Bi-allelic variants in the ESAM tight-junction gene cause a neurodevelopmental disorder associated with fetal intracranial hemorrhage.

The blood-brain barrier (BBB) is an essential gatekeeper for the central nervous system and incidence of neurodevelopmental disorders (NDDs) is higher in infants with a history of intracerebral hemorrhage (ICH). We discovered a rare disease trait in thirteen individuals, including four fetuses, from eight unrelated families associated with homozygous loss-of-function variant alleles of ESAM which encodes an endothelial cell adhesion molecule. The c.115del (p.Arg39Glyfs∗33) variant, identified in six individuals from four independent families of Southeastern Anatolia, severely impaired the in vitro tubulogenic process of endothelial colony-forming cells, recapitulating previous evidence in null mice, and caused lack of ESAM expression in the capillary endothelial cells of damaged brain. Affected individuals with bi-allelic ESAM variants showed profound global developmental delay/unspecified intellectual disability, epilepsy, absent or severely delayed speech, varying degrees of spasticity, ventriculomegaly, and ICH/cerebral calcifications, the latter being also observed in the fetuses. Phenotypic traits observed in individuals with bi-allelic ESAM variants overlap very closely with other known conditions characterized by endothelial dysfunction due to mutation of genes encoding tight junction molecules. Our findings emphasize the role of brain endothelial dysfunction in NDDs and contribute to the expansion of an emerging group of diseases that we propose to rename as "tightjunctionopathies."

Novel molecular, structural and clinical findings in an Italian cohort of congenital cataract.

The current genetic diagnostic workup of congenital cataract (CC) is mainly based on NGS panels, whereas exome sequencing (ES) has occasionally been employed. In this multicentre study, we investigated by ES the detection yield, mutational spectrum and genotype-phenotype correlations in a CC cohort recruited between 2020 and mid-2022. The cohort consisted of 67 affected individuals from 51 unrelated families and included both non-syndromic (75%) and syndromic (25%) phenotypes, with extra-CC ocular/visual features present in both groups (48% and 76%, respectively). The functional effect of variants was predicted by 3D modelling and hydropathy properties changes. Variant clustering was used for the in-depth assessment of genotype-phenotype correlations. A diagnostic (pathogenic or likely pathogenic) variant was identified in 19 out of 51 probands/families (~37%). In a further 14 probands/families a candidate variant was identified: in 12 families a VUS was detected, of which 9 were considered plausibly pathogenic (i.e., 4 or 5 points according to ACMG criteria), while in 2 probands ES identified a single variant in an autosomal recessive gene associated with CC. Eighteen probands/families, manifesting primarily non-syndromic CC (15/18, 83%), remained unsolved. The identified variants (8 P, 12 LP, 10 VUS-PP, and 5 VUS), half of which were unreported in the literature, affected five functional categories of genes involved in transcription/splicing, lens formation/homeostasis (i.e., crystallin genes), membrane signalling, cell-cell interaction, and immune response. A phenotype-specific variant clustering was observed in four genes (KIF1A, MAF, PAX6, SPTAN1), whereas variable expressivity and potential phenotypic expansion in two (BCOR, NHS) and five genes (CWC27, KIF1A, IFIH1, PAX6, SPTAN1), respectively. Finally, ES allowed to detect variants in six genes not commonly included in commercial CC panels. These findings broaden the genotype-phenotype correlations in one of the largest CC cohorts tested by ES, providing novel insights into the underlying pathogenetic mechanisms and emphasising the power of ES as first-tier test.

Identification of bi-allelic LFNG variants in three patients and further clinical and molecular refinement of spondylocostal dysostosis 3.

Spondylocostal dysostosis (SCD), a condition characterized by multiple segmentation defects of the vertebrae and rib malformations, is caused by bi-allelic variants in one of the genes involved in the Notch signaling pathway that tunes the "segmentation clock" of somitogenesis: DLL3, HES7, LFNG, MESP2, RIPPLY2, and TBX6. To date, seven individuals with LFNG variants have been reported in the literature. In this study we describe two newborns and one fetus with SCD, who were found by trio-based exome sequencing (trio-ES) to carry homozygous (c.822-5C>T) or compound heterozygous (c.[863dup];[1063G>A]) and (c.[521G>T];[890T>G]) variants in LFNG. Notably, the c.822-5C>T change, affecting the polypyrimidine tract of intron 5, is the first non-coding variant reported in LFNG. This study further refines the clinical and molecular features of spondylocostal dysostosis 3 and adds to the numerous investigations supporting the usefulness of trio-ES approach in prenatal and neonatal settings.

Further evidence supporting the role of GTDC1 in glycine metabolism and neurodevelopmental disorders.

Copy number variants (CNVs) represent the genetic cause of about 15-20% of neurodevelopmental disorders (NDDs). We identified a ~67 kb de novo intragenic deletion on chromosome 2q22.3 in a female individual showing a developmental encephalopathy characterised by epilepsy, severe intellectual disability, speech delay, microcephaly, and thin corpus callosum with facial dysmorphisms. The microdeletion involved exons 5-6 of GTDC1, encoding a putative glycosyltransferase, whose expression is particularly enriched in the nervous system. In a previous study, a balanced de novo translocation encompassing GTDC1 was reported in a male child with global developmental delay and delayed speech and language development. Based on these premises, we explored the transcriptomic profile of our proband to evaluate the functional consequences of the novel GTDC1 de novo intragenic deletion in relation to the observed neurodevelopmental phenotype. RNA-seq on the proband's lymphoblastoid cell line (LCL) showed expression changes of glycine/serine and cytokine/chemokine signalling pathways, which are related to neurodevelopment and epileptogenesis. Subsequent analysis by ELISA (enzyme-linked immunosorbent assay) and HPLC (high-performance liquid chromatography) revealed increased levels of glycine in the proband's LCL and serum compared to matched controls. Given that an increased level of glycine has been observed in the plasma samples of individuals with Rett syndrome, a condition sharing epilepsy, microcephaly, and intellectual disability with our proband, we proposed that the GTDC1 downregulation is implicated in neurodevelopmental impairment by altering glycine metabolism. Furthermore, our findings expanded the phenotypic spectrum of the novel GTDC1-related condition, including microcephaly and epilepsy among relevant clinical features.