Long QT syndrome (LQTS) is a rare and potentially life-threatening arrhythmia characterized by delayed repolarization on electrocardiogram (EKG) evaluation. Although LQTS is primarily associated with defects in ion channels, in approximately 20

Epstein-Barr virus (EBV) is causally associated with various malignancies and autoimmune diseases. It establishes a lifelong latent infection in host B-lymphocytes, and can strategically manipulate host cell metabolism and cell-to-cell communication via extracellular particles (EPs). While EBV-induced changes in fatty acid and mevalonate pathways are documented, the role of the sphingolipid network in this process remains poorly understood. In this study, we characterized both the EP and the cellular sphingolipid signatures of three EBV-infected (EBV+) lymphoblastoid cell lines compared to three EBV-negative (EBV-) counterparts. Lipidomic analysis revealed a profound metabolic redirection in EBV+ cells, characterized by reduced ceramide and sphingomyelin levels, and a significant upregulation of glycosphingolipids (GSLs), particularly hexosylceramide and globotrialosylceramide. Furthermore, EBV+ cells released larger, more heterogeneous EPs that are significantly enriched in GSLs and in the viral non-coding RNA EBER1. Our findings suggest that EBV-induced reshaping of sphingolipid metabolism correlates with altered physical properties of the secretome, potentially contributing to viral pathogenesis and microenvironmental manipulation.

Cornelia de Lange syndrome (CdLS) is a rare genetic disorder that affects almost any organ, including the central nervous system. It leads to a wide range of neurodevelopmental delays, and there are currently no available clinical treatments. CdLS is caused by pathogenic variants in one of the 7 genes coding for the cohesin complex, a multimeric structure responsible for sister chromatid cohesion, or for cohesin ring-interacting proteins. Additionally, altered regulation of molecular pathways during development, including the canonical WNT pathway, can cause CdLS malformations. In our study, we evaluated the positive effects of using lithium as an activator of the canonical WNT pathway to ameliorate neural CdLS phenotype. We have exploited accurate two-dimensional (2D) and three-dimensional (3D) human central nervous system in vitro models representing disease-related neurobiological phenotypes: induced pluripotent stem cells of human origin (hiPSCs) differentiated into neural precursors, neurons, and brain organoids (BOs). CdLS models demonstrate alterations in proliferation and differentiation capabilities when mimicking HDAC8 haploinsufficiency. Furthermore, RNA-seq analysis of BOs revealed that both neuronal differentiation and the WNT pathway are downregulated when treated with the HDAC8 inhibitor alone. Following lithium treatment, cells show an enhanced ability to differentiate into the neuronal lineage. Additionally, our working hypothesis is that a specific mechanism may exist that, by connecting lipid metabolism, canonical WNT pathway, and cell death, results in typical CdLS neurodevelopmental deficits.

The cohesin complex performs essential cellular functions including regulation of chromatin organization and DNA repair. Somatic pathogenetic variants in cohesin genes, such as STAG2, have been associated with cancer, but their contribution to brain tumorigenesis is unclear. Here, we report the presence of STAG2 variants in patients with glioblastoma and medulloblastoma and determined the effects of loss of STAG2 in human cells and of the homolog SA1 in Drosophila tissues. Reduction of SA1 expression during fly brain development led to defects in neural stem cell differentiation and promotion of tumorigenesis, both in the presence and absence of oncogenic activity. Treatment with inhibitors of poly ADP-ribose polymerase (PARP), which are used to treat forms of cancer with defects in DNA repair, in combination with STAG2/SA1 depletion resulted in apoptosis in vitro and in vivo. In flies, reduction of PARP activity ameliorated the tumor-associated phenotypes of SA1-deficient tissue. Our in vivo and in vitro data suggest that impairment of PARP activity compensates for reduced cohesin activity, highlighting a vulnerability that could be pharmacologically exploited in brain tumors.

Background: Defining a genetic diagnosis is a challenging and stepwise process, often limited by current knowledge perspectives. Alterations in non-coding RNAs and in their modulating mechanisms are now emerging as one of the main possible explanations underpinning many unsolved cases of neurodevelopmental disorders. The latest description concerns the PUS7 gene, which encodes a pseudouridine synthase (PUS) enzyme, which has been causatively associated with an extremely rare recessive disease characterized by postnatal microcephaly, neurodevelopmental impairment, and minor dysmorphisms. Here, we report a novel case of this poorly known disorder, which was diagnosed in a young girl referred to our center following a misdiagnosis of Kleefstra Syndrome type 2. Methods: Whole-genome sequencing detected two composite heterozygous deletions in the PUS7 gene, which led us to dismiss the former diagnosis. Guided by the gestaltic evaluation of our case, we reviewed the current knowledge about the PUS7-related disorder and, driven by the biological role of this enzyme, investigated its similarities with a main spliceosomopathy, ReNU syndrome. Results: Data obtained from this exploratory analysis, together with the recent evidence from the literature, support the affinity between these conditions, suggesting that disorders of pseudouridylation might possibly be included among spliceosomopathies. Conclusions: Aside from expanding the current knowledge about the PUS7-related disorder and exploring its nosological classification, one of the main aims of our report is to raise awareness of the impact of genetic labelling and misdiagnoses. The use of whole-genome sequencing technologies requires the effective collaboration between laboratories and physicians to reach the correct diagnosis.

The cohesin complex plays crucial roles in DNA repair, chromatid separation, and gene transcription regulation. Pathogenic variants in cohesins or dysfunctional transcriptional regulators lead to cohesinopathies, a broader group of disorders including Cornelia de Lange Spectrum (CdLSp), for which the prevalence of cancer cases remains unclear. Here, we aimed to assess the prevalence of oncological events in CdLSp and elucidate the role of cohesin variants in cancer predisposition. We developed a custom next-generation sequencing (NGS) panel targeting predisposition and pathogenic genes, which we applied on N = 120 samples of pediatric patients with acute lymphoblastic leukemia (ALL), identifying 11 out of 229 total-10 germline and 1 somatic-variants in cohesin genes. Data of N = 205 brain tumors were extracted by bioinformatic analysis of data from open-source databases carrying 19 somatic variants. In a cohort of 54 CdLSp patients, the largest cohort from a single center, with a median age of 13 years, the hypothesis of an increased prevalence of cancer in CdLSp was not confirmed. Our findings highlight a significant involvement of germline NIPBL variants in CdLSp, whereas RAD21 and STAG1/2 are predominantly found as somatic variants in neoplasms. However, a distinct genetic or molecular pattern distinguishing variants leading to CdLSp from tumors was not identified. Hence, we advocate for further investigation into the relationship between cohesin variants and cancer predisposition in a larger cohort of patients, with a longer observation time and including different types of malignancies, with more focus on epigenetic approaches.

Cellular senescence is a biological process in which the cell cycle is arrested in response to DNA damage caused by different endogenous and exogenous stimuli. In senescent cells, activation of intracellular cascade induces epigenetic, morphological and metabolic changes. Among them, senescent status is characterized by an alteration of the epigenome and the establishment of a peculiar senescence-associated secretory phenotype (SASP), which contributes to the extracellular matrix remodeling and senescence spreading. Growing interest is directed towards senescence relevance both in physiological processes and in pathological ones, including rare progeroid syndromes. However, little is known about senescence contribution to the onset and development of rare diseases in which aging traits are not manifested. Here, we review the current knowledge about senescence involvement in four rare mendelian disorders of the epigenetic machinery (i.e. chromatinopathies) and four rare lung diseases, that can be considered a paradigm for understanding how epigenome alteration and aberrant microenvironment modification in senescence process might drive disease onset and progression. First, we report the main characteristics of chromatinopathies and the relation between the chromatin-related epigenetic defects and the senescence features in Sotos syndrome, Cornelia de Lange syndrome, Rett syndrome, and Kleefstra syndromes. Thereafter, we describe the pathological alteration and senescence involvement in cystic fibrosis, idiopathic pulmonary fibrosis, pulmonary arterial hypertension and lymphangioleiomyomatosis, considering them as models of rare lung diseases in which accumulation of senescent cells and their proinflammatory SASP have a central role. Exploring the role of senescence in different and less common diseases might promote the understanding of the senescent process as a novel player in rare disorders, for a more comprehensive vision of their complexity and the suggestion of novel possible therapeutical targets.

Cellular senescence represents a permanent state of cell cycle arrest, also observed in neurodegenerative disorders. As p300 has been identified as an epigenetic driver of replicative senescence, we aimed to investigate whether in vitro p300 inhibition could rescue the stress-induced premature senescence (SIPS) phenotype. We exploited 2D and 3D (brain organoids) in vitro models of SIPS using two different stressor agents. In addition, we combined the treatment with a p300 inhibitor and validated p300 role in SIPS by analyzing different senescence markers and the transcriptome in our models. Interestingly, p300 inhibition can counteract the DNA damage and SIPS phenotype, detecting a dysregulation of gene expression and protein translation associated with the senescence program. These findings highlight both the molecular mechanisms underlying senescence and p300 as a possible pharmacological target. Thus, targeting p300 and, by extension, senescent cells could represent a promising therapeutic strategy for age-related diseases such as neurodegenerative disorders.

The cohesin complex performs essential cellular functions including regulation of chromosome cohesion, chromatin organization and DNA repair. Somatic pathogenetic variants in cohesin genes, such as STAG2 , have been associated with cancer, but their contribution to brain tumorigenesis is unclear. Here, we report the presence of STAG2 variants in glioblastoma and medulloblastoma patients and determine that loss of STAG2 in human cells leads to DNA damage and apoptosis. Treatment with inhibitors of the Poly ADP-ribose polymerase (PARP), which are used to treat forms of cancer with defects in DNA repair, increased the amount of apoptosis, confirming that synthetic lethality between reduced cohesin and PARP activity could be observed in vitro . Similar results were obtained in vivo by reducing expression of SA1, the Drosophila melanogaster homolog of STAG1/2. Cohesin gene silencing during fly brain development leads to defects in neural stem cells differentiation and tumorigenesis both in the presence of oncogenic activity and per se . Our in vivo and in vitro data suggests that impairment of PARP activity might induce synthetic lethality in cohesin-dependent tumors, highlighting a vulnerability that can be pharmacologically exploited. ### Competing Interest Statement The authors have declared no competing interest. AIRC, , 20661 WCR, , 18-399

Chromatinopathies (CPs) are an expanding group of rare genetic disorders affecting epigenetic machinery. Besides an intricate genotypic spectrum, these conditions share overlapping phenotypes characterized by neurocognitive impairment, growth defects and distinctive, but often convergent, facial features. Although individually rare, the landscape of CPs is increasingly growing and represents an emerging and possibly underestimated cause of disability. Due to their complexity and rarity, accurate diagnosis and management pose significant difficulties. To address these challenges and gain a deeper overview of these diseases' spectrum, we retrospectively collected clinical characteristics of 239 patients diagnosed with CPs and critically analyzed their diagnostic journey, growth charts, neurological and gestaltic features. Starting from the largest collection of CPs to date, our data point to wide sequencing analyses as the best shortcut to diagnosis. We have also demonstrated the importance of growth defects in this group of disorders that require dedicated growth tables, and we have delved into the great variability of neurological and clinical burden in these conditions. This retrospective study provides a significant advance in our understanding of these rare diseases and will help to improve diagnostic, therapeutic, and clinical approaches to CPs and to develop personalized multidisciplinary care plans for affected patients.

Histone deacetylases (HDACs) are enzymes pivotal for histone modification (i.e. acetylation marks removal), chromatin accessibility and gene expression regulation. Class I HDACs (including HDAC1, 2, 3, 8) are ubiquitously expressed and they often participate in multi-molecular protein complexes. To date, three neurodevelopmental disorders caused by mutations in genes encoding for HDACs (HDAC4, HDAC6 and HDAC8) and thus belonging to the group of chromatinopathies, have been described. We performed whole exome sequencing (WES) for a patient (#249) clinically diagnosed with the chromatinopathy Rubinstein-Taybi syndrome (RSTS) but negative for mutations in RSTS genes, identifying a de novo frameshift variant in HDAC2 gene. We then investigated its molecular effects in lymphoblastoid cell lines (LCLs) derived from the patient compared to LCLs from healthy donors (HD). As the variant was predicted to be likely pathogenetic and to affect the sequence of nuclear localization signal, we performed immunocytochemistry and lysates fractionation, observing a nuclear mis-localization of HDAC2 compared to HD LCLs. In addition, HDAC2 total protein abundance resulted altered in patient, and we found that newly identified variant in HDAC2 affects also acetylation levels, with significant difference in acetylation pattern among patient #249, HD and RSTS cells and in expression of a known molecular target. Remarkably, RNA-seq performed on #249, HD and RSTS cells shows differentially expressed genes (DEGs) common to #249 and RSTS. Interestingly, our reported patient was clinically diagnosed with RSTS, a chromatinopathy which known causative genes encode for enzymes antagonizing HDACs. These results support the role of HDAC2 as causative gene for chromatinopathies, strengthening the genotype-phenotype correlations in this relevant group of disorders.

ABSTRACT Limited therapeutic advancements in Schizophrenia (SCZ) depend on the heterogeneous nature of the disorder, impacting drug development and clinical trials that assume uniform therapy response, neglecting individual genetic and epigenomic variability. Disease modeling using human induced pluripotent stem cells (hiPSCs) is ideally suited for precision medicine, enabling individualized treatment approaches. Here, we describe the generation of patient-specific lines from somatic cells of SCZ individuals with well-defined diverse clinical trajectories using a Sendai virus-based reprogramming system. Karyotypically and CGH-array validated, the generated hiPSCs expressed diagnostic markers and demonstrated functional pluripotency. Converting these hiPSCs into neural progenitor cells enables the identification of aberrant cellular phenotypes associated with specific pathologically relevant neural phenotypes. This collection of hiPSC lines serves as a platform for developing therapeutic compounds targeting neural populations, potentially addressing early-stage disease alterations.

SMC1A epilepsy syndrome or developmental and epileptic encephalopathy-85 with or without midline brain defects (DEE85, OMIM #301044) is an X-linked neurologic disorder associated with mutations of the SMC1A gene, which is also responsible for about 5% of patients affected by Cornelia de Lange syndrome spectrum (CdLS). Only described in female patients, SMC1A epilepsy syndrome is characterized by the onset of severe refractory epileptic seizures in the first year of life, global developmental delay, a variable degree of intellectual disability, and dysmorphic facial features not typical of CdLS. This was a descriptive observational study for the largest international cohort with this specific disorder. The main goal of this study was to improve the knowledge of the natural history of this phenotype with particular attention to the psychomotor development and the epilepsy data. The analyzed cohort shows normal prenatal growth with the subsequent development of postnatal microcephaly. The incidence of neonatal problems (seizures and respiratory compromise) is considerable (51.4%). There is a significant prevalence of central nervous system (20%) and cardiovascular malformations (20%). Motor skills are generally delayed. The presence of drug-resistant epilepsy is confirmed; the therapeutic role of a ketogenic diet is still uncertain. The significant regression of previously acquired skills following the onset of seizures has been observed. Facial dysmorphisms are variable and no patient shows a classic CdLS phenotype. To sum up, SMC1A variants caused drug-resistant epilepsy in these patients, more than two-thirds of whom were shown to progress to developmental and epileptic encephalopathy. The SMC1A gene variants are all different from each other (apart from a couple of monozygotic twins), demonstrating the absence of a mutational hotspot in the SMC1A gene. Owing to the absence of phenotypic specificity, whole-exome sequencing is currently the diagnostic gold standard.

BACKGROUND:the protein phosphatase 3 catalytic subunit alpha (PPP3CA) gene encodes for the alpha isoform of the calcineurin catalytic subunit, which controls the phosphorylation status of many targets. Currently, 23 pathogenic variants of PPP3CA are known, with clinical manifestations varying by mutation type and domain. RESULTS:through whole exome sequencing, we found two de novo variants in PPP3CA: a frameshift variant predicted leading to a truncated protein in Pt.1 and a splicing variant in Pt.2 associated with mild phenotype. PPP3CA is ubiquitously expressed with tissue-specificity of; namely, splicing isoform 1 prevailing over isoform 2 in the central nervous system. By analyzing isoform distribution in patient-derived cell lines, we highlight a skewed expression of both isoforms in Pt.1, whereas only isoform 2 shows a moderate reduction in Pt.2. In contrast, we did not observe significant abundance changes at the protein level. Cell lines derived from Pt.1 showed a reduced proliferation, associated with an increase in cell death and the upregulation of the unfolded protein response (UPR) pathway. CONCLUSION:data suggest that an aberrant PPP3CA protein in Pt.1 could lead to UPR activation resulting in increased cell death. In Pt.2 an imbalance between the two main isoforms possibly explains the peculiar pathological manifestations, such as a moderate developmental delay.