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.

Beckwith-Wiedemann syndrome mimicking the classical form of congenital adrenal hyperplasia in newborn screening.

Beckwith-Wiedemann syndrome (BWS) is a common genetic congenital disease characterized by somatic overgrowth and its broad clinical spectrum includes pre- and post-natal macrosomia, macroglossia, visceromegaly, increased risk of neonatal hypoglycemia, and development of embryonic tumors. BWS occurs due to genetic/epigenetic changes involving growth-regulating genes, located on region 11p15, with an important genotype-phenotype correlation. Congenital adrenal hyperplasia (CAH) comprises a spectrum of autosomal recessive diseases presenting a variety of clinical manifestations due to a deficiency in one of the enzymes involved in cortisol secretion. Early diagnosis based on newborn screening prevents the adrenal crisis and early infant death. However, high 17-hydroxyprogesterone (17-OHP) levels can occur in newborns or premature infants without CAH, in situations of stress due to maternal or neonatal factors. Here, we report new cases of false-positive diagnosis of 21-hydroxylase deficiency during newborn screening - two girls and one boy with BWS. Methylation-specific multiplex ligation-dependent probe amplification revealed a gain of methylation in the H19 differentially methylated region. Notably, all three cases showed a complete normalization of biochemical changes, highlighting the transient nature of these hormonal findings that imitate the classical form of CAH. This report sheds light on a new cause of false-positive 21-hydroxylase deficiency diagnosis during newborn screening: Beckwith-Wiedemann syndrome.

A supervised learning method for classifying methylation disorders.

BACKGROUND: DNA methylation is one of the most stable and well-characterized epigenetic alterations in humans. Accordingly, it has already found clinical utility as a molecular biomarker in a variety of disease contexts. Existing methods for clinical diagnosis of methylation-related disorders focus on outlier detection in a small number of CpG sites using standardized cutoffs which differentiate healthy from abnormal methylation levels. The standardized cutoff values used in these methods do not take into account methylation patterns which are known to differ between the sexes and with age. RESULTS: Here we profile genome-wide DNA methylation from blood samples drawn from within a cohort composed of healthy controls of different age and sex alongside patients with Prader-Willi syndrome (PWS), Beckwith-Wiedemann syndrome, Fragile-X syndrome, Angelman syndrome, and Silver-Russell syndrome. We propose a Generalized Additive Model to perform age and sex adjusted outlier analysis of around 700,000 CpG sites throughout the human genome. Utilizing z-scores among the cohort for each site, we deployed an ensemble based machine learning pipeline and achieved a combined prediction accuracy of 0.96 (Binomial 95% Confidence Interval 0.868[Formula: see text]0.995). CONCLUSION: We demonstrate a method for age and sex adjusted outlier detection of differentially methylated loci based on a large cohort of healthy individuals. We present a custom machine learning pipeline utilizing this outlier analysis to classify samples for potential methylation associated congenital disorders. These methods are able to achieve high accuracy when used with machine learning methods to classify abnormal methylation patterns.

Development of disease-specific growth charts for Korean children with Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann syndrome (BWS) is an epigenetic overgrowth syndrome. Despite its distinctive growth pattern, the detailed growth trajectories of children with BWS remain largely unknown. We retrospectively analyzed 413 anthropometric measurements over an average of 4.4 years of follow-up in 51 children with BWS. We constructed sex-specific percentile curves for height, weight, and head circumference using a generalized additive model for location, scale, and shape. Males with BWS exhibited greater height at all ages evaluated, weight before the age of 10, and head circumference before the age of 9 than those of the general population. Females with BWS showed greater height before the age of 7, weight before the age of 4.5, and head circumference before the age of 7 than those of the general population. At the latest follow-up visit at a mean 8.4 years of age, bone age was significantly higher than chronological age. Compared to paternal uniparental disomy (pUPD), males with imprinting center region 2-loss of methylation (IC2-LOM) had higher standard deviation score (SDS) for height and weight, while females with IC2-LOM showed larger SDS for head circumference. These disease-specific growth charts can serve as valuable tools for clinical monitoring of children with BWS.

Pediatric cancer incidence among individuals with overgrowth syndromes and overgrowth features: A population-based assessment in seven million children.

BACKGROUND: Overgrowth syndromes (e.g., Beckwith-Wiedemann) are associated with an increased risk of pediatric cancer, although there are few population-based estimates of risk. There are also limited studies describing associations between other overgrowth features (e.g., hepatosplenomegaly) and pediatric cancer. Therefore, cancer risk among children with these conditions was evaluated with data from a large, diverse population-based registry linkage study. METHODS: This study includes all live births in Texas during the years 1999-2017. Children with overgrowth features and syndromes were identified from the Texas Birth Defects Registry; children with cancer were identified by linkage to the Texas Cancer Registry. Cox regression models were used to estimate the hazard ratio (HR) and 95% confidence interval (CI) for the association between each overgrowth syndrome/feature and cancer, which were adjusted for infant sex and maternal age. RESULTS: In the total birth cohort (n = 6,997,422), 21,207 children were identified as having an overgrowth syndrome or feature. Children with Beckwith-Wiedemann syndrome were 42 times more likely to develop pediatric cancer (95% CI, 24.20-71.83), with hepatoblastoma being the most common, followed by Wilms tumor. The presence of any isolated overgrowth feature was associated with increased cancer risk (HR, 4.70; 95% CI, 3.83-5.77); associations were strongest for hepatosplenomegaly (HR, 23.04; 95% CI, 13.37-39.69) and macroglossia (HR, 11.18; 95% CI, 6.35-19.70). CONCLUSIONS: This population-based assessment confirmed prior findings that children with either overgrowth syndromes or features were significantly more likely to develop cancer. Overall, this study supports recommendations for cancer surveillance in children with these conditions and may also inform future research into cancer etiology.

Cancer predisposition signaling in Beckwith-Wiedemann Syndrome drives Wilms tumor development.

BACKGROUND: Wilms tumor (WT) exhibits structural and epigenetic changes at chromosome 11p15, which also cause Beckwith-Wiedemann Syndrome (BWS). Children diagnosed with BWS have increased risk for WT. The aim of this study is to identify the molecular signaling signatures in BWS driving these tumors. METHODS: We performed whole exome sequencing, methylation array analysis, and gene expression analysis on BWS-WT samples. Our data were compared to publicly available nonBWS data. We categorized WT from BWS and nonBWS patients by assessment of 11p15 methylation status and defined 5 groups- control kidney, BWS-nontumor kidney, BWS-WT, normal-11p15 nonBWS-WT, altered-11p15 nonBWS-WT. RESULTS: BWS-WT samples showed single nucleotide variants in BCORL1, ASXL1, ATM and AXL but absence of recurrent gene mutations associated with sporadic WT. We defined a narrow methylation range stratifying nonBWS-WT samples. BWS-WT and altered-11p15 nonBWS-WT showed enrichment of common and unique molecular signatures based on global differential methylation and gene expression analysis. CTNNB1 overexpression and broad range of interactions were seen in the BWS-WT interactome study. CONCLUSION: While WT predisposition in BWS is well-established, as are 11p15 alterations in nonBWS-WT, this study focused on stratifying tumor genomics by 11p15 status. Further investigation of our findings may identify novel therapeutic targets in WT oncogenesis.

Preneoplastic liver colonization by 11p15.5 altered mosaic cells in young children with hepatoblastoma.

Pediatric liver tumors are very rare tumors with the most common diagnosis being hepatoblastoma. While hepatoblastomas are predominantly sporadic, around 15% of cases develop as part of predisposition syndromes such as Beckwith-Wiedemann (11p15.5 locus altered). Here, we identify mosaic genetic alterations of 11p15.5 locus in the liver of hepatoblastoma patients without a clinical diagnosis of Beckwith-Wiedemann syndrome. We do not retrieve these alterations in children with other types of pediatric liver tumors. We show that mosaic 11p15.5 alterations in liver FFPE sections of hepatoblastoma patients display IGF2 overexpression and H19 downregulation together with an alteration of the liver zonation. Moreover, mosaic livers' microenvironment is enriched in extracellular matrix and angiogenesis. Spatial transcriptomics and single-nucleus RNAseq analyses identify a 60-gene signature in 11p15.5 altered hepatocytes. These data provide insights for 11p15.5 mosaicism detection and its functional consequences during the early steps of carcinogenesis.

Beckwith-Wiedemann syndrome and twinning: case report and brief review of literature.

BACKGROUND: Beckwith-Wiedemann syndrome (BWS, OMIM #130,650) is a pediatric overgrowth disorder involving a predisposition to tumor development. Although the clinical management of affected patients is well established, it is less clear how to handle with the cases of siblings of affected patients, since the prevalence of the condition in twins (1:1000) is ten times higher than in singletones (1:10000). CASE PRESENTATION: We report the case of a premature twin patient who during her follow-up develops a clinical phenotype compatible with BWS, genetically confirmed in blood. However, the methylation alteration characteristic of the condition was also found in the almost phenotypically normal sibling, making it challening her management. CONCLUSION: Through our case report we highlight how the diagnosis of BWS can be made without any prenatal suspicion and we propose a review of the literature on how to manage siblings of affected patients in twinning situation.

Beckwith-Wiedemann Syndrome in Newborn of Mother with HELLP Syndrome/Preeclampsia: An Analysis of Literature and Case Report with Fetal Growth Restriction and Absence of CDKN1C Typical Pathogenic Genetic Variation.

Beckwith-Wiedemann Syndrome (BWS) is an imprinting disorder, which manifests by overgrowth and predisposition to embryonal tumors. The evidence on the relationship between maternal complications such as HELLP (hemolysis, elevated liver enzymes, and low platelet count) and preeclampsia and the development of BWS in offspring is scarce. A comprehensive clinical evaluation, with genetic testing focused on screening for mutations in the CDKN1C gene, which is commonly associated with BWS, was conducted in a newborn diagnosed with BWS born to a mother with a history of preeclampsia and HELLP syndrome. The case study revealed typical clinical manifestations of BWS in the newborn, including hemihyperplasia, macroglossia, midfacial hypoplasia, omphalocele, and hypoglycemia. Surprisingly, the infant also exhibited fetal growth restriction, a finding less commonly observed in BWS cases. Genetic analysis, however, showed no mutations in the CDKN1C gene, which contrasts with the majority of BWS cases. This case report highlights the complex nature of BWS and its potential association with maternal complications such as preeclampsia and HELLP syndrome. The atypical presence of fetal growth restriction in the newborn and the absence of CDKN1C gene mutations have not been reported to date in BWS.

Associations between the timing of tongue reduction surgery, (Epi)genotype, and dentoskeletal development in patients with Beckwith-Wiedemann syndrome.

Tongue reduction surgery is often pursued to manage the adverse effects of macroglossia in patients with Beckwith-Wiedemann syndrome (BWS). This study characterized dental outcomes in patients with BWS based on surgical timing and molecular diagnosis. A retrospective study was designed to include patients with BWS over the age of two who had clinical or radiographic documentation of dental development. Patients were grouped by history of tongue reduction surgery and surgical timing (early: <12 months). One hundred three patients were included (55 no tongue reduction, 18 early, 30 late). Patients who underwent late surgery had lower odds of class I occlusion (OR 0.11, 95% CI 0.02-0.58, p = 0.009) and higher odds of anterior open bite (OR 7.5, 95% CI 1.14-49.4, p = 0.036). Patients with clinical diagnoses and negative molecular testing had anterior open bite less frequently than patients with imprinting center 2 loss of methylation and paternal uniparental isodisomy of 11p15.5 (p = 0.009). Compared to reference values, patients who had tongue reductions had an increased mandibular plane angle (32.0 ± 4.5° versus 36.9 ± 5.0°, p = 0.001), indicative of hyperdivergent growth. The results of this study help to understand the complex nature of dentoskeletal growth in BWS and shed insight on how surgical timing and molecular diagnosis influence prognosis.

First-time application of droplet digital PCR for methylation testing of the 11p15.5 imprinting regions.

BACKGROUND: Beckwith-Wiedemann syndrome and Silver-Russel syndrome are two imprinting disorders caused by opposite molecular alterations in 11p15.5. With the current diagnostic tests, their molecular diagnosis is challenging due to molecular heterogeneity and mosaic occurrence of the most frequent alterations. As the determination of precise (epi)genotype of patients is relevant as the basis for a personalized treatment, different approaches are needed to increase the sensitivity of diagnostic testing of imprinting disorders. METHODS: We established methylation-specific droplet digital PCR approaches (MS-ddPCR) for the two imprinting centers in 11p15.5, and analyzed patients with paternal uniparental disomy of chromosome 11p15.5 (upd(11)pat) and other imprinting defects in the region. The results were compared to those from MS-MLPA (multiplex ligation-dependent probe amplification) and MS-pyrosequencing. RESULTS: MS-ddPCR confirmed the molecular alterations in all patients and the results matched well with MS-MLPA. The results of MS-pyrosequencing varied between different runs, whereas MS-ddPCR results were reproducible. CONCLUSION: We show for the first time that MS-ddPCR is a reliable and easy applicable method for determination of MS-associated changes in imprinting disorders. It is therefore an additional tool for multimethod diagnostics of imprinting disorders suitable to improve the diagnostic yield.

Tandem Triplication 11p15.5-ICR1 (H19/IGF2) Detected by Microarray and Optical Genome Mapping in a Prenatal Beckwith-Wiedemann Case.

Optical genome mapping (OGM) appears as a new tool for matching standard cytogenetic methods (karyotype and microarray) into a single assay. The chromosomal region 11p15.5 harbours two differentially methylated regions, the imprinting centre regions 1 and 2 (ICR1, ICR2). Disturbances in both regions alter human growth and are associated with two imprinting disorders, Beckwith-Wiedemann (BWS) and Silver-Russell syndromes. Herein, we present a prenatal case with a triplication in 11p15.5, including the H19/IGF2 imprinted region, detected by microarray and OGM. A 30-year-old pregnant woman of 17 weeks of gestation was referred for prenatal karyotype and microarray study because of increased nuchal translucency, short femur, megabladder, hyperechogenic bowel, and renal ectasia. Microarray, OGM, and MS-MLPA were performed, and a tandem cis-triplication in 11p15.5 and hypermethylation of the ICR1 region, compatible with BWS was detected. OGM, with its power to detect all classes of structural variants, including copy number variants, at a higher resolution than traditional cytogenetic methods can play a significant role in prenatal care and management as a next-generation cytogenomic tool. This study further supports the hypotheses that the amplification/duplication-triplication of the H19/IGF2 region could be related to BWS if it is of paternal origin.

Adult experiences in Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann syndrome (BWS) is an overgrowth and epigenetic disorder caused by changes on chromosome 11p15. The primary features requiring management in childhood include macroglossia, omphalocele, lateralized overgrowth, hyperinsulinism, and embryonal tumors. Management guidelines have not been developed for adults with BWS and there have been few studies to assess the clinical needs of these patients. Furthermore, there have been few studies on the psychosocial implications of BWS in children or adults. Here, we present a descriptive summary of data gathered from two separate adult BWS cohorts. The first, a patient-based survey cohort, includes self-reported health information and recollections about BWS experiences, while the second provides results of a medical record-based assessment from patients in an overgrowth registry. Results highlight the clinical features and medical issues affecting two large independent cohorts of adults with BWS while noting similarities. Open-ended questions asked of the survey cohort yielded themes to guide future qualitative studies. Finally, the study demonstrated the reliability of patient-reported data and the utility of international partnerships in this context.

Concurrent Hepatoblastoma and Wilms Tumor Leading to Diagnosis of Beckwith-Wiedemann Syndrome.

Beckwith-Wiedemann syndrome (BWS) is an epigenetic overgrowth disorder and cancer predisposition syndrome caused by imprinting defects of chromosome 11p15.5-11p15.4. BWS should be considered in children with atypical presentations of embryonal tumors regardless of clinical phenotype. Risk of malignancy correlates with specific molecular subgroups of BWS making molecular subclassification important for appropriate cancer screening. We report the first case of concurrent embryonal tumors in a phenotypically normal child, leading to the diagnosis of BWS with paternal uniparental disomy and describe the molecular classification of BWS as it relates to malignancy risk, along with approach to management.

Lateralized overgrowth as a guiding sign of abdominal neoplasms for pediatric orthopedic surgeons.

OBJECTIVES: This study aims to increase the awareness of the association between lateralized overgrowth (LO) and abdominal tumor among the pediatric orthopedic community and to evaluate its incidence in our center. PATIENTS AND METHODS: Between January 1997 and December 2021, a total of 166 patients with Wilms tumors and hepatoblastomas were retrospectively analyzed. Data including age, sex, initial clinical signs (hematuria, abdominal mass with or without general discomfort), type of asymmetric regional body overgrowth (isolated or in relation with any syndrome), and tumor stage at diagnosis were recorded. In addition, age at which asymmetric regional body overgrowth was described and age at the time of tumor diagnosis were noted. RESULTS: Of a total of 166 patients, 133 were diagnosed with Wilms tumors (nephroblastomas) and 33 were diagnosed with hepatoblastomas. In 94% of the cases, the initial clinical signs were an abdominal mass and/or hematuria. Overall, five (3%) patients presented with LO. Four patients with Wilms tumor presented it at the initial clinical examinations. In three of these cases (2.3%), we found it isolated and, in the remaining patient (0.75%), it was associated with Beckwith-Wiedemann spectrum. Only one patient affected from hepatoblastoma (3%) presented with an isolated LO at the time of tumor diagnosis. CONCLUSION: Our study results show an incidence of LO in relation to intra-abdominal tumors of 3%. The latest updates recommend genetic testing to identify subgroups with a higher risk for tumor development that are more likely to benefit from tumor protocol surveillance.

Investigation of a pervasive immune, cardiac, and behavioral phenotype in Beckwith-Wiedemann syndrome: A case report.

Beckwith-Wiedemann syndrome (BWS) is an imprinting disorder caused by genetic and epigenetic changes in the chromosome 11p15 region. The syndrome is characterized by a wide range of features including macrosomia, lateralized overgrowth, abdominal wall defects, and hypoglycemia. BWS presentation is variable across the entire patient population, but certain areas including immunology, cardiology, and behavioral differences are not well characterized. We present a case of a male patient with BWS due to the most common cause of BWS, loss of methylation at imprinting center 2 with a variable phenotype, including classical features (macrosomia, macroglossia, omphalocele, placentomegaly and mild lateralized overgrowth) in addition to uncommon features (immune deficiency, developmental delays, and pulmonary stenosis) not typically seen in BWS. This study defines a patient's clinical presentation and course and highlights the need to consider atypical organ systems in BWS as either an expansion of the phenotype or co-existing conditions to develop personalized care models.

Mosaic genome-wide paternal uniparental disomy after discordant results from primary fetal samples and cultured cells.

Mosaic genome-wide paternal uniparental disomy (GWpUPD) is a rare condition in which two euploid cell lines coexist in the same individual, one with biparental content and one with genome-wide paternal isodisomy. We report a complex prenatal diagnosis with discordant results from cultured and uncultured samples. A pregnant woman was referred for placental mesenchymal dysplasia and fetal omphalocele. Karyotype, array-CGH and Beckwith-Wiedemann Syndrome (BWS) testing (methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) of 11p15) performed on amniocytes were negative. After intrauterine fetal demise, the clinical suspicion persisted and BWS MS-MLPA was repeated on cultured cells from umbilical cord and amniotic fluid, revealing a mosaicism for KvH19 hypermethylation/KCNQ1OT1:TSS:DMR hypomethylation. These results, along with microsatellite analysis of the BWS region, were consistent with mosaic paternal 11p15 isodisomy. A concurrent maternal contamination exclusion test, analyzing polymorphic microsatellite markers on multiple chromosomes, showed an imbalance in favor of paternal alleles at all examined loci on cultured amniocytes and umbilical cord samples. This led to suspicion of mosaic GWpUPD, later confirmed by SNP-array, identifying a mosaic genome-wide paternal isodisomy affecting 60% of fetal cells. The assessment of mosaic GWpUPD requires multiple approaches beyond the current established diagnostic processes, also entertaining possible low-rate mosaicism. Clinical acumen and an integrated testing approach are the key to a successful diagnosis.

A 132 bp deletion affecting the KCNQ1OT1 gene associated with Silver-Russell syndrome clinical phenotype.

BACKGROUND: Imprinting centre 2 (IC2) in the chromosomal region 11p15.5 regulates the monoallelic expression of imprinted genes by differential methylation of paternal and maternal chromosomes. Copy number variants in IC2 are associated with Beckwith-Wiedemann syndrome and Silver-Russell syndrome (SRS). Clinical outcome of IC2 deletions seems to depend on the parental origin of the chromosome, deletion size and inclusion or exclusion of enhancer and promoter regions. RESULTS: A paternally inherited 132 bp deletion within the KCNQ1OT1 gene was found in a proband with an SRS clinical phenotype. The patient's father and paternal grandmother, who both carry the deletion on their maternal chromosome, are unaffected. Review of other IC2 deletions and their associated clinical presentation was useful in understanding the genetic-phenotypic correlation. CONCLUSION: Only six cases have been reported with deletions involving exclusively IC2, one being identical to our proband's 132 bp deletion. Our study, which is based on more extensive segregation data than the previous 132 bp deletion report, confirms the association of this deletion with growth restriction when paternally inherited. Remarkably, even though our patient has the same deletion, he has more pronounced phenotypic features; our findings thus suggest that some degree of clinical variability may be associated with this loss.