A specific serum lipid signature characterizes patients with glycogen storage disease type Ia.

Glycogen storage disease type Ia (GSDIa) is a rare, inherited glucose-6-phosphatase-α (G6Pase-α) deficiency-induced carbohydrate metabolism disorder. Although hyperlipidemia is a hallmark of GSDI, the extent of lipid metabolism disruption remains incompletely understood. Lipidomic analysis was performed to characterize the serum lipidome in patients with GSDIa, by including age- and sex-matched healthy controls and age-matched hypercholesterolemic controls. Metabolic control and dietary information biochemical markers were obtained from patients with GSDIa. Patients with GSDIa showed higher total serum lysophosphatidylcholine (Fold Change, (FC) 2.2, P < 0.0001), acyl-acyl-phosphatidylcholine (FC 2.1, P < 0.0001), and ceramide (FC 2.4, P < 0.0001) levels and bile acid (FC 0.7, P < 0.001), acylcarnitines (FC 0.7, P < 0.001), and cholesterol esters (FC 1.0, P < 0.001) than those of healthy controls, and higher di- (FC 1.1, P < 0.0001; FC 0.9, P < 0.01) and triacylglycerol (FC 6.3, P < 0.0001; FC 3.9, P < 0.01) levels than those of healthy controls and hypercholesterolemic subjects. Both total cholesterol and triglyceride values correlated with Cer (d16:1/22:0), Cer (d18:1/20:0), Cer (d18:1/20:0(OH)), Cer (d18:1/22:0), Cer (d18:1/23:0), Cer (d18:1/24:1), Cer (d18:2/22:0), Cer (d18:2/24:1). Total cholesterol also correlated with Cer (d18:1/24:0), Cer (d18:2/20:0), HexCer (d16:1/22:0), HexCer (d18:1/18:0), and Hex2Cer (d18:1/20:0). Triglyceridelevels correlated with Cer (d18:0/24:1). Alanine aminotransferase values correlated with Cer (d18:0/22:0), insulin with Cer (d18:1/22:1) and Cer (d18:1/24:1), and HDL with hexosylceramide (HexCer) (d18:2/23:0). These results expand on the currently known involvement of lipid metabolism in GSDIa. Circulating Cer may allow for refined dietary assessment compared with traditional biomarkers. Because specific lipid species are relatively easy to assess, they represent potential novel biomarkers of GSDIa.

Heterozygosity for loss-of-function variants in LZTR1 is associated with isolated multiple café-au-lait macules.

PURPOSE: Pathogenic LZTR1 variants cause schwannomatosis and dominant/recessive Noonan syndrome (NS). We aim to establish an association between heterozygous loss-of-function LZTR1 alleles and isolated multiple café-au-lait macules (CaLMs). METHODS: A total of 849 unrelated participants with multiple CaLMs, lacking pathogenic/likely pathogenic NF1 and SPRED1 variants, underwent RASopathy gene panel sequencing. Data on 125 individuals with heterozygous LZTR1 variants were collected for characterizing their clinical features and the associated molecular spectrum. In vitro functional assessment was performed on a representative panel of missense variants and small in-frame deletions. RESULTS: Analysis revealed heterozygous LZTR1 variants in 6.0% (51/849) of participants, exceeding the general population prevalence. LZTR1-related CaLMs varied in number, displayed sharp or irregular borders, and were generally isolated but occasionally associated with features recurring in RASopathies. In 2 families, CaLMs and schwannomas co-occurred. The molecular spectrum mainly consisted of truncating variants, indicating loss-of-function. These variants substantially overlapped with those occurring in schwannomatosis and recessive NS. Functional characterization showed accelerated protein degradation or mislocalization, and failure to downregulate mitogen-activated protein kinase signaling. CONCLUSION: Our findings expand the phenotypic variability associated with LZTR1 variants, which, in addition to conferring susceptibility to schwannomatosis and causing dominant and recessive NS, occur in individuals with isolated multiple CaLMs.

SMAD4 mutations causing Myhre syndrome are under positive selection in the male germline.

While it is widely thought that de novo mutations (DNMs) occur randomly, we previously showed that some DNMs are enriched because they are positively selected in the testes of aging men. These "selfish" mutations cause disorders with a shared presentation of features, including exclusive paternal origin, significant increase of the father's age, and high apparent germline mutation rate. To date, all known selfish mutations cluster within the components of the RTK-RAS-MAPK signaling pathway, a critical modulator of testicular homeostasis. Here, we demonstrate the selfish nature of the SMAD4 DNMs causing Myhre syndrome (MYHRS). By analyzing 16 informative trios, we show that MYHRS-causing DNMs originated on the paternally derived allele in all cases. We document a statistically significant epidemiological paternal age effect of 6.3 years excess for fathers of MYHRS probands. We developed an ultra-sensitive assay to quantify spontaneous MYHRS-causing SMAD4 variants in sperm and show that pathogenic variants at codon 500 are found at elevated level in sperm of most men and exhibit a strong positive correlation with donor's age, indicative of a high apparent germline mutation rate. Finally, we performed in vitro assays to validate the peculiar functional behavior of the clonally selected DNMs and explored the basis of the pathophysiology of the different SMAD4 sperm-enriched variants. Taken together, these data provide compelling evidence that SMAD4, a gene operating outside the canonical RAS-MAPK signaling pathway, is associated with selfish spermatogonial selection and raises the possibility that other genes/pathways are under positive selection in the aging human testis.

Immunological Aspects of Kabuki Syndrome: A Retrospective Multicenter Study of the Italian Primary Immunodeficiency Network (IPINet).

Kabuki Syndrome (KS) is a multisystemic genetic disorder. A portion of patients has immunological manifestations characterized by increased susceptibility to infections and autoimmunity. Aiming to describe the clinical and laboratory immunological aspects of KS, we conducted a retrospective multicenter observational study on patients with KS treated in centers affiliated to the Italian Primary Immunodeficiency Network.Thirty-nine patients were enrolled, with a median age at evaluation of 10 years (range: 3 m-21y). All individuals had organ malformations of variable severity. Congenital heart defect (CHD) was present in 19/39 patients (49%) and required surgical correction in 9/39 (23%), with associated thymectomy in 7/39 (18%). Autoimmune cytopenia occurred in 6/39 patients (15%) and was significantly correlated with thymectomy (p < 0.002), but not CHD. Individuals with cytopenia treated with mycophenolate as long-term immunomodulatory treatment (n = 4) showed complete response. Increased susceptibility to infections was observed in 22/32 patients (69%). IgG, IgA, and IgM were low in 13/29 (45%), 13/30 (43%) and 4/29 (14%) patients, respectively. Immunoglobulin substitution was required in three patients. Lymphocyte subsets were normal in all patients except for reduced naïve T-cells in 3/15 patients (20%) and reduced memory switched B-cells in 3/17 patients (18%). Elevated CD3 + TCRαß + CD4-CD8-T-cells were present in 5/17 individuals (23%) and were correlated with hematological and overall autoimmunity (p < 0.05).In conclusion, immunological manifestations of KS in our cohort include susceptibility to infections, antibody deficiency, and autoimmunity. Autoimmune cytopenia is correlated with thymectomy and elevated CD3 + TCRαß + CD4-CD8-T-cells, and benefits from treatment with mycophenolate.

Endocrine involvement in hepatic glycogen storage diseases: pathophysiology and implications for care.

Hepatic glycogen storage diseases constitute a group of disorders due to defects in the enzymes and transporters involved in glycogen breakdown and synthesis in the liver. Although hypoglycemia and hepatomegaly are the primary manifestations of (most of) hepatic GSDs, involvement of the endocrine system has been reported at multiple levels in individuals with hepatic GSDs. While some endocrine abnormalities (e.g., hypothalamic­pituitary axis dysfunction in GSD I) can be direct consequence of the genetic defect itself, others (e.g., osteopenia in GSD Ib, insulin-resistance in GSD I and GSD III) may be triggered by the (dietary/medical) treatment. Being aware of the endocrine abnormalities occurring in hepatic GSDs is essential (1) to provide optimized medical care to this group of individuals and (2) to drive research aiming at understanding the disease pathophysiology. In this review, a thorough description of the endocrine manifestations in individuals with hepatic GSDs is presented, including pathophysiological and clinical implications.

The role of unidentified bright objects in the neurocognitive profile of neurofibromatosis type 1 children: a volumetric MRI analysis.

PURPOSE: Cognitive impairment is described in 80% of Neurofibromatosis type 1 (NF1) patients. Brain focal areas of T2w increased signal intensity on MRI, the so-called Unidentified Bright Objects (UBOs) have been hypothesized to be related to cognitive dysfunction, although conflicting results are available in literature. Here, we investigated the possible relation between UBOs' volume, cognitive impairment, and language disability in NF1 patients. MATERIAL AND METHODS: In this retrospective study, clinical and MRI data of 21 NF1 patients (M/F = 12/9; mean age 10.1 ± 4.5) were evaluated. Brain intellectual functioning and language abilities were assessed with specific scales, while the analyzed MRI sequences included axial 2D-T2-weighted and FLAIR sequences. These images were used independently for UBOs segmentation with a semiautomatic approach and obtained volumes were normalized for biparietal diameters to take into account for brain volume. Possible differences in terms of normalized UBOs volumes were probed between cognitively affected and preserved patients, as well as between subjects with or without language impairment. RESULTS: Patients cognitively affected were not different in terms of UBOs volume compared to those preserved (p = 0.35 and p = 0.30, for T2-weighted and FLAIR images, respectively). Similarly, no differences were found between patients with and without language impairment (p = 0.47 and p = 0.40, for the two sequences). CONCLUSIONS: The relation between UBOs and cognition in children with NF1 has been already investigated in literature, although leading to conflicting results. Our study expands the current knowledge, showing a lack of correlation between UBOs volume and both cognitive impairment and language disability in NF1 patients.

Mitochondrial reprogramming in peripheral blood mononuclear cells of patients with glycogen storage disease type Ia.

BACKGROUND: Glycogen storage disease type Ia (GSDIa) is an inborn metabolic disorder caused by the deficiency of glucose-6-phospatase-α (G6Pase-α) leading to mitochondrial dysfunction. It remains unclear whether mitochondrial dysfunction is present in patients' peripheral blood mononuclear cells (PBMC) and whether dietary treatment can play a role. The aim of this study was to investigate mitochondrial function in PBMC of GSDIa patients. METHODS: Ten GSDIa patients and 10 age-, sex- and fasting-time matched controls were enrolled. Expression of genes involved in mitochondrial function and activity of key fatty acid oxidation (FAO) and Krebs cycle proteins were assessed in PBMC. Targeted metabolomics and assessment of metabolic control markers were also performed. RESULTS: Adult GSDIa patients showed increased CPT1A, SDHB, TFAM, mTOR expression (p < 0.05) and increased VLCAD, CPT2 and citrate synthase activity in PBMC (p < 0.05). VLCAD activity directly correlated with WC (p < 0.01), BMI (p < 0.05), serum malonycarnitine levels (p < 0.05). CPT2 activity directly correlated with BMI (p < 0.05). CONCLUSION: Mitochondrial reprogramming is detectable in PBMC of GSDIa patients. This feature may develop as an adaptation to the liver enzyme defect and may be triggered by dietary (over)treatment in the frame of G6Pase-α deficiency. PBMC can represent an adequate mean to assess (diet-induced) metabolic disturbances in GSDIa.

FOXI3 pathogenic variants cause one form of craniofacial microsomia.

Craniofacial microsomia (CFM; also known as Goldenhar syndrome), is a craniofacial developmental disorder of variable expressivity and severity with a recognizable set of abnormalities. These birth defects are associated with structures derived from the first and second pharyngeal arches, can occur unilaterally and include ear dysplasia, microtia, preauricular tags and pits, facial asymmetry and other malformations. The inheritance pattern is controversial, and the molecular etiology of this syndrome is largely unknown. A total of 670 patients belonging to unrelated pedigrees with European and Chinese ancestry with CFM, are investigated. We identify 18 likely pathogenic variants in 21 probands (3.1%) in FOXI3. Biochemical experiments on transcriptional activity and subcellular localization of the likely pathogenic FOXI3 variants, and knock-in mouse studies strongly support the involvement of FOXI3 in CFM. Our findings indicate autosomal dominant inheritance with reduced penetrance, and/or autosomal recessive inheritance. The phenotypic expression of the FOXI3 variants is variable. The penetrance of the likely pathogenic variants in the seemingly dominant form is reduced, since a considerable number of such variants in affected individuals were inherited from non-affected parents. Here we provide suggestive evidence that common variation in the FOXI3 allele in trans with the pathogenic variant could modify the phenotypic severity and accounts for the incomplete penetrance.

Clinical profiling of MRD48 and functional characterization of two novel pathogenic RAC1 variants.

RAC1 is a member of the Rac/Rho GTPase subfamily within the RAS superfamily of small GTP-binding proteins, comprising 3 paralogs playing a critical role in actin cytoskeleton remodeling, cell migration, proliferation and differentiation. De novo missense variants in RAC1 are associated with a rare neurodevelopmental disorder (MRD48) characterized by DD/ID and brain abnormalities coupled with a wide range of additional features. Structural and functional studies have documented either a dominant negative or constitutively active behavior for a subset of mutations. Here, we describe two individuals with previously unreported de novo missense RAC1 variants. We functionally demonstrate their pathogenicity proving a gain-of-function (GoF) effect for both. By reviewing the clinical features of these two individuals and the previously published MRD48 subjects, we further delineate the clinical profile of the disorder, confirming its phenotypic variability. Moreover, we compare the main features of MRD48 with the neurodevelopmental disease caused by GoF variants in the paralog RAC3, highlighting similarities and differences. Finally, we review all previously reported variants in RAC proteins and in the closely related CDC42, providing an updated overview of the spectrum and hotspots of pathogenic variants affecting these functionally related GTPases.

Performance Metrics of the Scoring System for the Diagnosis of the Beckwith-Wiedemann Spectrum (BWSp) and Its Correlation with Cancer Development.

Different scoring systems for the clinical diagnosis of the Beckwith-Wiedemann spectrum (BWSp) have been developed over time, the most recent being the international consensus score. Here we try to validate and provide data on the performance metrics of these scoring systems of the 2018 international consensus and the previous ones, relating them to BWSp features, molecular tests, and the probability of cancer development in a cohort of 831 patients. The consensus scoring system had the best performance (sensitivity 0.85 and specificity 0.43). In our cohort, the diagnostic yield of tests on blood-extracted DNA was low in patients with a low consensus score (~20% with a score = 2), and the score did not correlate with cancer development. We observed hepatoblastoma (HB) in 4.3% of patients with UPD(11)pat and Wilms tumor in 1.9% of patients with isolated lateralized overgrowth (ILO). We validated the efficacy of the currently used consensus score for BWSp clinical diagnosis. Based on our observation, a first-tier analysis of tissue-extracted DNA in patients with <4 points may be considered. We discourage the use of the consensus score value as an indicator of the probability of cancer development. Moreover, we suggest considering cancer screening for negative patients with ILO (risk ~2%) and HB screening for patients with UPD(11)pat (risk ~4%).

Genotypes and phenotypes heterogeneity in PIK3CA-related overgrowth spectrum and overlapping conditions: 150 novel patients and systematic review of 1007 patients with PIK3CA pathogenetic variants.

BACKGROUND: Postzygotic activating PIK3CA variants cause several phenotypes within the PIK3CA-related overgrowth spectrum (PROS). Variant strength, mosaicism level, specific tissue involvement and overlapping disorders are responsible for disease heterogeneity. We explored these factors in 150 novel patients and in an expanded cohort of 1007 PIK3CA-mutated patients, analysing our new data with previous literature to give a comprehensive picture. METHODS: We performed ultradeep targeted next-generation sequencing (NGS) on DNA from skin biopsy, buccal swab or blood using a panel including phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin pathway genes and GNAQ, GNA11, RASA1 and TEK. Additionally, 914 patients previously reported were systematically reviewed. RESULTS: 93 of our 150 patients had PIK3CA pathogenetic variants. The merged PROS cohort showed that PIK3CA variants span thorough all gene domains, some were exclusively associated with specific PROS phenotypes: weakly activating variants were associated with central nervous system (CNS) involvement, and strongly activating variants with extra-CNS phenotypes. Among the 57 with a wild-type PIK3CA allele, 11 patients with overgrowth and vascular malformations overlapping PROS had variants in GNAQ, GNA11, RASA1 or TEK. CONCLUSION: We confirm that (1) molecular diagnostic yield increases when multiple tissues are tested and by enriching NGS panels with genes of overlapping 'vascular' phenotypes; (2) strongly activating PIK3CA variants are found in affected tissue, rarely in blood: conversely, weakly activating mutations more common in blood; (3) weakly activating variants correlate with CNS involvement, strong variants are more common in cases without; (4) patients with vascular malformations overlapping those of PROS can harbour variants in genes other than PIK3CA.

Severe impact of late diagnosis of congenital adrenal hyperplasia on gender identity, sexual orientation and function: case report and review of the literature.

Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) represents the most frequent form of CAH and of 46, XX disorder of sex development in female newborns. In the majority of cases, particularly in developed countries, female patients suffering from the classic forms of CAH reach the diagnosis at birth or in the early childhood, allowing a prompt treatment with a correct gender assignment. The current manuscript describes an unusual case of an Italian 46-year-old woman, homeborn in the 60s, receiving an extraordinarily late diagnosis of simple virilising classic form of CAH due to 21-OHD, determining a relevant impairment of both physical and psychosexual development. The patient presented primary amenorrhea, height under target, overweight with visceral adiposity, hypercholesterolemia and insulin resistance, hirsutism with a typical male-pattern hair growth, external genital ambiguity, and a severe impairment in the entire series of psychological dimensions, particularly severe depressive symptoms, together with gender dysphoria relative to the female gender assigned at birth, cross-gender behaviours, and body image discomfort, which were associated with homosexual orientation, and sexual dysfunction. Following diagnosis and glucocorticoid (GC) replacement therapy, the hyperandrogenism control and familial and socio-cultural factors changes, particularly, living alone and the interruption of social isolation, were accompanied by menarche appearance, improvement in hirsutism and metabolic profile, and a resolution in all psychological dimensions, depressive symptoms, and gender dysphoria. The patient began to perceive homosexual orientation without discomfort, and ameliorating sexual function. Few cases of female patients with CAH due to 21-OHD receiving an extremely delayed diagnosis have been published. However, to the best of our knowledge, this is the first case including a complete psychosexual assessment at diagnosis with a detailed re-evaluation after 5 years of disease treatment.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) antigen detection in the Emergency Department: data from a pediatric cohort during the fourth COVID-19 wave in Italy.

BACKGROUND: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic has been challenging health care systems and made it necessary to use rapid and cost-effective testing methods, particularly in Emergency Department (ED) settings. Rapid Antigen Diagnostic Tests (RADTs) are a valid alternative to the gold standard RT-PCR, even in pediatric populations. This retrospective observational study has been conducted on a pediatric cohort afferent to the ED of the San Giovanni di Dio and Ruggi d'Aragona University Hospital in Salerno, tested at Point of Care with RADT Panbio® (Abbott), from September 1st, 2021 to February 28th, 2022, analyzing the positivity rate and clinical features of the cohort, also in reference to the rise of positive cases observed in the aforementioned period, and to the introduction in Italy of SARS-CoV-2 vaccination for children and teens on December 16th, 2021. METHODS: Data regarding access to the pediatric ED were extracted from the hospital's electronic database system. Parallel to this, we conducted a narrative literature search using PubMed database focusing on the use of RADT in pediatric ED and compared our data with the national pandemic trend. RESULTS: During the observation period, 1890 patients were tested for the presence of SARS-CoV-2 with RADT and the 2.7% of children resulted positive, with a peak in January 2022. The main symptoms in positive patients were: fever (n = 34; 66.7%), cough (n = 11; 21.5%), headache (n = 4; 7.8%), chest pain (n = 2; 3.9%) and abdominal pain (n = 1; 2%). Patients were divided into three different age groups (A, B, C) basing on the different access timing to vaccination; no statistically significant difference was detected in the distribution of positivity in these three groups (p > 0.05). Number of positive children in group A was greater in the post-vaccine group. Our data are concordant with the national trend of the pandemic showing a fourth wave peak in January 2022. CONCLUSION: The use of RADT as a first point-of-care screening may be helpful, time-saving and cost-sparing. Our study shows that, during the observation period, most children admitted to the ED for fever, actually tested positive for SARS-CoV-2 with a statistically greater difference than negative children. Instead, number of patients admitted for cough was statistically higher among negative than positive ones, probably due to the circulation of other respiratory viruses in children.

Clinical and molecular characterization of patients affected by Beckwith-Wiedemann spectrum conceived through assisted reproduction techniques.

The prevalence of Beckwith-Wiedemann spectrum (BWSp) is tenfold increased in children conceived through assisted reproductive techniques (ART). More than 90% of ART-BWSp patients reported so far display imprinting center 2 loss-of-methylations (IC2-LoM), versus 50% of naturally conceived BWSp patients. We describe a cohort of 74 ART-BWSp patients comparing their features with a cohort of naturally conceived BWSp patients, with the ART-BWSp patients previously described in literature, and with the general population of children born from ART. We found that the distribution of UPD(11)pat was not significantly different in ART and naturally conceived patients. We observed 68.9% of IC2-LoM and 16.2% of mosaic UPD(11)pat in our ART cohort, that strongly differ from the figure reported in other cohorts so far. Since UPD(11)pat likely results from post-fertilization recombination events, our findings allows to hypothesize that more complex molecular mechanisms, besides methylation disturbances, may underlie BWSp increased risk in ART pregnancies. Moreover, comparing the clinical features of ART and non-ART BWSp patients, we found that ART-BWSp patients might have a milder phenotype. Finally, our data show a progressive increase in the prevalence of BWSp over time, paralleling that of ART usage in the last decades.

Bone metabolism in patients with type 1 neurofibromatosis: key role of sun exposure and physical activity.

Bone metabolism has been rarely investigated in children affected by Neurofibromatosis type 1 (NF1). Aim of the present study was to assess bone mineral metabolism in children and adults NF1 patients, to determine the relevant factors potentially involved in the development of reduced bone mineral density (BMD), and provide possible therapeutic intervention in NF1 patients. 114 NF1 patients and sex and age matched controls were enrolled into the study. Clinical and biochemical factors reflecting bone metabolism were evaluated. Factors potentially affecting BMD were also investigated including: physical activity, sun exposure, vitamin D intake. Whenever the presence of vitamin D deficiency was recorded, cholecalciferol supplementation was started and z-score data obtained at Dual-Energy X-ray Absorptiometry (DXA) during supplementation were compared with previous ones. NF1 patients showed lower Z-scores at Dual-Energy X-ray Absorptiometry DXA than controls. Physical activity was significantly reduced in NF1 patients than in controls. Sun exposure was significantly lower in NF1 compared to control subjects. At linear regression analysis vitamin D was the most predictive factor of reduced z-score at DXA (p = 0.0001). Cholecalciferol supplementation significantly increased BMD z-score (p < 0.001). We speculated that a combination of different factors, including reduced sun exposure, possibly associated with reduced serum vitamin D levels, and poor physical activity, concur to the impaired bone status in NF1 patients. We also demonstrated that treatment with vitamin D can be effective in improving z-score value in NF1 patients, including children. In conclusion, the findings of the current study are expected to have important implications for the follow-up and prevention of osteopenia/osteoporosis in this common genetic disease.

DNA methylation episignature testing improves molecular diagnosis of Mendelian chromatinopathies.

PURPOSE: Chromatinopathies include more than 50 disorders caused by disease-causing variants of various components of chromatin structure and function. Many of these disorders exhibit unique genome-wide DNA methylation profiles, known as episignatures. In this study, the methylation profile of a large cohort of individuals with chromatinopathies was analyzed for episignature detection. METHODS: DNA methylation data was generated on extracted blood samples from 129 affected individuals with the Illumina Infinium EPIC arrays and analyzed using an established bioinformatic pipeline. RESULTS: The DNA methylation profiles matched and confirmed the sequence findings in both the discovery and validation cohorts. Twenty-five affected individuals carrying a variant of uncertain significance, did not show a methylation profile matching any of the known episignatures. Three additional variant of uncertain significance cases with an identified KDM6A variant were re-classified as likely pathogenic (n = 2) or re-assigned as Wolf-Hirschhorn syndrome (n = 1). Thirty of the 33 Next Generation Sequencing negative cases did not match a defined episignature while three matched Kabuki syndrome, Rubinstein-Taybi syndrome and BAFopathy respectively. CONCLUSION: With the expanding clinical utility of the EpiSign assay, DNA methylation analysis should be considered part of the testing cascade for individuals presenting with clinical features of Mendelian chromatinopathy disorders.

Clinical heterogeneity of Kabuki syndrome in a cohort of Italian patients and review of the literature.

Kabuki syndrome (KS) is a well-recognized disorder characterized by postnatal growth deficiency, dysmorphic facial features, skeletal anomalies, and intellectual disability. The syndrome is caused by KMT2D gene mutations or less frequently KDM6A gene mutations or deletions. We report a systematic evaluation of KS patients from Campania region of Italy; data were also compared with literature ones. We collected data of 15 subjects (8 males and 7 females with age range 10-26 years; mean age 16.9 years) with confirmed diagnosis of KS, representing the entire cohort of patients from Campania Region. Each patient performed biochemical testing and instrumental investigation. Neuro-intellectual development, cranio-facial dysmorphisms, and multisystem involvement data were collected retrospectively. For each category, type of defects and frequency of the anomalies were analyzed. Our observation shows that KS patients from Campania region have some particular and previously underscored, neurological and immunological findings. We found high prevalence of EEG's abnormalities (43%) and MRI brain abnormalities (60%). Microcephaly resulted more common in our series (33%), if compared with major cohorts described in literature. Biochemical features of immunodeficiency and autoimmune diseases including thyroid autoimmunity, polyserositis, and vitiligo were observed with high prevalence (54.5%). Low immunoglobulins levels were a frequent finding. Lymphocyte class investigation showed significantly reduced CD8 levels in one patient.Conclusions: These data confirm great heterogeneity of clinical manifestations in KS and suggest to introduce further clinical diagnostic criteria in order to perform a correct and precocious diagnosis. What is Known • Kabuki syndrome is characterized by growth deficiency, dysmorphic facial features, skeletal anomalies, and intellectual disability • Immune dysfunction is a common finding but autoimmune diseases are rarely seen • Neurological features are common What is New • Some particular facial features could help gestalt diagnosis (hypertelorism, broad nasal bridge, micrognathia, tooth agenesis, cutaneous haemangiomas and strabismus) • Higher prevalence of autoimmune disorders than previously reported • Particular neurological features are present in this cohort (EEG and MRI brain abnormalities).

RASopathies and hemostatic abnormalities: key role of platelet dysfunction.

BACKGROUND: Bleeding anomalies have been reported in patients affected by Noonan syndrome. No study has been performed in patients with molecularly confirmed RASopathy. We aimed to characterize the frequency and types of bleeding disorders in patients with RASopathies and evaluate any significant association with laboratory findings. PATIENTS AND METHODS: Forty-nine individuals (PTPN11, n = 27; SOS1, n = 7; RIT1, n = 3; SPRED1, n = 1; LZTR1, N = 3; RAF1, n = 2; BRAF, n = 4; MEK1, n = 1; MEK2, n = 1), and 49 age- and sex-matched controls were enrolled. The "Paediatric Bleeding Questionnaire Scoring Key" was administered to patients and families. Laboratory screening tests including clotting factors dosing, platelet count, Prothrombin Time and Partial Thromboplastin Time, were employed both in patients and controls to characterize the bleeding diathesis. A subgroup of 29/49 patients and 29/49 controls was also tested for platelet function. RESULTS: Regardless of the gene involved, pathological paediatric bleeding scores were recorded in 14/49 (28.5%) patients. Indeed, 7 were mutated in PTPN11, 3 in SOS1, 2 in RIT1, 1 in BRAF, and 1 in MEK1. Compared to patients with normal bleeding scores, those with pathologic bleeding score showed higher prevalence of splenomegaly (p = 0.006), prolonged aPTT (p = 0.04), lower levels of coagulation factor V (FV, p = 0.001), FVII (p = 0.003), FX (p = 0.0008) and FXIII (p = 0.002), higher vWAg (p = 0.04), and lower platelet sensitivity to Ristocetin (p = 0.001), arachidonic acid (AA) (p = 0.009) and collagen (p = 0.01). The presence of hematomas inversely correlated with factor V (p = 0.002), factor VII (p = 0.003), factor X (p = 0.002) and factor XIII (p = 0.004) levels, and directly correlated with platelet response to collagen (p = 0.02) and AA (p = 0.01). The presence of splenomegaly directly correlated with the presence of hematoma (p = 0.006), platelet response to Ristocetin (p = 0.04) and AA (p = 0.04), and inversely correlated with factor V levels (p = 0.03). CONCLUSIONS: Patients with RASopathies and a bleeding tendency exhibit multiple laboratory abnormalities, including platelet-related disorders. Splenomegaly is frequently detected and might be a suggestive sign for qualitative platelet dysfunction. A comprehensive clinical assessment should be carried out at diagnosis, during the follow-up and before any surgical procedures. Since there is currently no consensus on management of bleeding complications, it is important that physicians closely monitor these patients.

The SGLT2-inhibitor dapagliflozin improves neutropenia and neutrophil dysfunction in a mouse model of the inherited metabolic disorder GSDIb.

Glycogen Storage Disease type 1b (GSDIb) is a genetic disorder with long term severe complications. Accumulation of the glucose analog 1,5-anhydroglucitol-6-phosphate (1,5AG6P) in neutrophils inhibits the phosphorylation of glucose in these cells, causing neutropenia and neutrophil dysfunctions. This condition leads to serious infections and inflammatory bowel disease (IBD) in GSDIb patients. We show here that dapagliflozin, an inhibitor of the renal sodium-glucose co-transporter-2 (SGLT2), improves neutrophil function in an inducible mouse model of GSDIb by reducing 1,5AG6P accumulation in myeloid cells.

Clinical variability of neurofibromatosis 1: A modifying role of cooccurring PTPN11 variants and atypical brain MRI findings.

Neurofibromatosis 1 (NF1) is a disorder characterized by variable expressivity caused by loss-of-function variants in NF1, encoding neurofibromin, a protein negatively controlling RAS signaling. We evaluated whether concurrent variation in proteins functionally linked to neurofibromin contribute to the variable expressivity of NF1. Parallel sequencing of a RASopathy gene panel in 138 individuals with molecularly confirmed clinical diagnosis of NF1 identified missense variants in PTPN11, encoding SHP2, a positive regulator of RAS signaling, in four subjects from three unrelated families. Three subjects were heterozygous for a gain-of-function variant and showed a severe expression of NF1 (developmental delay, multiple cerebral neoplasms and peculiar cortical MRI findings), and features resembling Noonan syndrome (a RASopathy caused by activating variants in PTPN11). Conversely, the fourth subject, who showed an attenuated presentation, carried a previously unreported PTPN11 variant that had a hypomorphic behavior in vitro. Our findings document that functionally relevant PTPN11 variants occur in a small but significant proportion of subjects with NF1 modulating disease presentation, suggesting a model in which the clinical expression of pathogenic NF1 variants is modified by concomitant dysregulation of protein(s) functionally linked to neurofibromin. We also suggest targeting of SHP2 function as an approach to treat evolutive complications of NF1.