included in this scoping review. These 96 studies collectively provided insights into 167 patients who exhibited some form of oral and/or maxillofacial involvement. Notably, 25 % of these patients were within the age range of 0-6 months. It is worth highlighting that only 22 out of the 96 studies (22.9 %), had the primary objective of evaluating dental and/or maxillofacial deformities. Among the patients studied, a prevalent pattern emerged, revealing that severe anomalies such as micrognathia (56 %), high-arched palate (29 %), cleft palate (40 %), limited mouth opening (31 %), and dental anomalies (28 %) were frequently observed. Importantly, many of these patients were found to have more than one of these anomalies. Even though these maxillofacial impairments are known to be associated with dental problems (e.g., cleft palate is associated with oligodontia, hypodontia, and malocclusion), their secondary effects on the dental phenotype were not reported in the studies. CONCLUSION: Our findings have uncovered a notable deficiency in existing literature concerning dental and maxillofacial manifestations in AMC. This underscores the need for interdisciplinary collaboration and the undertaking of extensive prospective cohort studies focused on AMC. These studies should assess the oral and maxillofacial abnormalities that can impact daily functioning and overall quality of life.

Variants in DOK7 results in fetal akinesia deformation sequence: A case report and review of literature.

We report the third case of FADS due to biallelic DOK7 variants, which further strengthens the association of DOK7 with this lethal phenotype and lack of genotype phenotype correlation.

Two novel cases of biallelic SMPD4 variants with brain structural abnormalities.

Sphingomyelin phosphodiesterase 4 (SMPD4) encodes a member of the Mg2+-dependent, neutral sphingomyelinase family that catalyzes the hydrolysis of the phosphodiester bond of sphingomyelin to form phosphorylcholine and ceramide. Recent studies have revealed that biallelic loss-of-function variants of SMPD4 cause syndromic neurodevelopmental disorders characterized by microcephaly, congenital arthrogryposis, and structural brain anomalies. In this study, three novel loss-of-function SMPD4 variants were identified using exome sequencing (ES) in two independent patients with developmental delays, microcephaly, seizures, and brain structural abnormalities. Patient 1 had a homozygous c.740_741del, p.(Val247Glufs*21) variant and showed profound intellectual disability, hepatomegaly, a simplified gyral pattern, and a thin corpus callosum without congenital dysmorphic features. Patient 2 had a compound heterozygous nonsense c.2124_2125del, p.(Phe709*) variant and splice site c.1188+2dup variant. RNA analysis revealed that the c.1188+2dup variant caused exon 13 skipping, leading to a frameshift (p.Ala406Ser*6). In vitro transcription analysis using minigene system suggested that mRNA transcribed from mutant allele may be degraded by nonsense-mediated mRNA decay system. He exhibited diverse manifestations, including growth defects, muscle hypotonia, respiratory distress, arthrogryposis, insulin-dependent diabetes mellitus, sensorineural hearing loss, facial dysmorphism, and various brain abnormalities, including cerebral atrophy, hypomyelination, and cerebellar hypoplasia. Here, we review previous literatures and discuss the phenotypic diversity of SMPD4-related disorders.

Biallelic truncating TTN variants in M-band encoding exons cause a fetal lethal titinopathy.

To report two novel TTN variants associated with fetal recessive titinopathy, thereby broadening the range of TTN variants that can lead to titinopathy. Clinical information on the fetus and parents was gathered, and genomic DNAs were extracted from the fetal tissue and family members' peripheral blood samples. Exome sequencing on fetal DNA was performed and following bioinformatics analysis, the suspected pathogenic variants were confirmed through Sanger sequencing. Prenatal ultrasound performed at 29 weeks of gestation revealed hydrops fetalis, decreased fetal movements, multiple joint contractures and polyhydramnios. Intrauterine fetal death was noted in the third trimester. Exome sequencing revealed compound heterozygous variants in the TTN gene: a paternally inherited allele c.101227C>T (p.Arg33743Ter) and a maternally inherited c.104254C>T (p.Gln34752Ter) allele. These variants have not been previously reported and are evaluated to be likely pathogenic according to the American College of Medical Genetics and Genomics guidelines. We report a fetus with hydrops fetalis and arthrogryposis multiplex congenita associated with a compound heterozygote in the TTN gene. Our report broadens the clinical and genetic spectrum associated with the TTN-related conditions.

Biallellic variants in CACNA1S cause fetal akinesia sequence, progressive hydrops and stillbirth.

Fetal arthrogryposis is a well-recognised ultrasonographic phenotype, caused by both genetic, maternal and extrinsic factors. When present with fetal growth restriction, pulmonary hypoplasia and multiple joint contractures, it is often referred to as fetal akinesia deformation sequence (FADS). Historically, elucidating genetic causes of arthryogryposis/FADS has been challenging; there are now more than 150 genes known to cause arthrogryposis through myopathic, neuromuscular and metabolic pathways affecting fetal movement. FADS is associated with over 400 medical conditions making prenatal diagnosis challenging. Here we present a case of FADS diagnosed at 19 weeks gestation with progression to severe fetal hydrops and stillbirth at 26-weeks gestation. Initial investigations including combined first trimester screening, TORCH (infection) screen and chromosomal microarray were normal. Trio whole exome sequencing (WES) detected compound heterozygous likely pathogenic CACNA1S gene variants associated with autosomal dominant (AD) and autosomal recessive (AR) congenital myopathy and FADS. To our knowledge, this is the first prenatal diagnosis of this condition.

Foetal Akinesia Deformation Sequence: A Rare Lethal Entity.

Foetal akinesia deformation sequence (FADS) represents a group of disorders resulting from absent or diminished in utero foetal mobility. The aetiology is multifactorial, including genetic, environmental, maternal, and foetal causes. The absence of foetal movements leading to multiple joint contractures, pulmonary hypoplasia, and intrauterine growth restriction are the key features of foetal akinesia deformation sequence. Herein we describe the case of a 30-year-old gravida 4 (para 2+1) who came for foetal ultrasound at 28 weeks of gestation due to decreased foetal movements. Ultrasound showed features of FADS with fixed flexed position of foetal limbs, pulmonary hypoplasia, polyhydramnios, and intrauterine growth restriction. The timely use of ultrasound enables early detection of these cases and aids in appropriate counselling and management.

Muscular phenotype description of abnormal THOC2 splicing.

Until recently, the disease known to be associated with THOC2 mutations was Intellectual developmental disorder, X-linked 12 (MIM300957). However, recently, fetal arthrogryposis multiplex congenita has been associated with a specific splice site mutation in the THOC2 gene. We report a family with the same splice site mutation in the THOC2 gene involved in fetal arthrogryposis as well. We provide the first description of the muscular phenotype of this disease which reveals the presence of cytoplasmic bodies. Our findings expand the clinical phenotype of THOC2 gene related defects.

Features of Congenital Arthrogryposis Due to Abnormalities in Collagen Homeostasis, a Scoping Review.

Congenital arthrogryposis (CA) refers to the presence of multiple contractures at birth. It is a feature of several inherited syndromes, notable amongst them are disorders of collagen formation. This review aims to characterize disorders that directly or indirectly impact collagen structure and function leading to CA in search for common phenotypic or pathophysiological features, possible genotype-phenotype correlation, and potential novel treatment approaches based on a better understanding of the underlying pathomechanism. Nine genes, corresponding to five clinical phenotypes, were identified after a literature search. The most notable trend was the extreme phenotype variability. Clinical features across all syndromes ranged from subtle with minimal congenital contractures, to severe with multiple congenital contractures and extra-articular features including skin, respiratory, or other manifestations. Five of the identified genes were involved in the function of the Lysyl Hydroxylase 2 or 3 enzymes, which enable the hydroxylation and/or glycosylation of lysyl residues to allow the formation of the collagen superstructure. Whilst current treatment approaches are post-natal surgical correction, there are also potential in-utero therapies being developed. Cyclosporin A showed promise in treating collagen VI disorders although there is an associated risk of immunosuppression. The treatments that could be in the clinical trials soon are the splice correction therapies in collagen VI-related disorders.

Identification of two novel MYH3 variants causing different phenotypes in prenatal diagnosis.

The MYH3 gene encodes the embryonic myosin heavy chain, which is crucial for the skeletal and muscular development. The MYH3 variants are associated with distal arthrogryposis type 2A (Freeman-Sheldon syndrome), distal arthrogryposis type 2B3 (Sheldon-Hall syndrome), CPSFS1A (Contractures, pterygia, and spondylocarpostarsal fusion syndrome 1A) and CPSFS1B, which have some shared characteristics and great variability of clinical phenotypes. In this study, we report two novel MYH3 missense variants c.1024T>G (p.Phe342Val) and c.3872A>C (p.Gln1291Pro), demonstrating different phenotypes in the prenatal setting. This study expands the spectrum of MYH3 variants and supports the domain-specific genotype-phenotype correlation of MYH3.

Mutation In Fkbp10 Gene Cause Bruck Syndrome 1 (Brks1) In A Pakistani Family Of Pashtun Origin.

BACKGROUND: Bruck syndrome or BRKS1 is an extremely rare condition characterized by the onset of fractures in infancy, joint contractures, short stature, severe limb deformity, and progressive scoliosis. Less than fifty cases of BRKS1 have been reported so far. Here, we report Bruck syndrome 1 in two siblings who belong to a consanguineous Pashtun family living in Karachi. Our first case is a seven years old boy who presented with recurrent fractures, lower limb deformity, and unable to walk. He had markedly reduced bone mineral density (BMD) and a normal bone profile. The other sibling presented at one week of age with arthrogryposis multiplex congenita, post-axial polydactyly of both feet and spontaneous fracture of the right proximal femur. Genetic testing of our cases was performed in which genomic DNA was enriched for targeted regions using the hybridization-based protocol, and DNA sequencing was done using Illumina technology; both cases were found homozygous for pathogenic variant c.344G>A (p.Arg115Gln) in FKBP10 gene leading to the diagnosis of BRKS1. FKBP10 gene mutation has been reported earlier in association with BRKS1, but in our case report, we have reported the first case of BRKS1, particularly in the Pakistani population of Pashtun ethnicity. We have reported post-axial polydactyly of both feet and spina bifida for the first time in association with FKBP10 mutation. In addition, the skeletal survey of patients with BRKS 1 is elaborated in detail in this report.

Optic nerve abnormalities in female-restricted Wieacker-Wolff syndrome by a novel variant in the ZC4H2 gene.

BACKGROUND: Wieacker-Wolff syndrome is an ultra-rare disease with X-linked inheritance characterized by arthrogryposis, intellectual disability, microcephaly, and distal limb muscle atrophy. Ophthalmic abnormalities such as ptosis, strabismus, and oculomotor apraxia have been reported in half of the patients. Wieacker-Wolff syndrome female-restricted (WRWFFR) is an even rarer disease recently used for females with a more severe phenotype. MATERIALS AND METHODS: Clinical geneticist and ophthalmic examination, neuroimaging, and exome sequencing. RESULTS: A 4 years-old girl with developmental and language delay, microcephaly, camptodactyly, digital pads, and arthrogryposis was identified by the clinical geneticist. Ophthalmic examination revealed deep-set eyes, high hyperopic astigmatism in both eyes, and reduced retinal nerve fiber layer thickness measured by optical coherence tomography. Exome sequencing identified a novel, probably pathogenic variant in the ZC4H2 gene NM_018684.3:c.145A>T p. (Lys49*) in heterozygosis. DISCUSSION: WRWFFR is an ultra-rare disease with X-linked inheritance by variants in the ZC4H2 gene. This case reports a girl with a novel nonsense variant in the ZC4H2 gene and a severe phenotype; previous reports have identified WRWFFR in females with large deletions and nonsense mutations which could explain the manifestations in the current case report. A complete ophthalmic examination should be considered in patients with WRWFFR to detect the possibly associated optic nerve involvement and other previously described manifestations such as ptosis and strabismus.

DST variants are responsible for neurogenic arthrogryposis multiplex congenita enlarging the spectrum of type VI hereditary sensory autonomic neuropathy.

Arthrogryposis multiplex congenita (AMC) is a developmental condition characterized by multiple joint contractures resulting from reduced or absent fetal movements. Through whole-exome sequencing combined with arrayCGH from DNA of a fetus presenting with early onset AMC, we identified biallelic loss of function variants in Dystonin (DST): a stop gain variant (NM_001144769.5:c.12208G > T:p.(Glu4070Ter)) on the neuronal isoform and a 175 kb microdeletion including exons 25-96 of this isoform on the other allele [NC_000006.11:g.(56212278_56323554)_(56499398_56507586)del]. Transmission electron microscopy of the sciatic nerve revealed abnormal morphology of the peripheral nerve with severe hypomyelination associated with dramatic reduction of fiber density which highlights the critical role of DST in peripheral nerve axonogenesis during development in human. Variants in the neuronal isoforms of DST cause hereditary sensory and autonomic neuropathy which has been reported in several unrelated families with highly variable age of onset from fetal to adult onset. Our data enlarge the disease mechanisms of neurogenic AMC.

Neonatal developmental and epileptic encephalopathy with movement disorders and arthrogryposis: A case report with a novel missense variant of SCN1A.

Variants of SCN1A represent the archetypal channelopathy associated with several epilepsy syndromes. The clinical phenotypes have recently expanded from Dravet syndrome. CASE REPORT: We present a female patient with the de novo SCN1A missense variant, c.5340G > A (p. Met1780Ile). The patient had various clinical features with neonatal onset SCN1A epileptic encephalopathy, arthrogryposis multiplex congenita, thoracic hypoplasia, thoracic scoliosis, and hyperekplexia. CONCLUSION: Our findings are compatible with neonatal developmental and epileptic encephalopathy with movement disorders and arthrogryposis; the most severe phenotype probably caused by gain-of-function variant of SCN1A. The efficacy of sodium channel blocker was also discussed. Further exploration of the phenotype-genotype relationship of SCN1A variants may lead to better pharmacological treatments and family guidance.

Variants in ACTC1 underlie distal arthrogryposis accompanied by congenital heart defects.

Contraction of the human sarcomere is the result of interactions between myosin cross-bridges and actin filaments. Pathogenic variants in genes such as MYH7, TPM1, and TNNI3 that encode parts of the cardiac sarcomere cause muscle diseases that affect the heart, such as dilated cardiomyopathy and hypertrophic cardiomyopathy. In contrast, pathogenic variants in homologous genes such as MYH2, TPM2, and TNNI2 that encode parts of the skeletal muscle sarcomere cause muscle diseases affecting skeletal muscle, such as distal arthrogryposis (DA) syndromes and skeletal myopathies. To date, there have been few reports of genes (e.g., MYH7) encoding sarcomeric proteins in which the same pathogenic variant affects skeletal and cardiac muscle. Moreover, none of the known genes underlying DA have been found to contain pathogenic variants that also cause cardiac abnormalities. We report five families with DA because of heterozygous missense variants in the gene actin, alpha, cardiac muscle 1 (ACTC1). ACTC1 encodes a highly conserved actin that binds to myosin in cardiac and skeletal muscle. Pathogenic variants in ACTC1 have been found previously to underlie atrial septal defect, dilated cardiomyopathy, hypertrophic cardiomyopathy, and left ventricular noncompaction. Our discovery delineates a new DA condition because of variants in ACTC1 and suggests that some functions of ACTC1 are shared in cardiac and skeletal muscle.