Pathogenic variants in HGF give rise to childhood-to-late onset primary lymphoedema by loss of function.

Developmental and functional defects in the lymphatic system are responsible for primary lymphoedema (PL). PL is a chronic debilitating disease caused by increased accumulation of interstitial fluid, predisposing to inflammation, infections and fibrosis. There is no cure, only symptomatic treatment is available. Thirty-two genes or loci have been linked to PL, and another 22 are suggested, including Hepatocyte Growth Factor (HGF). We searched for HGF variants in 770 index patients from the Brussels PL cohort. We identified ten variants predicted to cause HGF loss-of-function (six nonsense, two frameshifts, and two splice-site changes; 1.3% of our cohort), and 14 missense variants predicted to be pathogenic in 17 families (2.21%). We studied co-segregation within families, mRNA stability for non-sense variants, and in vitro functional effects of the missense variants. Analyses of the mRNA of patient cells revealed degradation of the nonsense mutant allele. Reduced protein secretion was detected for nine of the 14 missense variants expressed in COS-7 cells. Stimulation of lymphatic endothelial cells with these 14 HGF variant proteins resulted in decreased activation of the downstream targets AKT and ERK1/2 for three of them. Clinically, HGF-associated PL was diverse, but predominantly bilateral in the lower limbs with onset varying from early childhood to adulthood. Finally, aggregation study in a second independent cohort underscored that rare likely pathogenic variants in HGF explain about 2% of PL. Therefore, HGF signalling seems crucial for lymphatic development and/or maintenance in human beings and HGF should be included in diagnostic genetic screens for PL.

Biallelic variants in Plexin B2 (PLXNB2) cause amelogenesis imperfecta, hearing loss and intellectual disability.

BACKGROUND: Plexins are large transmembrane receptors for the semaphorin family of signalling proteins. Semaphorin-plexin signalling controls cellular interactions that are critical during development as well as in adult life stages. Nine plexin genes have been identified in humans, but despite the apparent importance of plexins in development, only biallelic PLXND1 and PLXNA1 variants have so far been associated with Mendelian genetic disease. METHODS: Eight individuals from six families presented with a recessively inherited variable clinical condition, with core features of amelogenesis imperfecta (AI) and sensorineural hearing loss (SNHL), with variable intellectual disability. Probands were investigated by exome or genome sequencing. Common variants and those unlikely to affect function were excluded. Variants consistent with autosomal recessive inheritance were prioritised. Variant segregation analysis was performed by Sanger sequencing. RNA expression analysis was conducted in C57Bl6 mice. RESULTS: Rare biallelic pathogenic variants in plexin B2 (PLXNB2), a large transmembrane semaphorin receptor protein, were found to segregate with disease in all six families. The variants identified include missense, nonsense, splicing changes and a multiexon deletion. Plxnb2 expression was detected in differentiating ameloblasts. CONCLUSION: We identify rare biallelic pathogenic variants in PLXNB2 as a cause of a new autosomal recessive, phenotypically diverse syndrome with AI and SNHL as core features. Intellectual disability, ocular disease, ear developmental abnormalities and lymphoedema were also present in multiple cases. The variable syndromic human phenotype overlaps with that seen in Plxnb2 knockout mice, and, together with the rarity of human PLXNB2 variants, may explain why pathogenic variants in PLXNB2 have not been reported previously.

Guidelines for the diagnosis and management of adult aplastic anaemia: A British Society for Haematology Guideline.

Pancytopenia with hypocellular bone marrow is the hallmark of aplastic anaemia (AA) and the diagnosis is confirmed after careful evaluation, following exclusion of alternate diagnosis including hypoplastic myelodysplastic syndromes. Emerging use of molecular cyto-genomics is helpful in delineating immune mediated AA from inherited bone marrow failures (IBMF). Camitta criteria is used to assess disease severity, which along with age and availability of human leucocyte antigen compatible donor are determinants for therapeutic decisions. Supportive care with blood and platelet transfusion support, along with anti-microbial prophylaxis and prompt management of opportunistic infections remain key throughout the disease course. The standard first-line treatment for newly diagnosed acquired severe/very severe AA patients is horse anti-thymocyte globulin and ciclosporin-based immunosuppressive therapy (IST) with eltrombopag or allogeneic haemopoietic stem cell transplant (HSCT) from a matched sibling donor. Unrelated donor HSCT in adults should be considered after lack of response to IST, and up front for young adults with severe infections and a readily available matched unrelated donor. Management of IBMF, AA in pregnancy and in elderly require special attention. In view of the rarity of AA and complexity of management, appropriate discussion in multidisciplinary meetings and involvement of expert centres is strongly recommended to improve patient outcomes.

Redefining WILD syndrome: a primary lymphatic dysplasia with congenital multisegmental lymphoedema, cutaneous lymphovascular malformation, CD4 lymphopaenia and warts.

BACKGROUND: Primary lymphoedema (PL) syndromes are increasingly recognised as presentations of complex genetic disease, with at least 20 identified causative genes. Recognition of clinical patterns is key to diagnosis, research and therapeutics. The defining criteria for one such clinical syndrome, 'WILD syndrome' (Warts, Immunodeficiency, Lymphoedema and anogenital Dysplasia), have previously depended on a single case report. METHODS AND RESULTS: We present 21 patients (including the first described case) with similar clinical and immunological phenotypes. All had PL affecting multiple segments, with systemic involvement (intestinal lymphangiectasia/pleural or pericardial effusions) in 70% (n=14/20). Most (n=20, 95%) had a distinctive cutaneous lymphovascular malformation on the upper anterior chest wall. Some (n=10, 48%) also had hyperpigmented lesions resembling epidermal naevi (but probably lymphatic in origin). Warts were common (n=17, 81%) and often refractory. In contrast to the previous case report, anogenital dysplasia was uncommon-only found in two further cases (total n=3, 14%). Low CD4 counts and CD4:CD8 ratios typified the syndrome (17 of 19, 89%), but monocyte counts were universally normal, unlike GATA2 deficiency. CONCLUSION: WILD syndrome is a previously unrecognised, underdiagnosed generalised PL syndrome. Based on this case series, we redefine WILD as 'Warts, Immunodeficiency, andLymphatic Dysplasia' and suggest specific diagnostic criteria. The essential criterion is congenital multisegmental PL in a 'mosaic' distribution. The major diagnostic features are recurrent warts, cutaneous lymphovascular malformations, systemic involvement (lymphatic dysplasia), genital swelling and CD4 lymphopaenia with normal monocyte counts. The absence of family history suggests a sporadic condition, and the random distribution of swelling implicates mosaic postzygotic mutation as the cause.

Overlapping cortical malformations in patients with pathogenic variants in GRIN1 and GRIN2B.

BACKGROUND: Malformations of cortical development (MCDs) have been reported in a subset of patients with pathogenic heterozygous variants in GRIN1 or GRIN2B, genes which encode for subunits of the N-methyl-D-aspartate receptor (NMDAR). The aim of this study was to further define the phenotypic spectrum of NMDAR-related MCDs. METHODS: We report the clinical, radiological and molecular features of 7 new patients and review data on 18 previously reported individuals with NMDAR-related MCDs. Neuropathological findings for two individuals with heterozygous variants in GRIN1 are presented. We report the clinical and neuropathological features of one additional individual with homozygous pathogenic variants in GRIN1. RESULTS: Heterozygous variants in GRIN1 and GRIN2B were associated with overlapping severe clinical and imaging features, including global developmental delay, epilepsy, diffuse dysgyria, dysmorphic basal ganglia and hippocampi. Neuropathological examination in two fetuses with heterozygous GRIN1 variants suggests that proliferation as well as radial and tangential neuronal migration are impaired. In addition, we show that neuronal migration is also impaired by homozygous GRIN1 variants in an individual with microcephaly with simplified gyral pattern. CONCLUSION: These findings expand our understanding of the clinical and imaging features of the 'NMDARopathy' spectrum and contribute to our understanding of the likely underlying pathogenic mechanisms leading to MCD in these patients.

Short amplicon reverse transcription-polymerase chain reaction detects aberrant splicing in genes with low expression in blood missed by ribonucleic acid sequencing analysis for clinical diagnosis.

Use of blood RNA sequencing (RNA-seq) as a splicing analysis tool for clinical interpretation of variants of uncertain significance (VUSs) found via whole-genome and exome sequencing can be difficult for genes that have low expression in the blood due to insufficient read count coverage aligned to specific genes of interest. Here, we present a short amplicon reverse transcription-polymerase chain reaction(RT-PCR) for the detection of genes with low blood expression. Short amplicon RT-PCR, is designed to span three exons where an exon harboring a variant is flanked by one upstream and one downstream exon. We tested short amplicon RT-PCRs for genes that have median transcripts per million (TPM) values less than one according to the genotype-tissue expression database. Median TPM values of genes analyzed in this study are SYN1 = 0.8549, COL1A1 = 0.6275, TCF4 = 0.4009, DSP = .2894, TTN = 0.2851, COL5A2 = 0.1036, TERT = 0.04452, NTRK2 = 0.0344, ABCA4 = 0.00744, PRPH = 0, and WT1 = 0. All these genes show insufficient exon-spanning read coverage in our RNA-seq data to allow splicing analysis. We successfully detected all genes tested except PRPH and WT1. Aberrant splicing was detected in SYN1, TCF4, NTRK2, TTN, and TERT VUSs. Therefore, our results show short amplicon RT-PCR is a useful alternative for the analysis of splicing events in genes with low TPM in blood RNA for clinical diagnostics.

The seventh international RASopathies symposium: Pathways to a cure-expanding knowledge, enhancing research, and therapeutic discovery.

RASopathies are a group of genetic disorders that are caused by genes that affect the canonical Ras/mitogen-activated protein kinase (MAPK) signaling pathway. Despite tremendous progress in understanding the molecular consequences of these genetic anomalies, little movement has been made in translating these findings to the clinic. This year, the seventh International RASopathies Symposium focused on expanding the research knowledge that we have gained over the years to enhance new discoveries in the field, ones that we hope can lead to effective therapeutic treatments. Indeed, for the first time, research efforts are finally being translated to the clinic, with compassionate use of Ras/MAPK pathway inhibitors for the treatment of RASopathies. This biannual meeting, organized by the RASopathies Network, brought together basic scientists, clinicians, clinician scientists, patients, advocates, and their families, as well as representatives from pharmaceutical companies and the National Institutes of Health. A history of RASopathy gene discovery, identification of new disease genes, and the latest research, both at the bench and in the clinic, were discussed.

SHORT Syndrome: Systematic Appraisal of the Medical and Dental Phenotype.

INTRODUCTION: SHORT syndrome is a rare autosomal dominant condition described by its acronym of short stature, hyperextensibility of joints and/or inguinal hernia, ocular depression, Rieger abnormality, and teething delay. Individuals have a distinct progeroid craniofacial appearance with a triangular face, frontal bossing, hypoplastic or thin alae nasi, large low-set ears, and mandibular retrognathia. OBJECTIVES: To systematically appraise the literature and update the clinical phenotype with emphasis on the dental condition. DESIGN: A systematic literature search was carried out to update the clinical phenotype, identifying reports of individuals with SHORT syndrome published after August 2015. The same search strategy but not limited to publication date was carried out to identify reports of the dental phenotype. Two independent reviewers screened 1937 articles with 55 articles identified for full-text review. RESULTS: Nineteen individuals from 11 families were identified. Facial dysmorphism including ocular depression, triangular shaped face, frontal bossing, large low-set ears, and micrognathia were the most consistent features followed by lipodystrophy, insulin resistance, and intrauterine growth restriction. Teething delay, microdontia, hypodontia, and enamel hypoplasia have all been reported. CONCLUSION: Features that comprise the SHORT acronym do not accurately or completely describe the clinical phenotype. The craniofacial appearance is one of the most consistent features. Lipodystrophy and insulin resistance may also be considered cardinal features. After teething delay, enamel hypoplasia and microdontia are the most common dental manifestations. We present recommendations for the dental and orthodontic/orthognathic management of individuals with SHORT syndrome.

Janus-faced EPHB4-associated disorders: novel pathogenic variants and unreported intrafamilial overlapping phenotypes.

PURPOSE: Several clinical phenotypes including fetal hydrops, central conducting lymphatic anomaly or capillary malformations with arteriovenous malformations 2 (CM-AVM2) have been associated with EPHB4 (Ephrin type B receptor 4) variants, demanding new approaches for deciphering pathogenesis of novel variants of uncertain significance (VUS) identified in EPHB4, and for the identification of differentiated disease mechanisms at the molecular level. METHODS: Ten index cases with various phenotypes, either fetal hydrops, CM-AVM2, or peripheral lower limb lymphedema, whose distinct clinical phenotypes are described in detail in this study, presented with a variant in EPHB4. In vitro functional studies were performed to confirm pathogenicity. RESULTS: Pathogenicity was demonstrated for six of the seven novel EPHB4 VUS investigated. A heterogeneity of molecular disease mechanisms was identified, from loss of protein production or aberrant subcellular localization to total reduction of the phosphorylation capability of the receptor. There was some phenotype-genotype correlation; however, previously unreported intrafamilial overlapping phenotypes such as lymphatic-related fetal hydrops (LRFH) and CM-AVM2 in the same family were observed. CONCLUSION: This study highlights the usefulness of protein expression and subcellular localization studies to predict EPHB4 variant pathogenesis. Our accurate clinical phenotyping expands our interpretation of the Janus-faced spectrum of EPHB4-related disorders, introducing the discovery of cases with overlapping phenotypes.

Update and audit of the St George's classification algorithm of primary lymphatic anomalies: a clinical and molecular approach to diagnosis.

Primary lymphatic anomalies may present in a myriad of ways and are highly heterogenous. Careful consideration of the presentation can lead to an accurate clinical and/or molecular diagnosis which will assist with management. The most common presentation is lymphoedema, swelling resulting from failure of the peripheral lymphatic system. However, there may be internal lymphatic dysfunction, for example, chylous reflux, or lymphatic malformations, including the thorax or abdomen. A number of causal germline or postzygotic gene mutations have been discovered. Some through careful phenotyping and categorisation of the patients based on the St George's classification pathway/algorithm. The St George's classification algorithm is aimed at providing an accurate diagnosis for patients with lymphoedema based on age of onset, areas affected by swelling and associated clinical features. This has enabled the identification of new causative genes. This update brings the classification of primary lymphatic disorders in line with the International Society for the Study of Vascular Anomalies 2018 classification for vascular anomalies. The St George's algorithm considers combined vascular malformations and primary lymphatic anomalies. It divides the types of primary lymphatic anomalies into lymphatic malformations and primary lymphoedema. It further divides the primary lymphoedema into syndromic, generalised lymphatic dysplasia with internal/systemic involvement, congenital-onset lymphoedema and late-onset lymphoedema. An audit and update of the algorithm has revealed where new genes have been discovered and that a molecular diagnosis was possible in 26% of all patients overall and 41% of those tested.

The role of automated breast ultrasound in the assessment of the local extent of breast cancer.

Automated breast ultrasound (ABUS) is a non-invasive advanced ultrasound modality. The degree of extension of the cancer within the breast is very important to choose the appropriate kind of surgery/therapy. In the current work, the aim was to evaluate the role of the ABUS in the assessment of the local extent of the breast cancer before management. This is a prospective analysis that studied 562 female patients with proved breast cancers. Evaluation was in regard of the size, multiplicity, and the stromal invasion (ie, the presence of tumor emboli or tumor masses within the stroma of the breast tissue) around the tumor. Cases were subjected to automated breast ultrasound performed in the axial and coronal planes. ABUS showed high accuracy of assessment of the tumor multiplicity (82.2%) and the stromal involvement (93.5%). There was a statistical significance (P < .001) between the ABUS and the pathology regarding the measurement of the size of the index cancer. In conclusion, ABUS could be used for determination of the intramammary extend of the breast cancer. ABUS provided accurate assessment of the peritumor stromal involvement and multiplicity of the cancer which is required to choose the proper choice of surgery.

Goldberg-Shprintzen syndrome is determined by the absence, or reduced expression levels, of KIFBP.

Goldberg-Shprintzen syndrome (GOSHS) is caused by loss of function variants in the kinesin binding protein gene (KIFBP). However, the phenotypic range of this syndrome is wide, indicating that other factors may play a role. To date, 37 patients with GOSHS have been reported. Here, we document nine new patients with variants in KIFBP: seven with nonsense variants and two with missense variants. To our knowledge, this is the first time that missense variants have been reported in GOSHS. We functionally investigated the effect of the variants identified, in an attempt to find a genotype-phenotype correlation. We also determined whether common Hirschsprung disease (HSCR)-associated single nucleotide polymorphisms (SNPs), could explain the presence of HSCR in GOSHS. Our results showed that the missense variants led to reduced expression of KIFBP, while the truncating variants resulted in lack of protein. However, no correlation was found between the severity of GOSHS and the location of the variants. We were also unable to find a correlation between common HSCR-associated SNPs, and HSCR development in GOSHS. In conclusion, we show that reduced, as well as lack of KIFBP expression can lead to GOSHS, and our results suggest that a threshold expression of KIFBP may modulate phenotypic variability of the disease.

Detection of structural mosaicism from targeted and whole-genome sequencing data.

Structural mosaic abnormalities are large post-zygotic mutations present in a subset of cells and have been implicated in developmental disorders and cancer. Such mutations have been conventionally assessed in clinical diagnostics using cytogenetic or microarray testing. Modern disease studies rely heavily on exome sequencing, yet an adequate method for the detection of structural mosaicism using targeted sequencing data is lacking. Here, we present a method, called MrMosaic, to detect structural mosaic abnormalities using deviations in allele fraction and read coverage from next-generation sequencing data. Whole-exome sequencing (WES) and whole-genome sequencing (WGS) simulations were used to calculate detection performance across a range of mosaic event sizes, types, clonalities, and sequencing depths. The tool was applied to 4911 patients with undiagnosed developmental disorders, and 11 events among nine patients were detected. For eight of these 11 events, mosaicism was observed in saliva but not blood, suggesting that assaying blood alone would miss a large fraction, possibly >50%, of mosaic diagnostic chromosomal rearrangements.

Expanding the phenotypic spectrum consequent upon de novo WDR37 missense variants.

Structural eye disorders are increasingly recognised as having a genetic basis, although current genetic testing is limited in its success. De novo missense variants in WDR37 are a recently described cause of a multisystemic syndromic disorder featuring ocular coloboma. This study characterises the phenotypic spectrum of this disorder and reports 2 de novo heterozygous variants (p.Thr115Ile, p.Ser119Tyr) in three unrelated Caucasian individuals. All had a clinical phenotype consisting of bilateral iris and retinal coloboma, developmental delay and additional, variable multisystem features. The variants fall within a highly conserved region upstream of the WD-repeat domains, within an apparent mutation cluster. Consistent with the literature, intellectual disability, structural eye disorders, epilepsy, congenital heart disease, genitorenal anomalies and dysmorphic facial features were observed. In addition, a broader developmental profile is reported with a more specific musculoskeletal phenotype described in association with the novel variant (p.Thr115Ile). We further expand the phenotypic spectrum of WDR37-related disorders to include those with milder developmental delay and strengthen the association of ocular coloboma and musculoskeletal features. We promote the inclusion of WDR37 on gene panels for intellectual disability, epilepsy and structural eye disorders.

Abnormal Rab11-Rab8-vesicles cluster in enterocytes of patients with microvillus inclusion disease.

Microvillus inclusion disease (MVID) is a congenital enteropathy characterized by accumulation of vesiculo-tubular endomembranes in the subapical cytoplasm of enterocytes, historically termed "secretory granules." However, neither their identity nor pathophysiological significance is well defined. Using immunoelectron microscopy and tomography, we studied biopsies from MVID patients (3× Myosin 5b mutations and 1× Syntaxin3 mutation) and compared them to controls and genome-edited CaCo2 cell models, harboring relevant mutations. Duodenal biopsies from 2 patients with novel Myosin 5b mutations and typical clinical symptoms showed unusual ultrastructural phenotypes: aberrant subapical vesicles and tubules were prominent in the enterocytes, though other histological hallmarks of MVID were almost absent (ectopic intra-/intercellular microvilli, brush border atrophy). We identified these enigmatic vesiculo-tubular organelles as Rab11-Rab8-positive recycling compartments of altered size, shape and location harboring the apical SNARE Syntaxin3, apical transporters sodium-hydrogen exchanger 3 (NHE3) and cystic fibrosis transmembrane conductance regulator. Our data strongly indicate that in MVID disrupted trafficking between cargo vesicles and the apical plasma membrane is the primary cause of a defect of epithelial polarity and subsequent facultative loss of brush border integrity, leading to malabsorption. Furthermore, they support the notion that mislocalization of transporters, such as NHE3 substantially contributes to the reported sodium loss diarrhea.

Null variants and deletions in BRWD3 cause an X-linked syndrome of mild-moderate intellectual disability, macrocephaly, and obesity: A series of 17 patients.

BRWD3 has been described as a cause of X-linked intellectual disability, but relatively little is known about the specific phenotype. We report the largest BRWD3 patient series to date, comprising 17 males with 12 distinct null variants and 2 partial gene deletions. All patients presented with intellectual disability, which was classified as moderate (65%) or mild (35%). Behavioral issues were present in 75% of patients, including aggressive behavior, attention deficit/hyperactivity and/or autistic spectrum disorders. Mean head circumference was +2.8 SD (2.8 standard deviations above the mean), and mean BMI was +2.0 SD (in the context of a mean height of +1.3 SD), indicating a predominant macrocephaly/obesity phenotype. Shared facial features included a tall chin, prognathism, broad forehead, and prominent supraorbital ridge. Additional features, reported in a minority (<30%) of patients included cryptorchidism, neonatal hypotonia, and small joint hypermobility. This study delineates the clinical features associated with BRWD3 null variants and partial gene deletions, and suggests that BRWD3 should be included in the differential diagnosis of patients with an overgrowth-intellectual disability (OGID) phenotype, particularly in male patients with a mild or moderate intellectual disability associated with macrocephaly and/or obesity.

The phenotype of Sotos syndrome in adulthood: A review of 44 individuals.

Sotos syndrome is an overgrowth-intellectual disability (OGID) syndrome caused by NSD1 pathogenic variants and characterized by a distinctive facial appearance, an intellectual disability, tall stature and/or macrocephaly. Other associated clinical features include scoliosis, seizures, renal anomalies, and cardiac anomalies. However, many of the published Sotos syndrome clinical descriptions are based on studies of children; the phenotype in adults with Sotos syndrome is not yet well described. Given that it is now 17 years since disruption of NSD1 was shown to cause Sotos syndrome, many of the children first reported are now adults. It is therefore timely to investigate the phenotype of 44 adults with Sotos syndrome and NSD1 pathogenic variants. We have shown that adults with Sotos syndrome display a wide spectrum of intellectual ability with functioning ranging from fully independent to fully dependent. Reproductive rates are low. In our cohort, median height in adult women is +1.9 SD and men +0.5 SD. There is a distinctive facial appearance in adults with a tall, square, prominent chin. Reassuringly, adults with Sotos syndrome are generally healthy with few new medical issues; however, lymphedema, poor dentition, hearing loss, contractures and tremor have developed in a small number of individuals.

PURA syndrome: clinical delineation and genotype-phenotype study in 32 individuals with review of published literature.

BACKGROUND: De novo mutations in PURA have recently been described to cause PURA syndrome, a neurodevelopmental disorder characterised by severe intellectual disability (ID), epilepsy, feeding difficulties and neonatal hypotonia. OBJECTIVES: To delineate the clinical spectrum of PURA syndrome and study genotype-phenotype correlations. METHODS: Diagnostic or research-based exome or Sanger sequencing was performed in individuals with ID. We systematically collected clinical and mutation data on newly ascertained PURA syndrome individuals, evaluated data of previously reported individuals and performed a computational analysis of photographs. We classified mutations based on predicted effect using 3D in silico models of crystal structures of Drosophila-derived Pur-alpha homologues. Finally, we explored genotype-phenotype correlations by analysis of both recurrent mutations as well as mutation classes. RESULTS: We report mutations in PURA (purine-rich element binding protein A) in 32 individuals, the largest cohort described so far. Evaluation of clinical data, including 22 previously published cases, revealed that all have moderate to severe ID and neonatal-onset symptoms, including hypotonia (96%), respiratory problems (57%), feeding difficulties (77%), exaggerated startle response (44%), hypersomnolence (66%) and hypothermia (35%). Epilepsy (54%) and gastrointestinal (69%), ophthalmological (51%) and endocrine problems (42%) were observed frequently. Computational analysis of facial photographs showed subtle facial dysmorphism. No strong genotype-phenotype correlation was identified by subgrouping mutations into functional classes. CONCLUSION: We delineate the clinical spectrum of PURA syndrome with the identification of 32 additional individuals. The identification of one individual through targeted Sanger sequencing points towards the clinical recognisability of the syndrome. Genotype-phenotype analysis showed no significant correlation between mutation classes and disease severity.

Human phenotypes caused by PIEZO1 mutations; one gene, two overlapping phenotypes?

PIEZO1 is a large mechanosensitive ion channel protein. Diseases associated with PIEZO1 include autosomal recessive generalised lymphatic dysplasia of Fotiou (GLDF) and autosomal dominant dehydrated hereditary stomatocytosis with or without pseudohyperkalemia and/or perinatal oedema (DHS). The two disorders show overlapping features, fetal hydrops/perinatal oedema have been reported in both. Electrophysiological studies suggest opposite mechanisms of action: the mutations identified in GLDF patients cause a loss-of-function mechanism of disease and mutations in DHS patients cause gain of function. This raises the question: Is the pathogenic disease mechanism behind the fetal oedema the same in the two phenotypes? In this Symposium Review, we will discuss the two conditions and highlight key questions that remain to be answered. For instance, the perinatal oedema often resolves soon after birth and we are still at a loss to understand why. Are there any mechanisms which could compensate for the faulty PIEZO1 in these patients? Are there physiological changes at birth that are less reliant on the function of PIEZO1? Thus, there is a clear need for further studies into the two disorders, in order to fully understand the role of PIEZO1 in health and disease.

Rapid prenatal diagnosis using targeted exome sequencing: a cohort study to assess feasibility and potential impact on prenatal counseling and pregnancy management.

PURPOSE: Unexpected fetal abnormalities occur in 2-5% of pregnancies. While traditional cytogenetic and microarray approaches achieve diagnosis in around 40% of cases, lack of diagnosis in others impedes parental counseling, informed decision making, and pregnancy management. Postnatally exome sequencing yields high diagnostic rates, but relies on careful phenotyping to interpret genotype results. Here we used a multidisciplinary approach to explore the utility of rapid fetal exome sequencing for prenatal diagnosis using skeletal dysplasias as an exemplar. METHODS: Parents in pregnancies undergoing invasive testing because of sonographic fetal abnormalities, where multidisciplinary review considered skeletal dysplasia a likely etiology, were consented for exome trio sequencing (both parents and fetus). Variant interpretation focused on a virtual panel of 240 genes known to cause skeletal dysplasias. RESULTS: Definitive molecular diagnosis was made in 13/16 (81%) cases. In some cases, fetal ultrasound findings alone were of sufficient severity for parents to opt for termination. In others, molecular diagnosis informed accurate prediction of outcome, improved parental counseling, and enabled parents to terminate or continue the pregnancy with certainty. CONCLUSION: Trio sequencing with expert multidisciplinary review for case selection and data interpretation yields timely, high diagnostic rates in fetuses presenting with unexpected skeletal abnormalities. This improves parental counseling and pregnancy management.