K acetyltransferase 2B (KAT2B) variants can be responsible for early onset steroid-resistant nephrotic syndrome.

In children, 15% of nephrotic syndromes are steroid-resistant (SRNS); approximately 30% of early onset SRNS have a genetic origin, with more than 100 causal genes described so far. SRNS can be syndromic, if associated with signs and symptoms affecting other organs or systems, such as the central nervous system, the heart or the eyes. Patients with SRNS are at high risk of chronic kidney disease and progressive renal failure, and as such need multidisciplinary care, centred on renal protection. Recently, K acetyltransferase 2B (KAT2B) loss of function was identified as a risk factor for morphological and functional defects in Drosophila nephrocytes; in vitro knockdown of KAT2B also impaired the adhesion and migration ability of human podocytes.Here we provide the first clinical description of a family affected by a loss of function mutation of KAT2B Clinically, both siblings presented with early onset SRNS and bilateral cataract, without neurological or heart defects. Renal function was maintained in the teenage years; nephrotic-range proteinuria was insensitive to immunosuppressive therapies. Therefore, mutations of KAT2B should be sought in patients with unexplained syndromic SRNS affecting the eye.

Classification of PTEN germline non-truncating variants: a new approach to interpretation.

BACKGROUND: PTEN hamartoma tumour syndrome (PHTS) encompasses distinct syndromes, including Cowden syndrome resulting from PTEN pathogenic variants. Missense variants account for 30% of PHTS cases, but their classification remains challenging. To address these difficulties, guidelines were published by the Clinical Genome Resource PTEN Variant Curation Expert Panel. METHODS: Between 2010 and 2020, the Bergonie Institute reference laboratory identified 76 different non-truncating PTEN variants in 166 patients, 17 of which have not previously been reported. Variants were initially classified following the current guidelines. Subsequently, a new classification method was developed based on four main criteria: functional exploration, phenotypic features and familial segregation, in silico modelling, and allelic frequency. RESULTS: This new method of classification is more discriminative and reclassifies 25 variants, including 8 variants of unknown significance. CONCLUSION: This report proposes a revision of the current PTEN variant classification criteria which at present rely on functional tests evaluating only the phosphatase activity of PTEN and apply a particularly stringent clinical PHTS score.The classification of non-truncating variants of PTEN is facilitated by taking into consideration protein stability for variants with intact phosphatase activity, clinical and segregation criteria adapted to the phenotypic variability of PHTS and by specifying the allelic frequency of variants in the general population. This novel method of classification remains to be validated in a prospective cohort.

Christianson syndrome across the lifespan: genetic mutations and longitudinal study in children, adolescents, and adults.

OBJECTIVES: Mutations in the X-linked endosomal Na+/H+ exchanger 6 (NHE6) cause Christianson syndrome (CS). Here, in the largest study to date, we examine genetic diversity and clinical progression in CS into adulthood. METHOD: Data were collected as part of the International Christianson Syndrome and NHE6 (SLC9A6) Gene Network Study. 44 individuals with 31 unique NHE6 mutations, age 2-32 years, were followed prospectively, herein reporting baseline, 1 year follow-up and retrospective natural history. RESULTS: We present data on the CS phenotype with regard to physical growth and adaptive and motor regression across the lifespan including information on mortality. Longitudinal data on body weight and height were examined using a linear mixed model. The rate of growth across development was slow and resulted in prominently decreased age-normed height and weight by adulthood. Adaptive functioning was longitudinally examined; a majority of adult participants (18+ years) lost gross and fine motor skills over a 1 year follow-up. Previously defined core diagnostic criteria for CS (present in>85%)-namely non-verbal status, intellectual disability, epilepsy, postnatal microcephaly, ataxia, hyperkinesia-were universally present in age 6-16; however, an additional core feature of high pain tolerance was added (present in 91%). While neurologic examinations were consistent with cerebellar dysfunction, importantly, a majority of individuals (>50% older than 10) also had corticospinal tract abnormalities. Three participants died during the period of the study. CONCLUSIONS: In this large and longitudinal study of CS, we begin to define the trajectory of symptoms and the adult phenotype thereby identifying critical targets for treatment.

Pathogenic SATB2 missense variants affecting p.Gly392 have variable functional implications and result in diverse clinical phenotypes.

SATB2-associated syndrome (SAS) is caused by pathogenic variants in SATB2, which encodes an evolutionarily conserved transcription factor. Despite the broad range of phenotypic manifestations and variable severity related to this syndrome, haploinsufficiency has been assumed to be the primary molecular explanation.In this study, we describe eight individuals with SATB2 variants that affect p.Gly392 (four women, age range 2-16 years; p.Gly392Arg, p.Gly392Glu and p.Gly392Val). Of these, individuals with p.Gly392Arg substitutions were found to have more severe neurodevelopmental phenotypes based on an established rubric scoring system when compared with individuals with p.Gly392Glu, p.Gly392Val and other previously reported causative SATB2 missense variants. Consistent with the observations at the phenotypic level, using human cell-based and model organism functional data, we documented that while all three described p.Gly392 variants affect the same residue and seem to all have a partial loss-of-function effect, some effects on SATB2 protein function appear to be variant-specific. Our results indicate that genotype-phenotype correlations in SAS are more complex than originally thought, and variant-specific genotype-phenotype correlations are needed.

Novel truncating germline variant reinforces TINF2 as a susceptibility gene for familial non-medullary thyroid cancer.

BACKGROUND: It has long been observed that there are families in which non-medullary thyroid cancer (NMTC) occurs, but few syndromes and genes have been described to date. Proteins in the shelterin complex have been implied in cancer. Here, we have studied shelterin genes in families affected by NMTC (FNMTC). METHODS: We performed whole-exome sequencing (WES) in 10 affected individuals from four families with at least three affected members. Polymerase chain reaction (PCR) and Sanger sequencing were performed to search for variants in the TINF2 gene in 40 FNMTC families. TINF2 transcripts and loss of heterozygosity (LOH) were studied in several affected patients of one family. RESULTS: We found the c.507G>T variant in heterozygosis in the TINF2 gene in one family, co-segregating in all five affected members. This variant affects the normal splicing. LOH was not observed. CONCLUSIONS: Our results reinforce the TINF2 gene as a susceptibility cause of FNMTC suggesting the importance of location of frameshift variants in TINF2. According to our data and previous literature, TINF2 pathogenic variants appear to be a significant risk factor for the development of NMTC and/or melanoma.

Genetic findings in people with schwannomas who do not meet clinical diagnostic criteria for NF2-related schwannomatosis.

BACKGROUND: Most schwannomas are isolated tumours occurring in otherwise healthy people. However, bilateral vestibular schwannomas (BVS) or multiple non-vestibular schwannomas indicate an underlying genetic predisposition. This is most commonly NF2-related schwannomatosis (SWN), but when BVS are absent, this can also indicate SMARCB1-related or LZTR1-related SWN. METHODS: We assessed the variant detection rates for the three major SWN genes (NF2, LZTR1 and SMARCB1) in 154 people, from 150 families, who had at least one non-vestibular schwannoma, but who did not meet clinical criteria for NF2-related SWN at the time of genetic testing. RESULTS: We found that 17 (11%) people from 13 families had a germline SMARCB1 variant and 19 (12%) unrelated individuals had a germline LZTR1 variant. 19 people had an NF2 variant, but 18 of these were mosaic and 17 were only detected when 2 tumours were available for testing. The overall detection rate was 25% using blood alone, but increased to 36% when tumour analysis was included. Another 12 people had a germline variant of uncertain significance (VUS). CONCLUSIONS: There were similar proportions of LZTR1, SMARCB1 or mosaic NF2. However, since an NF2 variant was detected in tumours from 103 people, it is likely that further cases of mosaicism would be detected if more people had additional tumours available for analysis. In addition, if further evidence becomes available to show that the VUSs are pathogenic, this would significantly increase the proportion of people with a genetic diagnosis. Our results indicate the importance of comprehensive genetic testing and improved variant classification.

Autosomal Recessive Hyper-IgE Syndrome in a Child With Beta Thalassemia Trait: A Case Report.

Autoimmune diseases are multifaceted disorders, and their coexistence with other conditions can present unique challenges in diagnosis and management. Here, we report a rare case of autosomal recessive hyper-IgE syndrome (AR-HIES) in a child with beta thalassemia trait. AR-HIES is a distinct immunodeficiency disorder characterized by severe eczema and recurrent bacterial and viral infections, particularly affecting the sinopulmonary system. This case highlights the importance of recognizing and managing the co-occurrence of rare genetic conditions, as it can impact treatment strategies and familial counseling. This unique case of AR-HIES in a child with beta thalassemia trait underscores the complexity of autoimmune disorders and the need for comprehensive evaluation in patients presenting with multiple clinical manifestations.

Expanding the genetic and clinical spectrum of Tatton-Brown-Rahman syndrome in a series of 24 French patients.

BACKGROUND: Tatton-Brown-Rahman syndrome (TBRS; OMIM 615879), also known as DNA methyltransferase 3 alpha (DNMT3A)-overgrowth syndrome (DOS), was first described by Tatton-Brown in 2014. This syndrome is characterised by overgrowth, intellectual disability and distinctive facial features and is the consequence of germline loss-of-function variants in DNMT3A, which encodes a DNA methyltransferase involved in epigenetic regulation. Somatic variants of DNMT3A are frequently observed in haematological malignancies, including acute myeloid leukaemia (AML). To date, 100 individuals with TBRS with de novo germline variants have been described. We aimed to further characterise this disorder clinically and at the molecular level in a nationwide series of 24 French patients and to investigate the correlation between the severity of intellectual disability and the type of variant. METHODS: We collected genetic and medical information from 24 individuals with TBRS using a questionnaire released through the French National AnDDI-Rares Network. RESULTS: Here, we describe the first nationwide French cohort of 24 individuals with germline likely pathogenic/pathogenic variants in DNMT3A, including 17 novel variants. We confirmed that the main phenotypic features were intellectual disability (100% of individuals), distinctive facial features (96%) and overgrowth (87%). We highlighted novel clinical features, such as hypertrichosis, and further described the neurological features and EEG results. CONCLUSION: This study of a nationwide cohort of individuals with TBRS confirms previously published data and provides additional information and clarifies clinical features to facilitate diagnosis and improve care. This study adds value to the growing body of knowledge on TBRS and broadens its clinical and molecular spectrum.

Genetic evidence for splicing-dependent structural and functional plasticity in CASK protein.

BACKGROUND: Pontocerebellar hypoplasia (PCH) may present with supratentorial phenotypes and is often accompanied by microcephaly. Damaging mutations in the X-linked gene CASK produce self-limiting microcephaly with PCH in females but are often lethal in males. CASK deficiency leads to early degeneration of cerebellar granule cells but its role in other regions of the brain remains uncertain. METHOD: We generated a conditional Cask knockout mice and deleted Cask ubiquitously after birth at different times. We examined the clinical features in several subjects with damaging mutations clustered in the central part of the CASK protein. We have performed phylogenetic analysis and RT-PCR to assess the splicing pattern within the same protein region and performed in silico structural analysis to examine the effect of splicing on the CASK's structure. RESULT: We demonstrate that deletion of murine Cask after adulthood does not affect survival but leads to cerebellar degeneration and ataxia over time. Intriguingly, damaging hemizygous CASK mutations in boys who display microcephaly and cerebral dysfunction but without PCH are known. These mutations are present in two vertebrate-specific CASK exons. These exons are subject to alternative splicing both in forebrain and hindbrain. Inclusion of these exons differentially affects the molecular structure and hence possibly the function/s of the CASK C-terminus. CONCLUSION: Loss of CASK function disproportionately affects the cerebellum. Clinical data, however, suggest that CASK may have additional vertebrate-specific function/s that play a role in the mammalian forebrain. Thus, CASK has an ancient function shared between invertebrates and vertebrates as well as novel vertebrate-specific function/s.

Novel mutation leading to splice donor loss in a conserved site of DMD gene causes Duchenne muscular dystrophy with cryptorchidism.

BACKGROUND: As one of the most common congenital abnormalities in male births, cryptorchidism has been found to have a polygenic aetiology according to previous studies of common variants. However, little is known about genetic predisposition of rare variants for cryptorchidism, since rare variants have larger effective size on diseases than common variants. METHODS: In this study, a cohort of 115 Chinese probands with cryptorchidism was analysed using whole-genome sequencing, alongside 19 parental controls and 2136 unaffected men. Additionally, CRISPR-Cas9 editing of a conserved variant was performed in a mouse model, with MRI screening used to observe the phenotype. RESULTS: In 30 of 115 patients (26.1%), we identified four novel genes (ARSH, DMD, MAGEA4 and SHROOM2) affecting at least five unrelated patients and four known genes (USP9Y, UBA1, BCORL1 and KDM6A) with the candidate rare pathogenic variants affecting at least two cases. Burden tests of rare variants revealed the genome-wide significances for newly identified genes (p<2.5×10-6) under the Bonferroni correction. Surprisingly, novel and known genes were mainly found on X chromosome (seven on X and one on Y) and all rare X-chromosomal segregating variants exhibited a maternal inheritance rather than de novo origin. CRISPR-Cas9 mouse modelling of a splice donor loss variant in DMD (NC_000023.11:g.32454661C>G), which resides in a conserved site across vertebrates, replicated bilateral cryptorchidism phenotypes, confirmed by MRI at 4 and 10 weeks. The movement tests further revealed symptoms of Duchenne muscular dystrophy (DMD) in transgenic mice. CONCLUSION: Our results revealed the role of the DMD gene mutation in causing cryptorchidism. The results also suggest that maternal-X inheritance of pathogenic defects could have a predominant role in the development of cryptorchidism.

Multimorbidity disease clusters are associated with venous thromboembolism: an extended cross-sectional national study.

Multimorbidity, i.e., two or more non-communicable diseases (NCDs), is an escalating challenge for society. Venous thromboembolism (VTE) is a common cardiovascular disease and it is unknown which multimorbidity clusters associates with VTE. Our aim was to examine the association between different common disease clusters of multimorbidity and VTE. The study is an extended (1997-2015) cross-sectional Swedish study using the National Patient Register and the Multigeneration Register. A total of 2,694,442 Swedish-born individuals were included in the study. Multimorbidity was defined by 45 NCDs. A principal component analysis (PCA) identified multimorbidity disease clusters. Odds ratios (OR) for VTE were calculated for the different multimorbidity disease clusters. There were 16% (n = 440,742) of multimorbid individuals in the study population. Forty-four of the individual 45 NCDs were associated with VTE. The PCA analysis identified nine multimorbidity disease clusters, F1-F9. Seven of these multimorbidity clusters were associated with VTE. The adjusted OR for VTE in the multimorbid patients was for the first three clusters: F1 (cardiometabolic diseases) 3.44 (95%CI 3.24-3.65), F2 (mental disorders) 2.25 (95%CI 2.14-2.37) and F3 (digestive system diseases) 4.35 (95%CI 3.63-5.22). There was an association between multimorbidity severity and OR for VTE. For instance, the occurrence of at least five diseases was in F1 and F2 associated with ORs for VTE: 8.17 (95%CI 6.32-10.55) and 6.31 (95%CI 4.34-9.17), respectively. In this nationwide study we have shown a strong association between VTE and different multimorbidity disease clusters that might be useful for VTE prediction.

Survey of service needs to embed genome sequencing for motor neuron disease in neurology in the English National Health Service.

All people with motor neuron disease (pwMND) in England are eligible for genome sequencing (GS), with panel-based testing. With the advent of genetically targeted MND treatments, and increasing demand for GS, it is important that clinicians have the knowledge and skills to support pwMND in making informed decisions around GS. We undertook an online survey of clinical genomic knowledge and genetic counselling skills in English clinicians who see pwMND. There were 245 respondents to the survey (160 neurology clinicians and 85 genetic clinicians). Neurology clinicians reported multiple, overlapping barriers to offering pwMND GS. Lack of time to discuss GS in clinic and lack of training in genetics were reported. Neurology clinicians scored significantly less well on self-rated genomic knowledge and genetic counselling skills than genetic clinicians. The majority of neurology clinicians reported that they do not have adequate educational or patient information resources to support GS discussions. We identify low levels of genomic knowledge and skills in the neurology workforce. This may impede access to GS and precision medicine for pwMND.

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.

Diagnosis and management in Rubinstein-Taybi syndrome: first international consensus statement.

Rubinstein-Taybi syndrome (RTS) is an archetypical genetic syndrome that is characterised by intellectual disability, well-defined facial features, distal limb anomalies and atypical growth, among numerous other signs and symptoms. It is caused by variants in either of two genes (CREBBP, EP300) which encode for the proteins CBP and p300, which both have a function in transcription regulation and histone acetylation. As a group of international experts and national support groups dedicated to the syndrome, we realised that marked heterogeneity currently exists in clinical and molecular diagnostic approaches and care practices in various parts of the world. Here, we outline a series of recommendations that document the consensus of a group of international experts on clinical diagnostic criteria for types of RTS (RTS1: CREBBP; RTS2: EP300), molecular investigations, long-term management of various particular physical and behavioural issues and care planning. The recommendations as presented here will need to be evaluated for improvements to allow for continued optimisation of diagnostics and care.

Dissecting genetic architecture of rare dystonia: genetic, molecular and clinical insights.

BACKGROUND: Dystonia is one of the most common movement disorders. To date, the genetic causes of dystonia in populations of European descent have been extensively studied. However, other populations, particularly those from the Middle East, have not been adequately studied. The purpose of this study is to discover the genetic basis of dystonia in a clinically and genetically well-characterised dystonia cohort from Turkey, which harbours poorly studied populations. METHODS: Exome sequencing analysis was performed in 42 Turkish dystonia families. Using co-expression network (CEN) analysis, identified candidate genes were interrogated for the networks including known dystonia-associated genes and genes further associated with the protein-protein interaction, animal model-based characteristics and clinical findings. RESULTS: We identified potentially disease-causing variants in the established dystonia genes (PRKRA, SGCE, KMT2B, SLC2A1, GCH1, THAP1, HPCA, TSPOAP1, AOPEP; n=11 families (26%)), in the uncommon forms of dystonia-associated genes (PCCB, CACNA1A, ALDH5A1, PRKN; n=4 families (10%)) and in the candidate genes prioritised based on the pathogenicity of the variants and CEN-based analyses (n=11 families (21%)). The diagnostic yield was found to be 36%. Several pathways and gene ontologies implicated in immune system, transcription, metabolic pathways, endosomal-lysosomal and neurodevelopmental mechanisms were over-represented in our CEN analysis. CONCLUSIONS: Here, using a structured approach, we have characterised a clinically and genetically well-defined dystonia cohort from Turkey, where dystonia has not been widely studied, and provided an uncovered genetic basis, which will facilitate diagnostic dystonia research.

Bi-allelic variants in chromatoid body protein TDRD6 cause spermiogenesis defects and severe oligoasthenoteratozoospermia in humans.

BACKGROUND: The association between the TDRD6 variants and human infertility remains unclear, as only one homozygous missense variant of TDRD6 was found to be associated with oligoasthenoteratozoospermia (OAT). METHODS: Whole-exome sequencing and Sanger sequencing were employed to identify potential pathogenic variants of TDRD6 in infertile men. Histology, immunofluorescence, immunoblotting and ultrastructural analyses were conducted to clarify the structural and functional abnormalities of sperm in mutated patients. Tdrd6-knockout mice were generated using the CRISPR-Cas9 system. Total RNA-seq and single-cell RNA-seq (scRNA-seq) analyses were used to elucidate the underlying molecular mechanisms, followed by validation through quantitative RT-PCR and immunostaining. Intracytoplasmic sperm injection (ICSI) was also used to assess the efficacy of clinical treatment. RESULTS: Bi-allelic TDRD6 variants were identified in five unrelated Chinese individuals with OAT, including homozygous loss-of-function variants in two consanguineous families. Notably, besides reduced concentrations and impaired motility, a significant occurrence of acrosomal hypoplasia was detected in multiple spermatozoa among five patients. Using the Tdrd6-deficient mice, we further elucidate the pivotal role of TDRD6 in spermiogenesis and acrosome identified. In addition, the mislocalisation of crucial chromatoid body components DDX4 (MVH) and UPF1 was also observed in round spermatids from patients harbouring TDRD6 variants. ScRNA-seq analysis of germ cells from a patient with TDRD6 variants revealed that TDRD6 regulates mRNA metabolism processes involved in spermatid differentiation and cytoplasmic translation. CONCLUSION: Our findings strongly suggest that TDRD6 plays a conserved role in spermiogenesis and confirms the causal relationship between TDRD6 variants and human OAT. Additionally, this study highlights the unfavourable ICSI outcomes in individuals with bi-allelic TDRD6 variants, providing insights for potential clinical treatment strategies.

De novo heterozygous missense variants in CELSR1 as cause of fetal pleural effusions and progressive fetal hydrops.

Fetal hydrops as detected by prenatal ultrasound usually carries a poor prognosis depending on the underlying aetiology. We describe the prenatal and postnatal clinical course of two unrelated female probands in whom de novo heterozygous missense variants in the planar cell polarity gene CELSR1 were detected using exome sequencing. Using several in vitro assays, we show that the CELSR1 p.(Cys1318Tyr) variant disrupted the subcellular localisation, affected cell-cell junction, impaired planar cell polarity signalling and lowered proliferation rate. These observations suggest that deleterious rare CELSR1 variants could be a possible cause of fetal hydrops.

Genotype and phenotype correlation of PHACTR1-related neurological disorders.

BACKGROUND: PHACTR1 (phosphatase and actin regulators) plays a key role in cortical migration and synaptic activity by binding and regulating G-actin and PPP1CA. This study aimed to expand the genotype and phenotype of patients with de novo variants in PHACTR1 and analyse the impact of variants on protein-protein interaction. METHODS: We identified seven patients with PHACTR1 variants by trio-based whole-exome sequencing. Additional two subjects were ascertained from two centres through GeneMatcher. The genotype-phenotype correlation was determined, and AlphaFold-Multimer was used to predict protein-protein interactions and interfaces. RESULTS: Eight individuals carried missense variants and one had CNV in the PHACTR1. Infantile epileptic spasms syndrome (IESS) was the unifying phenotype in eight patients with missense variants of PHACTR1. They could present with other types of seizures and often exhibit drug-resistant epilepsy with a poor prognosis. One patient with CNV displayed a developmental encephalopathy phenotype. Using AlphaFold-Multimer, our findings indicate that PHACTR1 and G-actin-binding sequences overlap with PPP1CA at the RPEL3 domain, which suggests possible competition between PPP1CA and G-actin for binding to PHACTR1 through a similar polymerisation interface. In addition, patients carrying missense variants located at the PHACTR1-PPP1CA or PHACTR1-G-actin interfaces consistently exhibit the IESS phenotype. These missense variants are mostly concentrated in the overlapping sequence (RPEL3 domain). CONCLUSIONS: Patients with variants in PHACTR1 can have a phenotype of developmental encephalopathy in addition to IESS. Moreover, our study confirmed that the variants affect the binding of PHACTR1 to G-actin or PPP1CA, resulting in neurological disorders in patients.