A novel frameshift TBX4 variant in a family with ischio-coxo-podo-patellar syndrome and variable severity.

BACKGROUND: Congenital anomalies of the knee are a spectrum of rare disorders with wide clinical and genetic variability, which are mainly due to the complex processes underlying knee development. Despite progresses in understanding pathomechanisms and associated genes, many patients remain undiagnosed. OBJECTIVE: To uncover the genetic bases of a congenital patellar dislocation affecting multiple family members with variable severity. METHODS: We performed ES in the proband and his father, both showing bilateral patellar dislocation, his sister with a milder similar condition, and his unaffected mother. Sanger sequencing was then performed in the proband's brother and paternal aunt, both affected as well. RESULTS: ES and Sanger sequencing identified the presence of the novel heterozygous frameshift mutation c.735delT in the TBX4 gene in all affected family members. TBX4 is associated with autosomal dominant ischio-coxo-podo-patellar syndrome with/without pulmonary arterial hypertension (ICPPS, #147891), reaching a diagnosis in the family. Intrafamilial clinical heterogeneity suggests that other factors might be involved, such as additional variants in TBX4 or in other modifier genes. Interestingly, we identified three additional variants in the TBX4 gene in the proband only, whose phenotype is more severe. Despite being classified as benign, one of these variants is predicted to disrupt a splicing protein binding site, and may therefore affect TBX4 alternative splicing, accounting for the more severe phenotype of the proband. CONCLUSION: We expand and further delineate the genotypic and phenotypic spectrum of ICPPS. Further studies are necessary to shed light on the potential effect of this variant and on the variable phenotypic expressivity of TBX4-related phenotypes.

Testing calpain inhibition in tumor endothelial cells: novel targetable biomarkers against glioblastoma malignancy.

Introduction: Glioblastoma IDH-wildtype (GBM) is the most malignant brain tumor in adults, with a poor prognosis of approximately 15 months after diagnosis. Most patients suffer from a recurrence in <1 year, and this renders GBM a life-threatening challenge. Among molecular mechanisms driving GBM aggressiveness, angiogenesis mediated by GBM endothelial cells (GECs) deserves consideration as a therapeutic turning point. In this scenario, calpains, a family of ubiquitously expressed calcium-dependent cysteine proteases, emerged as promising targets to be investigated as a novel therapeutic strategy and prognostic tissue biomarkers. Methods: To explore this hypothesis, GECs were isolated from n=10 GBM biopsies and characterized phenotypically by immunofluorescence. The expression levels of calpains were evaluated by qRT-PCR and Western blot, and their association with patients' prognosis was estimated by Pearson correlation and Kaplan-Meier survival analysis. Calpain targeting efficacy was assessed by a time- and dose-dependent proliferation curve, MTT assay for viability, caspase-3/7 activity, migration and angiogenesis in vitro, and gene and protein expression level modification. Results: Immunofluorescence confirmed the endothelial phenotype of our primary GECs. A significant overexpression was observed for calpain-1/2/3 (CAPN) and calpain-small-subunits-1/2 (CAPNS1), whereas calpastatin gene, the calpain natural inhibitor, was reported to be downregulated. A significant negative correlation was observed between CAPN1/CAPNS1 and patient overall survival. GEC challenging revealed that the inhibition of calpain-1 exerts the strongest proapoptotic efficacy, so GEC mortality reached the 80%, confirmed by the increased activity of caspase-3/7. Functional assays revealed a strong affection of in vitro migration and angiogenesis. Gene and protein expression proved a downregulation of MAPK, VEGF/VEGFRs, and Bcl-2, and an upregulation of caspases and Bax-family mediators. Conclusion: Overall, the differential expression of calpains and their correlation with patient survival suggest a novel promising target pathway, whose blockade showed encouraging results toward precision medicine strategies.

Exploring the Impact of Genetics in a Large Cohort of Moebius Patients by Trio Whole Exome Sequencing.

Moebius syndrome (MBS) is a rare congenital disorder characterized by non-progressive facial palsy and ocular abduction paralysis. Most cases are sporadic, but also rare familial cases with autosomal dominant transmission and incomplete penetrance/variable expressivity have been described. The genetic etiology of MBS is still unclear: de novo pathogenic variants in REV3L and PLXND1 are reported in only a minority of cases, suggesting the involvement of additional causative genes. With the aim to uncover the molecular causative defect and identify a potential genetic basis of this condition, we performed trio-WES on a cohort of 37 MBS and MBS-like patients. No de novo variants emerged in REV3L and PLXND1. We then proceeded with a cohort analysis to identify possible common causative genes among all patients and a trio-based analysis using an in silico panel of candidate genes. However, identified variants emerging from both approaches were considered unlikely to be causative of MBS, mainly due to the lack of clinical overlap. In conclusion, despite this large cohort, WES failed to identify mutations possibly associated with MBS, further supporting the heterogeneity of this syndrome, and suggesting the need for integrated omics approaches to identify the molecular causes underlying MBS development.

Pharmacogenetics in Italy: current landscape and future prospects.

Pharmacogenetics investigates sequence of genes that affect drug response, enabling personalized medication. This approach reduces drug-induced adverse reactions and improves clinical effectiveness, making it a crucial consideration for personalized medical care. Numerous guidelines, drawn by global consortia and scientific organizations, codify genotype-driven administration for over 120 active substances. As the scientific community acknowledges the benefits of genotype-tailored therapy over traditionally agnostic drug administration, the push for its implementation into Italian healthcare system is gaining momentum. This evolution is influenced by several factors, including the improved access to patient genotypes, the sequencing costs decrease, the growing of large-scale genetic studies, the rising popularity of direct-to-consumer pharmacogenetic tests, and the continuous improvement of pharmacogenetic guidelines. Since EMA (European Medicines Agency) and AIFA (Italian Medicines Agency) provide genotype information on drug leaflet without clear and explicit clinical indications for gene testing, the regulation of pharmacogenetic testing is a pressing matter in Italy. In this manuscript, we have reviewed how to overcome the obstacles in implementing pharmacogenetic testing in the clinical practice of the Italian healthcare system. Our particular emphasis has been on germline testing, given the absence of well-defined national directives in contrast to somatic pharmacogenetics.

Non-invasive biomarkers for sperm retrieval in non-obstructive patients: a comprehensive review.

Recent advancements in reproductive medicine have guided novel strategies for addressing male infertility, particularly in cases of non-obstructive azoospermia (NOA). Two prominent invasive interventions, namely testicular sperm extraction (TESE) and microdissection TESE (micro-TESE), have emerged as key techniques to retrieve gametes for assisted reproduction technologies (ART). Both heterogeneity and complexity of NOA pose a multifaceted challenge to clinicians, as the invasiveness of these procedures and their unpredictable success underscore the need for more precise guidance. Seminal plasma can be aptly regarded as a liquid biopsy of the male reproductive tract, encompassing secretions from the testes, epididymides, seminal vesicles, bulbourethral glands, and prostate. This fluid harbors a variety of cell-free nucleic acids, microvesicles, proteins, and metabolites intricately linked to gonadal activity. However, despite numerous investigations exploring potential biomarkers from seminal fluid, their widespread inclusion into the clinical practice remains limited. This could be partially due to the complex interplay of diverse clinical and genetic factors inherent to NOA that likely contributes to the absence of definitive biomarkers for residual spermatogenesis. It is conceivable that the integration of clinical data with biomarkers could increase the potential in predicting surgical procedure outcomes and their choice in NOA cases. This comprehensive review addresses the challenge of sperm retrieval in NOA through non-invasive biomarkers. Moreover, we delve into promising perspectives, elucidating innovative approaches grounded in multi-omics methodologies, including genomics, transcriptomics and proteomics. These cutting-edge techniques, combined with the clinical and genetics features of patients, could improve the use of biomarkers in personalized medical approaches, patient counseling, and the decision-making continuum. Finally, Artificial intelligence (AI) holds significant potential in the realm of combining biomarkers and clinical data, also in the context of identifying non-invasive biomarkers for sperm retrieval.

Dysfunction in IGF2R Pathway and Associated Perturbations in Autophagy and WNT Processes in Beckwith-Wiedemann Syndrome Cell Lines.

Beckwith-Wiedemann Syndrome (BWS) is an imprinting disorder characterized by overgrowth, stemming from various genetic and epigenetic changes. This study delves into the role of IGF2 upregulation in BWS, focusing on insulin-like growth factor pathways, which are poorly known in this syndrome. We examined the IGF2R, the primary receptor of IGF2, WNT, and autophagy/lysosomal pathways in BWS patient-derived lymphoblastoid cell lines, showing different genetic and epigenetic defects. The findings reveal a decreased expression and mislocalization of IGF2R protein, suggesting receptor dysfunction. Additionally, our results point to a dysregulation in the AKT/GSK-3/mTOR pathway, along with imbalances in autophagy and the WNT pathway. In conclusion, BWS cells, regardless of the genetic/epigenetic profiles, are characterized by alteration of the IGF2R pathway that is associated with the perturbation of the autophagy and lysosome processes. These alterations seem to be a key point of the molecular pathogenesis of BWS and potentially contribute to BWS's characteristic overgrowth and cancer susceptibility. Our study also uncovers alterations in the WNT pathway across all BWS cell lines, consistent with its role in growth regulation and cancer development.

Proceedings of the 14th European immunogenicity platform open symposium on immunogenicity of biopharmaceuticals.

Biologics have revolutionized disease management in many therapeutic areas by addressing unmet medical needs and overcoming resistance to standard-of-care treatment in numerous patients. However, the development of unwanted immune responses directed against these drugs, humoral and/or cellular, can hinder their efficacy and have safety consequences with various degrees of severity. Health authorities ask that a thorough immunogenicity risk assessment be conducted during drug development to incorporate an appropriate monitoring and mitigation plan in clinical studies. With the rapid diversification and complexification of biologics, which today include modalities such as multi-domain antibodies, cell-based products, AAV delivery vectors, and nucleic acids, developers are faced with the challenge of establishing a risk assessment strategy sometimes in the absence of specific regulatory guidelines. The European Immunogenicity Platform (EIP) Open Symposium on Immunogenicity of Biopharmaceuticals and its one-day training course gives experts and newcomers across academia, industry, and regulatory agencies an opportunity to share experience and knowledge to overcome these challenges. Here, we report the discussions that took place at the EIP's 14th Symposium, held in April 2023. The topics covered included immunogenicity monitoring and clinical relevance, non-clinical immunogenicity risk assessment, regulatory aspects of immunogenicity assessment and reporting, and the challenges associated with new modalities, which were discussed in a dedicated session.

Distinct Functional Humoral Immune Responses Are Induced after Live Attenuated and Inactivated Seasonal Influenza Vaccination.

Influenza viruses infect 5-30% of the world's population annually, resulting in millions of incidents of hospitalization and thousands of mortalities worldwide every year. Although annual vaccination has significantly reduced hospitalization rates in vulnerable populations, the current vaccines are estimated to offer a wide range of protection from 10 to 60% annually. Such incomplete immunity may be related to both poor antigenic coverage of circulating strains, as well as to the insufficient induction of protective immunity. Beyond the role of hemagglutinin (HA) and neuraminidase (NA), vaccine-induced Abs have the capacity to induce a broader array of Ab effector functions, including Ab-dependent cellular cytotoxicity, that has been implicated in universal immunity against influenza viruses. However, whether different vaccine platforms can induce functional humoral immunity in a distinct manner remains incompletely defined. In this study, we compared vaccine-induced humoral immune responses induced by two seasonal influenza vaccines in Homo sapiens, the i.m. inactivated vaccine (IIV/Fluzone) and the live attenuated mucosal vaccine (LAIV/FluMist). Whereas the inactivated influenza vaccine induced superior Ab titers and FcγR binding capacity to diverse HA and NA Ags, the live attenuated influenza mucosal vaccine induced a more robust functional humoral immune response against both the HA and NA domains. Multivariate Ab analysis further highlighted the significantly different overall functional humoral immune profiles induced by the two vaccines, marked by differences in IgG titers, FcR binding, and both NK cell-recruiting and opsonophagocytic Ab functions. These results highlight the striking differences in Ab Fc-effector profiles induced systemically by two distinct influenza vaccine platforms.

Case report: Chorea and cognitive decline in a young woman: instrumental and genetic assessment of a case originally diagnosed as multiple sclerosis.

We describe the case of a young woman affected by debilitating chorea and rapidly progressive cognitive decline. While her original diagnosis was multiple sclerosis, we performed a full instrumental and genetic assessement, though which we identified multiple genetic variants, including a novel variant of the APP gene. We propose some possible mechanisms by which such variants may contribute to neuroinflammation and ultimately lead to this devastating clinical course.

Pitfalls of whole exome sequencing in undefined clinical conditions with a suspected genetic etiology.

BACKGROUND: Whole-Exome Sequencing (WES) is a valuable tool for the molecular diagnosis of patients with a suspected genetic condition. In complex and heterogeneous diseases, the interpretation of WES variants is more challenging given the absence of diagnostic handles and other reported cases with overlapping clinical presentations. OBJECTIVE: To describe candidate variants emerging from trio-WES and possibly associated with the clinical phenotype in clinically heterogeneous conditions. METHODS: We performed WES in ten patients from eight families, selected because of the lack of a clear clinical diagnosis or suspicion, the presence of multiple clinical signs, and the negative results of traditional genetic tests. RESULTS: Although we identified ten candidate variants, reaching the diagnosis of these cases is challenging, given the complexity and the rarity of these syndromes and because affected genes are already associated with known genetic diseases only partially recapitulating patients' phenotypes. However, the identification of these variants could shed light into the definition of new genotype-phenotype correlations. Here, we describe the clinical and molecular data of these cases with the aim of favoring the match with other similar cases and, hopefully, confirm our diagnostic hypotheses. CONCLUSION: This study emphasizes the major limitations associated with WES data interpretation, but also highlights its clinical utility in unraveling novel genotype-phenotype correlations in complex and heterogeneous undefined clinical conditions with a suspected genetic etiology.

Mental health and the effects on methylation of stress-related genes in front-line versus other health care professionals during the second wave of COVID-19 pandemic: an Italian pilot study.

Healthcare workers experienced high degree of stress during COVID-19. Purpose of the present article is to compare mental health (depressive and Post-Traumatic-Stress-Disorders-PTSD-symptoms) and epigenetics aspects (degree of methylation of stress-related genes) in front-line healthcare professionals versus healthcare working in non-COVID-19 wards. Sixty-eight healthcare workers were included in the study: 39 were working in COVID-19 wards (cases) and 29 in non-COVID wards (controls). From all participants, demographic and clinical information were collected by an ad-hoc questionnaire. Depressive and PTSD symptoms were evaluated by the Patient Health Questionnaire-9 (PHQ-9) and the Impact of Event Scale-Revised (IES-R), respectively. Methylation analyses of 9 promoter/regulatory regions of genes known to be implicated in depression/PTSD (ADCYAP1, BDNF, CRHR1, DRD2, IGF2, LSD1/KDM1A, NR3C1, OXTR, SLC6A4) were performed on DNA from blood samples by the MassARRAY EpiTYPER platform, with MassCleave settings. Controls showed more frequent lifetime history of anxiety/depression with respect to cases (χ2 = 5.72, p = 0.03). On the contrary, cases versus controls presented higher PHQ-9 (t = 2.13, p = 0.04), PHQ-9 sleep item (t = 2.26, p = 0.03), IES-R total (t = 2.17, p = 0.03), IES-R intrusion (t = 2.46, p = 0.02), IES-R avoidance (t = 1.99, p = 0.05) mean total scores. Methylation levels at CRHR1, DRD2 and LSD1 genes was significantly higher in cases with respect to controls (p < 0.01, p = 0.03 and p = 0.03, respectively). Frontline health professionals experienced more negative effects on mental health during COVID-19 pandemic than non-frontline healthcare workers. Methylation levels were increased in genes regulating HPA axis (CRHR1) and dopamine neurotransmission (DRD2 and LSD1), thus supporting the involvement of these biological processes in depression/PTSD and indicating that methylation of these genes can be modulated by stress conditions, such as working as healthcare front-line during COVID-19 pandemic.

The Genetic and Epigenetic Footprint in Idiopathic Pulmonary Fibrosis and Familial Pulmonary Fibrosis: A State-of-the-Art Review.

Idiopathic pulmonary fibrosis (IPF) is a rare disease of the lung with a largely unknown etiology and a poor prognosis. Intriguingly, forms of familial pulmonary fibrosis (FPF) have long been known and linked to specific genetic mutations. There is little evidence of the possible role of genetics in the etiology of sporadic IPF. We carried out a non-systematic, narrative literature review aimed at describing the main known genetic and epigenetic mechanisms that are involved in the pathogenesis and prognosis of IPF and FPF. In this review, we highlighted the mutations in classical genes associated with FPF, including those encoding for telomerases (TERT, TERC, PARN, RTEL1), which are also found in about 10-20% of cases of sporadic IPF. In addition to the Mendelian forms, mutations in the genes encoding for the surfactant proteins (SFTPC, SFTPA1, SFTPA2, ABCA3) and polymorphisms of genes for the mucin MUC5B and the Toll-interacting protein TOLLIP are other pathways favoring the fibrogenesis that have been thoroughly explored. Moreover, great attention has been paid to the main epigenetic alterations (DNA methylation, histone modification and non-coding RNA gene silencing) that are emerging to play a role in fibrogenesis. Finally, a gaze on the shared mechanisms between cancer and fibrogenesis, and future perspectives on the genetics of pulmonary fibrosis have been analyzed.

A Regulatory Risk-Based Approach to ATMP/CGT Development: Integrating Scientific Challenges With Current Regulatory Expectations.

Cell and Gene Therapy Products (CGT), regulated as Advanced Therapy Medicinal Products (ATMP) in the European Union (EU), represent a novel and varied group of biotherapeutics developed to treat specific conditions for which there are limited or no effective treatments. The novelty and complexity of this product modality demands a regulatory risk-based approach to define a sound development plan, particularly, as most developers aim to target more than one market area simultaneously for clinical development and registration. This regulatory strategy should be built on solid scientific data that also addresses general regulatory recommendations to enable a benefit:risk analysis that is aligned with the particularities of each CGT product. This risk-based approach is especially helpful when no detailed product-specific regulatory guidelines are available. The goal of this article is to orient developers on how to build a combined EU/US regulatory strategy through the assessment of commonly understood quality (CMC), non-clinical, and clinical regulatory risks faced by ATMP/CGT.

Correction: Rondinone et al. Extensive Placental Methylation Profiling in Normal Pregnancies. Int. J. Mol. Sci. 2021, 22, 2136.

In the original publication [...].

Preferential X Chromosome Inactivation as a Mechanism to Explain Female Preponderance in Myasthenia Gravis.

Myasthenia gravis (MG) is a neuromuscular autoimmune disease characterized by prevalence in young women (3:1). Several mechanisms proposed as explanations for gender bias, including skewed X chromosome inactivation (XCI) and dosage or sex hormones, are often involved in the development of autoimmunity. The skewed XCI pattern can lead to an unbalanced expression of some X-linked genes, as observed in several autoimmune disorders characterized by female predominance. No data are yet available regarding XCI and MG. We hypothesize that the preferential XCI pattern may contribute to the female bias observed in the onset of MG, especially among younger women. XCI analysis was performed on blood samples of 284 women between the ages of 20 and 82. XCI was tested using the Human Androgen Receptor Assay (HUMARA). XCI patterns were classified as random (XCI < 75%) and preferential (XCI ≥ 75%). In 121 informative patients, the frequency of skewed XCI patterns was 47%, significantly higher than in healthy controls (17%; p ≤ 0.00001). Interestingly, the phenomenon was observed mainly in younger patients (<45 years; p ≤ 0.00001). Furthermore, considering the XCI pattern and the other clinical characteristics of patients, no significant differences were found. In conclusion, we observed preferential XCI in MG female patients, suggesting its potential role in the aetiology of MG, as observed in other autoimmune diseases in women.

Functional Compartmentalization of Antibodies in the Central Nervous System During Chronic HIV Infection.

The central nervous system (CNS) has emerged as a critical HIV reservoir. Thus, interventions aimed at controlling and eliminating HIV must include CNS-targeted strategies. Given the inaccessibility of the brain, efforts have focused on cerebrospinal fluid (CSF), aimed at defining biomarkers of HIV-disease in the CNS, including HIV-specific antibodies. However, how antibodies traffic between the blood and CNS, and whether specific antibody profiles track with HIV-associated neurocognitive disorders (HAND) remains unclear. Here, we comprehensively profiled HIV-specific antibodies across plasma and CSF from 20 antiretroviral therapy (ART) naive or treated persons with HIV. CSF was populated by IgG1 and IgG3 antibodies, with reduced Fc-effector profiles. While ART improved plasma antibody functional coordination, CSF profiles were unaffected by ART and were unrelated to HAND severity. These data point to a functional sieving of antibodies across the blood-brain barrier, providing previously unappreciated insights for the development of next-generation therapeutics targeting the CNS reservoir.

Assessment of pregnancy dietary intake and association with maternal and neonatal outcomes.

BACKGROUND: Maternal dietary habits are contributors of maternal and fetal health; however, available data are heterogeneous and not conclusive. METHODS: Nutrient intake during pregnancy was assessed in 503 women with uncomplicated pregnancies, using the validated Food Frequency Questionnaire developed by the European Prospective Investigation into Cancer and Nutrition (EPIC-FFQ). RESULTS: In all, 68% of women had a normal body mass index at the beginning of pregnancy, and 83% of newborns had an appropriate weight for gestational age. Maternal pre-pregnancy body mass index (BMI), gestational weight gain (GWG), and placental weight were independently correlated with birth weight. GWG was not related to the pre-pregnancy BMI. EPIC-FFQ evaluation showed that 30% of women adhered to the European Food Safety Authority (EFSA) ranges for macronutrient intake. In most pregnant women (98.1%), consumption of water was below recommendations. Comparing women with intakes within EFSA ranges for macronutrients with those who did not, no differences were found in BMI, GWG, and neonatal or placental weight. Neither maternal nor neonatal parameters were associated with the maternal dietary profiles. CONCLUSIONS: In our population, maternal pre-pregnancy BMI, GWG, and placental weight are determinants of birth weight percentile, while no association was found with maternal nutrition. Future studies should explore associations through all infancy. IMPACT: Maternal anthropometrics and nutrition status may affect offspring birth weight. In 503 healthy women, maternal pre-pregnancy body mass index (BMI), gestational weight gain (GWG), and placental weight were independently correlated to neonatal birth weight. GWG was not related to the pre-pregnancy BMI. In all, 30% of women respected the EFSA ranges for macronutrients. Neither maternal nor neonatal parameters were associated with maternal dietary profiles considered in this study. Maternal pre-pregnancy BMI, GWG, and placental weight are determinants of neonatal birth weight percentile, while a connection with maternal nutrition profiles was not found.

Novel Pathogenetic Variants in PTHLH and TRPS1 Genes Causing Syndromic Brachydactyly.

Skeletal disorders, including both isolated and syndromic brachydactyly type E, derive from genetic defects affecting the fine tuning of the network of pathways involved in skeletogenesis and growth-plate development. Alterations of different genes of this network may result in overlapping phenotypes, as exemplified by disorders due to the impairment of the parathyroid hormone/parathyroid hormone-related protein pathway, and obtaining a correct diagnosis is sometimes challenging without a genetic confirmation. Five patients with Albright's hereditary osteodystrophy (AHO)-like skeletal malformations without a clear clinical diagnosis were analyzed by whole-exome sequencing (WES) and novel potentially pathogenic variants in parathyroid hormone like hormone (PTHLH) (BDE with short stature [BDE2]) and TRPS1 (tricho-rhino-phalangeal syndrome [TRPS]) were discovered. The pathogenic impact of these variants was confirmed by in vitro functional studies. This study expands the spectrum of genetic defects associated with BDE2 and TRPS and demonstrates the pathogenicity of TRPS1 missense variants located outside both the nuclear localization signal and the GATA ((A/T)GATA(A/G)-binding zinc-containing domain) and Ikaros-like binding domains. Unfortunately, we could not find distinctive phenotypic features that might have led to an earlier clinical diagnosis, further highlighting the high degree of overlap among skeletal syndromes associated with brachydactyly and AHO-like features, and the need for a close interdisciplinary workout in these rare patients. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Cohesin Mutations Induce Chromatin Conformation Perturbation of the H19/IGF2 Imprinted Region and Gene Expression Dysregulation in Cornelia de Lange Syndrome Cell Lines.

Traditionally, Cornelia de Lange Syndrome (CdLS) is considered a cohesinopathy caused by constitutive mutations in cohesin complex genes. Cohesin is a major regulator of chromatin architecture, including the formation of chromatin loops at the imprinted IGF2/H19 domain. We used 3C analysis on lymphoblastoid cells from CdLS patients carrying mutations in NIPBL and SMC1A genes to explore 3D chromatin structure of the IGF2/H19 locus and evaluate the influence of cohesin alterations in chromatin architecture. We also assessed quantitative expression of imprinted loci and WNT pathway genes, together with DMR methylation status of the imprinted genes. A general impairment of chromatin architecture and the emergence of new interactions were found. Moreover, imprinting alterations also involved the expression and methylation levels of imprinted genes, suggesting an association among cohesin genetic defects, chromatin architecture impairment, and imprinting network alteration. The WNT pathway resulted dysregulated: canonical WNT, cell cycle, and WNT signal negative regulation were the most significantly affected subpathways. Among the deregulated pathway nodes, the key node of the frizzled receptors was repressed. Our study provides new evidence that mutations in genes of the cohesin complex have effects on the chromatin architecture and epigenetic stability of genes commonly regulated by high order chromatin structure.

A novel de novo DDX3X missense variant in a female with brachycephaly and intellectual disability: a case report.

BACKGROUND: De novo pathogenic variants in the DDX3X gene are reported to account for 1-3% of unexplained intellectual disability (ID) in females, leading to the rare disease known as DDX3X syndrome (MRXSSB, OMIM #300958). Besides ID, these patients manifest a variable clinical presentation, which includes neurological and behavioral defects, and abnormal brain MRIs. CASE PRESENTATION: We report a 10-year-old girl affected by delayed psychomotor development, delayed myelination, and polymicrogyria (PMG). We identified a novel de novo missense mutation in the DDX3X gene (c.625C > G) by whole exome sequencing (WES). The DDX3X gene encodes a DEAD-box ATP-dependent RNA-helicase broadly implicated in gene expression through regulation of mRNA metabolism. The identified mutation is located just upstream the helicase domain and is suggested to impair the protein activity, thus resulting in the altered translation of DDX3X-dependent mRNAs. The proband, presenting with the typical PMG phenotype related to the syndrome, does not show other clinical signs frequently reported in presence of missense DDX3X mutations that are associated with a most severe clinical presentation. In addition, she has brachycephaly, never described in female DDX3X patients, and macroglossia, that has never been associated with the syndrome. CONCLUSIONS: This case expands the knowledge of DDX3X pathogenic variants and the associated DDX3X syndrome phenotypic spectrum.