Beckwith-Wiedemann Syndrome Phenotype Research Articles

6775 Four Cases with the Beckwith-Wiedemann Syndrome Phenotype Caused by GNAS Defects

Abstract Disclosure: T. Urakawa: None. Y. Kanamaru: None. N. Amano: None. A. Uchida: None. J. Shin: None. M. Fukami: None. M. Kagami: None. Background: Beckwith-Wiedemann syndrome (BWS) is an overgrowth disorder with a heterogeneous phenotypic spectrum, including macroglossia (MG), lateralized overgrowth (LO), and hyperinsulinism (HI). According to the BWS clinical scoring system, the consensus statement recommends conducting genetic testing for patients with two or more scores and doing the clinical diagnosis of BWS for patients with four or more scores. The etiologies of BWS consist of methylation abnormalities of differentially methylated regions (DMRs) on 11p15.5, paternal uniparental disomy of chromosome 11 (UPD(11)pat), and maternal loss of function variants of CDKN1C. However, in 20% of BWS cases, the genetic causes remain unclear. Here, we report on four cases of the BWS phenotype with GNAS defects, known to cause pseudohypoparathyroidism (PHP). Case presentation: Case 1 was born at 37 weeks of gestation with birth weight (BW) of 2964 g (+0.8 SD) and birth length (BL) of 50.5 cm (+1.3 SD) and had MG, LO, HI, ear creases (EC), naevus flammeus, umbilical hernia (UH), and postnatal overgrowth (PO) together with intellectual disability (BWS score 9). Case 2 was born at 32 weeks with BW of 1415 g (-1.4 SD) and BL of 40.0 cm (-0.8 SD), showing MG, HI, UH, and PO (BWS score 5). Case 3 was born at 36 weeks with BW of 2366 g (-0.5 SD) and BL of 44.0 cm (-1.1 SD), exhibiting MG, EC, UH, and PO (BWS score 4). Case 4 was born at 36 weeks with BW of 2762 g (+0.6 SD) and BL of 46.0 cm (-0.4 SD), presenting MG and PO (BWS score 2). Cases 3 and 4 showed elevated intact PTH levels. Genetic analysis: Methylation-specific MLPA (MS-MLPA) for multiple DMRs showed abnormal methylation in DMRs on the GNAS locus in cases 1-3, but normal methylation levels in DMRs on 11p15.5. Case 1 had multi-locus imprinting disturbance (MLID) detected by comprehensive methylation analysis (EPIC, Illumina) and a 9pter-9p24.2 deletion identified by array CGH analysis. Case 2 also had MLID together with UPD(20)pat detected by microsatellite marker analysis. Case 3 showed only abnormal methylation in the DMRs on the GNAS locus confirmed by a comprehensive methylation analysis. Case 4 showed no abnormal methylation in the DMRs examined by MS-MLPA and had a pathogenic variant in GNAS (F219L) detected by exome sequencing. Discussion: We identified GNAS defects in four cases with the BWS phenotype. MG and PO were observed in all cases, and other BWS features in each case. Some clinical features observed in 9p terminal deletion syndrome and PHP overlap those in BWS, such as HI, MG, and PO. Therefore, these diseases should be considered as the differential diagnosis of BWS. MLID was identified in two cases, but the methylation status of DMRs on 11p15.5 was normal. The impact of MLID on the BWS phenotype is unknown. Conclusion:GNAS defects can cause the BWS phenotype. Further genetic analysis including GNAS should be considered in cases with the BWS phenotype without 11p15.5 abnormalities. Presentation: 6/2/2024

Comprehensive molecular and clinical findings in 29 patients with multi-locus imprinting disturbance

BackgroundMulti-locus imprinting disturbance (MLID) with methylation defects in various differentially methylated regions (DMRs) has recently been identified in approximately 150 cases with imprinting disorders (IDs), and deleterious variants have been found in genes related to methylation maintenance of DMRs, such as those encoding proteins constructing the subcortical maternal complex (SCMC), in a small fraction of patients and/or their mothers. However, integrated methylation analysis for DMRs and sequence analysis for MLID-causative genes in MLID cases and their mothers have been performed only in a single study focusing on Beckwith-Wiedemann syndrome (BWS) and Silver-Russell syndrome (SRS) phenotypes.ResultsOf 783 patients with various IDs we have identified to date, we examined a total of 386 patients with confirmed epimutation and 71 patients with epimutation or uniparental disomy. Consequently, we identified MLID in 29 patients with epimutation confirmed by methylation analysis for multiple ID-associated DMRs using pyrosequencing and/or methylation-specific multiple ligation-dependent probe amplification. MLID was detected in approximately 12% of patients with BWS phenotype and approximately 5% of patients with SRS phenotype, but not in patients with Kagami-Ogata syndrome, Prader-Willi syndrome, or Angelman syndrome phenotypes. We next conducted array-based methylation analysis for 78 DMRs and whole-exome sequencing in the 29 patients, revealing hypomethylation-dominant aberrant methylation patterns in various DMRs of all the patients, eight probably deleterious variants in genes for SCMC in the mothers of patients, and one homozygous deleterious variant in ZNF445 in one patient. These variants did not show gene-specific methylation disturbance patterns. Clinically, neurodevelopmental delay and/or intellectual developmental disorder (ND/IDD) was observed in about half of the MLID patients, with no association with the identified methylation disturbance patterns and genetic variants. Notably, seven patients with BWS phenotype were conceived by assisted reproductive technology (ART).ConclusionsThe frequency of MLID was 7.5% (29/386) in IDs caused by confirmed epimutation. Furthermore, we revealed diverse patterns of hypomethylation-dominant methylation defects, nine deleterious variants, ND/IDD complications in about half of the MLID patients, and a high frequency of MLID in ART-conceived patients.

Familial Beckwith-Wiedemann syndrome: Prenatal manifestation and a possible expansion of the phenotype

We describe a case of Beckwith-Wiedemann syndrome (BWS) demonstrating pre- and post-natal intra-familial variability. Our first encounter with the family occurred in the 1990s following the birth of 3 affected offspring. The first two pregnancies presented with exomphalos and elevated second trimester maternal serum alpha-fetoprotein (msAFP, 3.43 and 4.01 MOM, respectively) as well as elevated maternal human chorionic gonadotrophin (mhCG, 4.33 and 8.8 MOM, respectively). The diagnosis of BWS was confirmed postnatally in both cases. The third ongoing pregnancy presented only with elevated mhCG (7.09 MOM) and no malformation. Nonetheless BWS was suspected. The diagnosis was confirmed postnatally with clinical manifestations including macroglossia and cleft palate. Two affected female siblings were also diagnosed with Mullerian agenesis in adulthood.Suspecting a common genetic etiology, sequencing of the CDKN1C gene revealed a maternally inherited, likely pathogenic variant (NM_000076.2: c.367_385del; p.(Ala123Serfs*143)) causative of BWS. Chromosomal microarray and whole exome sequencing did not reveal any other pathogenic variant that would explain the Mullerian agenesis. One of the affected females underwent successful preimplantation genetic testing (PGT) with a surrogate and gave birth to a healthy female.To the best of our knowledge, this is the first report of Mullerian agenesis as a possible rare expansion of the BWS phenotype. In addition, this case highlights the potential role of abnormal second trimester biochemical markers (msAFP, mHCG) as possible indicators of BWS, especially in familial cases.

Unbalanced segregation of a paternal t(9;11)(p24.3;p15.4) translocation causing familial Beckwith-Wiedemann syndrome: a case report

BackgroundThe vast majority of cases with Beckwith-Wiedemann syndrome (BWS) are caused by a molecular defect in the imprinted chromosome region 11p15.5. The underlying mechanisms include epimutations, uniparental disomy, copy number variations, and structural rearrangements. In addition, maternal loss-of-function mutations in CDKN1C are found. Despite growing knowledge on BWS pathogenesis, up to 20% of patients with BWS phenotype remain without molecular diagnosis.Case presentationHerein, we report an Iranian family with two females affected with BWS in different generations. Bisulfite pyrosequencing revealed hypermethylation of the H19/IGF2: intergenic differentially methylated region (IG DMR), also known as imprinting center 1 (IC1) and hypomethylation of the KCNQ1OT1: transcriptional start site (TSS) DMR (IC2). Array CGH demonstrated an 8 Mb duplication on chromosome 11p15.5p15.4 (205,827-8,150,933) and a 1 Mb deletion on chromosome 9p24.3 (209,020-1,288,114). Chromosome painting revealed that this duplication-deficiency in both patients is due to unbalanced segregation of a paternal reciprocal t(9;11)(p24.3;p15.4) translocation.ConclusionsThis is the first report of a paternally inherited unbalanced translocation between the chromosome 9 and 11 short arms underlying familial BWS. Copy number variations involving the 11p15.5 region are detected by the consensus diagnostic algorithm. However, in complex cases which do not only affect the BWS region itself, characterization of submicroscopic chromosome rearrangements can assist to estimate the recurrence risk and possible phenotypic outcomes.

MON-LB061 Cushing Syndrome Due to an Adrenocortical Carcinoma in a Baby with Atypical Beckwith-Wiedemann Syndrome

Introduction: Beckwith-Wiedemann syndrome (BWS) is a congenital tumor-predisposition syndrome of which around 70% develops because of the methylation defects in the imprinted genes at chromosome 11p15.5. KCNQ1OT1 hypomethylation is the most common underlying genetic aberration in sporadic BWS, accounting for 50% of the sporadic cases but confers the least tumor risk. Case: We present a 5 month-old girl who presented with an excessive weight gain, arrested growth in height and head circumference and Cushingoid face starting from two months of age. She was born following an in vitro fertilization (IVF) pregnancy to non-consanguineous parents with negative family history for malignancies. She had signs of Cushing syndrome, nevus flammeus between eyebrows, microcephaly, a palpable mass on upper right abdomen and Tanner stage 2 pubic hair on physical examination. She had elevated free cortisol and suppressed ACTH and dexamethasone suppression test was consistent with Cushing Syndrome. Abdominal MRI revealed a right adrenocortical mass measuring 4x3x1.5 cm. Histopathology confirmed a diagnosis of adrenocortical carcinoma. There was no adrenal cortex cytomegaly at histopathological evaluation. Methylation analysis of the imprinted domains at chromosome 11p15.5 revealed hypomethylation at KvDMR (KCNQ1OT1 gene). Except for a nevus flammeus and adrenocortical carcinoma, no other features of BWS were present. Growth rate of height and head circumference normalised after tumor resection. Conclusion: This case establishes that KCNQ1OT1 hypomethylation should not only be considered in cases with a clear BWS phenotype but in all pediatric cases of apparently sporadic adrenocortical carcinoma. Unless otherwise noted, all abstracts presented at ENDO are embargoed until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. s presented at a news conference are embargoed until the date and time of the news conference. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.

Genome-wide methylation profiling of Beckwith-Wiedemann syndrome patients without molecular confirmation after routine diagnostics

Beckwith-Wiedemann syndrome (BWS) is caused due to the disturbance of imprinted genes at chromosome 11p15. The molecular confirmation of this syndrome is possible in approximately 85% of the cases, whereas in the remaining 15% of the cases, the underlying defect remains unclear. The goal of our research was to identify new epigenetic loci related to BWS. We studied a group of 25 patients clinically diagnosed with BWS but without molecular conformation after DNA diagnostics and performed a whole genome methylation analysis using the HumanMethylation450 Array (Illumina).We found hypermethylation throughout the methylome in two BWS patients. The hypermethylated sites in these patients overlapped and included both non-imprinted and imprinted regions. This finding was not previously described in any BWS-diagnosed patient.Furthermore, one BWS patient exhibited aberrant methylation in four maternally methylated regions—IGF1R, NHP2L1, L3MBTL, and ZDBF2—that overlapped with the differentially methylated regions found in BWS patients with multi-locus imprinting disturbance (MLID). This finding suggests that the BWS phenotype can result from MLID without detectable methylation defects in the primarily disease-associated loci (11p15). Another patient manifested small but significant aberrant methylation in disease-associated loci at 11p near H19, possibly confirming the diagnosis in this patient.

Beckwith-Wiedemann Syndrome (BWS) Current Status of Diagnosis and Clinical Management: Summary of the First International Consensus Statement

Beckwith-Wiedemann syndrome (BWS) belongs to the group of imprinting disorders and is characterized by variable clinical features, including overgrowth, macroglossia, abdominal wall defect, neonatal hypoglycemia, body asymmetry and an increased risk for embryonal tumors. In the majority of cases, molecular alterations of the Imprinting Center (IC) regions in the chromosomal region 11p15.5 can be detected, and a correlation of single clinical features with specific genomic and epigenetic changes is obvious. Therefore, the detailed molecular diagnosis is a prerequisite for a precise prediction of the tumor risk and the tumor spectrum. Furthermore, it is the basis for a well-directed genetic counselling of the families. Despite a huge number of comprehensive studies based on a large number of cases, standardized diagnostic criteria and advices for therapeutic management were missing. In the following, the recently published first international consensus guidelines drafted by 41 experts in the field of BWS from 11 European countries and the USA are summarized. Patients support groups had been included as well. In total, 72 consented recommendations for clinical and molecular diagnosis as well as for the clinical management of BWS have been published. They refer both to patients with the classical BWS phenotype and to those with "atypical" phenotypes which are summarized as BWS spectrum (BWSp). A modified clinical scoring system is now suggested, which represents the basis to initiate molecular diagnostics. Therapeutic recommendations comprise the major clinical questions in BWS/BWSp, i. e. early monitoring of an increased tumor risk, treatment of the macroglossia and the abdominal wall defects, and therapeutic interventions for hypoglycemia. However, though there was a broad consensus on the majority of therapeutic interventions, discussions on tumor monitoring are foreseeable. Thus, prospective studies to evaluate the consensus guidelines and their use are planned.

Beckwith-Wiedemann Syndrome Review: A Guide for the Neonatal Nurse.

Beckwith-Wiedemann syndrome (BWS) is the most common pediatric overgrowth syndrome. Features characteristic of the BWS phenotype include both physical attributes, such as macroglossia, abdominal wall defects, gigantism, nevus flammeus, visceromegaly, and mid-face hypoplasia, as well as biochemical abnormalities such as hypoglycemia. It is essential for the neonatal nurse to be able to recognize BWS in the patient's early years of life because of the increased frequency of medical complications, malformations, and the increased risk of embryonic malignancies. This article focuses on the presentation of BWS as an aid to early detection.

Mutations of the Imprinted CDKN1C Gene as a Cause of the Overgrowth Beckwith-Wiedemann Syndrome: Clinical Spectrum and Functional Characterization.

Beckwith-Wiedemann syndrome (BWS) is an imprinting disorder associating macroglossia, abdominal wall defects, visceromegaly, and a high risk of childhood tumor. Molecular anomalies are mostly epigenetic; however, mutations of CDKN1C are implicated in 8% of cases, including both sporadic and familial forms. We aimed to describe the phenotype of BWS patients with CDKN1C mutations and develop a functional test for CDKN1C mutations. For each propositus, we sequenced the three exons and intron-exon boundaries of CDKN1C in patients presenting a BWS phenotype, including abdominal wall defects, without 11p15 methylation defects. We developed a functional test based on flow cytometry. We identified 37 mutations in 38 pedigrees (50 patients and seven fetuses). Analysis of parental samples when available showed that all mutations tested but one was inherited from the mother. The four missense mutations led to a less severe phenotype (lower frequency of exomphalos) than the other 33 mutations. The following four tumors occurred: one neuroblastoma, one ganglioneuroblastoma, one melanoma, and one acute lymphoid leukemia. Cases of BWS caused by CDKN1C mutations are not rare. CDKN1C sequencing should be performed for BWS patients presenting with abdominal wall defects or cleft palate without 11p15 methylation defects or body asymmetry, or in familial cases of BWS.

Meningocele in a congolese female with beckwith-wiedemann phenotype.

Beckwith-Wiedemann syndrome (BWS) is a rare congenital syndrome characterized by an overgrowth, macroglossia, exomphalos, and predisposition to embryonal tumors. Central nervous abnormalities associated with BWS are rare. We describe a one-day-old Congolese female who presented meningocele associated with BWS phenotype.

Molecular Findings in Beckwith–Wiedemann Syndrome

Our understanding of Beckwith-Wiedemann syndrome (BWS) has recently been enhanced by advances in its molecular characterization. These advances have further delineated intricate (epi)genetic regulation of the imprinted gene cluster on chromosome 11p15.5 and the role of these genes in normal growth and development. Studies of the molecular changes associated with the BWS phenotype have been instrumental in elucidating critical molecular elements in this imprinted region. This review will provide updated information on the multiple new regulatory elements that have been recently found to contribute to in cis or in trans control of imprinted gene expression in the chromosome 11p15.5 region and the clinical expression of the BWS phenotype.

Brain abnormalities in patients with Beckwith–Wiedemann syndrome

Beckwith-Wiedemann syndrome (BWS) is an overgrowth disorder with variability in clinical manifestations and molecular causes. In most cases, patients with BWS have normal development. Cases with developmental delay are usually attributed to neonatal hypoglycemia or chromosome abnormalities involving copy number variation for genes beyond the critical BWS region at 11p15.5. Brain abnormalities have not previously been recognized within the BWS phenotypic spectrum. We report on seven cases of BWS associated with posterior fossa abnormalities. Of these, two cases presented with Blake's pouch cyst, two with Dandy-Walker variant (DWV; hypoplasia of the inferior part of the vermis), one with Dandy-Walker malformation (DWM) and one with a complex of DWM, dysgenesis of the corpus callosum and brain stem abnormality. In all these cases, molecular findings involved the centromeric imprinted domain on chromosome locus 11p15.5, which includes imprinting center 2 (IC2) and the imprinted growth suppressor gene, CDKN1C. Three cases had loss of methylation at IC2, two had CDKN1C mutations, and one had loss of methylation at IC2 and a microdeletion. In one case no mutation/methylation abnormality was detected. These findings together with previously reported correlations suggest that genes in imprinted domain 2 at 11p15.5 are involved in normal midline development of several organs including the brain. Our data suggest that brain malformations may present as a finding within the BWS phenotype when the molecular etiology involves imprinted domain 2. Brain imaging may be useful in identifying such malformations in individuals with BWS and neurodevelopmental issues.

Maternal gametic transmission of translocations or inversions of human chromosome 11p15.5 results in regional DNA hypermethylation and downregulation of CDKN1C expression

Beckwith–Wiedemann syndrome (BWS) is an overgrowth syndrome associated with genetic or epigenetic alterations in one of two imprinted domains on chromosome 11p15.5. Rarely, chromosomal translocations or inversions of chromosome 11p15.5 are associated with BWS but the molecular pathophysiology in such cases is not understood. In our series of 3 translocation and 2 inversion patients with BWS, the chromosome 11p15.5 breakpoints map within the centromeric imprinted domain, 2. We hypothesized that either microdeletions/microduplications adjacent to the breakpoints could disrupt genomic sequences important for imprinted gene regulation. An alternate hypothesis was that epigenetic alterations of as yet unknown regulatory DNA sequences, result in the BWS phenotype. A high resolution Nimblegen custom microarray was designed representing all non-repetitive sequences in the telomeric 33 Mb of the short arm of human chromosome 11. For the BWS-associated chromosome 11p15.5 translocations and inversions, we found no evidence of microdeletions/microduplications. DNA methylation was also tested on this microarray using the HpaII tiny fragment enrichment by ligation-mediated PCR (HELP) assay. This high-resolution DNA methylation microarray analysis revealed a gain of DNA methylation in the translocation/inversion patients affecting the p-ter segment of chromosome 11p15, including both imprinted domains. BWS patients that inherited a maternal translocation or inversion also demonstrated reduced expression of the growth suppressing imprinted gene, CDKN1C in Domain 2. In summary, our data demonstrate that translocations and inversions involving imprinted domain 2 on chromosome 11p15.5, alter regional DNA methylation patterns and imprinted gene expression in cis, suggesting that these epigenetic alterations are generated by an alteration in “chromatin context”.

The KCNQ1OT1 imprinting control region and non-coding RNA: new properties derived from the study of Beckwith–Wiedemann syndrome and Silver–Russell syndrome cases

A cluster of imprinted genes at chromosome 11p15.5 is associated with the growth disorders, Silver–Russell syndrome (SRS) and Beckwith–Wiedemann syndrome (BWS). The cluster is divided into two domains with independent imprinting control regions (ICRs). We describe two maternal 11p15.5 microduplications with contrasting phenotypes. The first is an inverted and in cis duplication of the entire 11p15.5 cluster associated with the maintenance of genomic imprinting and with the SRS phenotype. The second is a 160 kb duplication also inverted and in cis, but resulting in the imprinting alteration of the centromeric domain. It includes the centromeric ICR (ICR2) and the most 5′ 20 kb of the non-coding KCNQ1OT1 gene. Its maternal transmission is associated with ICR2 hypomethylation and the BWS phenotype. By excluding epigenetic mosaicism, cell clones analysis indicated that the two closely located ICR2 sequences resulting from the 160 kb duplication carried discordant DNA methylation on the maternal chromosome and supported the hypothesis that the ICR2 sequence is not sufficient for establishing imprinted methylation and some other property, possibly orientation-dependent, is needed. Furthermore, the 1.2 Mb duplication demonstrated that all features are present for correct imprinting at ICR2 when this is duplicated and inverted within the entire cluster. In the individuals maternally inheriting the 160 kb duplication, ICR2 hypomethylation led to the expression of a truncated KCNQ1OT1 transcript and to down-regulation of CDKN1C. We demonstrated by chromatin RNA immunopurification that the KCNQ1OT1 RNA interacts with chromatin through its most 5′ 20 kb sequence, providing a mechanism likely mediating the silencing activity of this long non-coding RNA.

A novel microdeletion in the IGF2/H19 imprinting centre region defines a recurrent mutation mechanism in familial Beckwith–Wiedemann syndrome

The overgrowth disorder Beckwith–Wiedemann syndrome (BWS) is associated with dysregulation of imprinted genes at chromosome 11p15.5. The molecular defects are heterogeneous but most of the cases are associated with defective DNA methylation at either one of two Imprinting Control Regions (IC1 and IC2) or Uniparental paternal Disomy (UPD) at 11p15.5. In rare cases, the BWS phenotype has been found associated with maternal transmission of IC1 microdeletions. We describe a family with a novel 1.8kb deletion that is associated with hypermethylation at IC1. The mutation results from recombination between highly homologous sequences containing target sites for the zinc-finger protein CTCF (CTSs). This finding supports the hypothesis that the function of IC1 and the penetrance of the clinical phenotype depend on the spacing of the CTSs resulting from recombination in the mutant allele.

Neonatal hepatoblastoma in a newborn with severe phenotype of Beckwith–Wiedemann syndrome

Beckwith-Wiedemann syndrome is an overgrowth disorder characterized by neonatal macrosomia, abdominal wall defects, macroglossia, renal anomalies, organomegaly, hypoglycemia, and cancer predisposition. Hepatoblastoma is the second most frequent tumor and periodic serum alpha-fetoprotein (αFP) dosage is the cornerstone of the tumor surveillance for its early detection. In this report, we describe the outstanding case of a Beckwith-Wiedemann syndrome (BWS) newborn with severe phenotype and paternal chromosome 11 uniparental disomy (UPD11) associated with a high tumor risk. Based on the clinical picture and previous reports, a close monitoring of αFP was commenced. The marker was normal immediately after birth, but rapidly raised in 20 days, leading to the diagnosis of an extremely aggressive hepatoblastoma. The latter was successfully treated with pre-surgical reductive chemotherapy, gross total mass resection, and subsequent chemotherapy. Based on this observation, the tumor surveillance routinely suggested every 3 months should be more intense and with closer time intervals in newborns with severe BWS phenotype. We suggest monitoring neonatal αFP every 20 days in such cases.

Beckwith–Wiedemann syndrome and uniparental disomy 11p: fine mapping of the recombination breakpoints and evaluation of several techniques

Beckwith-Wiedemann syndrome (BWS) is a phenotypically and genotypically heterogeneous overgrowth syndrome characterized by somatic overgrowth, macroglossia and abdominal wall defects. Other usual findings are hemihyperplasia, embryonal tumours, adrenocortical cytomegaly, ear anomalies, visceromegaly, renal abnormalities, neonatal hypoglycaemia, cleft palate, polydactyly and a positive family history. BWS is a complex, multigenic disorder associated, in up to 90% of patients, with alteration in the expression or function of one or more genes in the 11p15.5 imprinted gene cluster. There are several molecular anomalies associated with BWS and the large proportion of cases, about 85%, is sporadic and karyotypically normal. One of the major categories of BWS molecular alteration (10-20% of cases) is represented by mosaic paternal uniparental disomy (pUPD), namely patients with two paternally derived copies of chromosome 11p15 and no maternal contribution for that. In these patients, in addition to the effects of IGF2 overexpression, a decreased level of the maternally expressed gene CDKN1C may contribute to the BWS phenotype. In this paper, we reviewed a series of nine patients with BWS because of pUPD using several methods with the aim to evaluate the percentage of mosaicism, the methylation status at both loci, the extension of the pUPD at the short arm and the breakpoints of recombination. Fine mapping of mitotic recombination breakpoints by single-nucleotide polymorphism-array in individuals with UPD and fine estimation of epigenetic defects will provide a basis for understanding the aetiology of BWS, allowing more accurate prognostic predictions and facilitating management and surveillance of individuals with this disorder.

Multilocus methylation analysis in a large cohort of 11p15-related foetal growth disorders (Russell Silver and Beckwith Wiedemann syndromes) reveals simultaneous loss of methylation at paternal and maternal imprinted loci

Genomic imprinting plays an important role in mammalian development. Loss of imprinting (LOI) through loss (LOM) or gain (GOM) of methylation is involved in many human disorders and cancers. The imprinted 11p15 region is crucial for the control of foetal growth and LOI at this locus is implicated in two clinically opposite disorders: Beckwith Wiedemann syndrome (BWS) with foetal overgrowth associated with an enhanced tumour risk and Russell-Silver syndrome (RSS) with intrauterine and postnatal growth restriction. So far, only a few studies have assessed multilocus LOM in human imprinting diseases. To investigate multilocus LOI syndrome, we studied the methylation status of five maternally and two paternally methylated loci in a large series (n = 167) of patients with 11p15-related foetal growth disorders. We found that 9.5% of RSS and 24% of BWS patients showed multilocus LOM at regions other than ICR1 and ICR2 11p15, respectively. Moreover, over two third of multilocus LOM RSS patients also had LOM at a second paternally methylated locus, DLK1/GTL2 IG-DMR. No additional clinical features due to LOM of other loci were found suggesting an (epi)dominant effect of the 11p15 LOM on the clinical phenotype for this series of patients. Surprisingly, four patients displayed LOM at both ICR1 and ICR2 11p15. Three of them had a RSS and one a BWS phenotype. Our results show for the first time that multilocus LOM can also concern RSS patients. Moreover, LOM can involve both paternally and maternally methylated loci in the same patient.

Different mechanisms cause imprinting defects at the IGF2/H19 locus in Beckwith-Wiedemann syndrome and Wilms' tumour

The parent of origin-dependent expression of the IGF2 and H19 genes is controlled by the imprinting centre 1 (IC1) consisting in a methylation-sensitive chromatin insulator. Deletions removing part of IC1 have been found in patients affected by the overgrowth- and tumour-associated Beckwith-Wiedemann syndrome (BWS). These mutations result in the hypermethylation of the remaining IC1 region, loss of IGF2/H19 imprinting and fully penetrant BWS phenotype when maternally transmitted. We now report that 12 additional cases with IC1 hypermethylation have a similar clinical phenotype but showed neither a detectable deletion nor other mutation in the local vicinity. Likewise, no IC1 deletion was detected in 40 sporadic non-syndromic Wilms' tumours. A detailed analysis of the BWS patients showed that the hypermethylation variably affected the IC1 region and was generally mosaic. We observed that all these cases were sporadic and in at least two families affected and unaffected members shared the same maternal IC1 allele but not the abnormal maternal chromosome epigenotype. Furthermore, the chromosome with the imprinting defect derived from either the maternal grandfather or maternal grandmother. Overall, these results indicate that methylation-imprinting defects at the IGF2-H19 locus can result from inherited mutations of the IC and have high recurrence risk or arise independently from the sequence context and generally not transmitted to the progeny. Despite these differences, the epigenetic abnormalities are usually present in the patients in the mosaic form and probably acquired by post-zygotic de novo methylation. Distinguishing between these two groups of cases is important for genetic counselling.

Mechanisms causing imprinting defects in familial Beckwith–Wiedemann syndrome with Wilms' tumour

The imprinted expression of the IGF2 and H19 genes is controlled by the Imprinting Centre 1 (IC1) at chromosome 11p15.5. This is a methylation-sensitive chromatin insulator that works by binding the zinc-finger protein CTCF in a parent-specific manner. Microdeletions abolishing some of the CTCF target sites (CTSs) of IC1 have been associated with the Beckwith-Wiedemann syndrome (BWS). However, the link between these mutations and the molecular and clinical phenotypes was debated. We have identified two novel families with IC1 deletions, in which individuals with the clinical features of the BWS are present in multiple generations. By analysing the methylation pattern at the IGF2-H19 locus together with the clinical phenotypes in the individuals with maternal and those with paternal transmission of five different deletions, we demonstrate that maternal transmission of 1.4-1.8 kb deletions in the IC1 region co-segregates with the hypermethylation of the residual CTSs and BWS phenotype with complete penetrance, whereas normal phenotype is observed upon paternal transmission. Although gene expression could not be assayed in all cases, the methylation detected at the IGF2 DMR2 and H19 promoter suggests that IC1 hypermethylation is consistently associated with biallelic activation of IGF2 and biallelic silencing of H19. Comparison of these deletions with a 2.2 kb one previously reported by another group indicates that the spacing of the CTSs on the deleted allele is critical for the gain of the abnormal methylation and penetrance of the clinical phenotype. Furthermore, we observe that the hypermethylation resulting from the deletions is always mosaic, suggesting that the epigenetic defect at the IGF2-H19 locus is established post-zygotically and may cause body asymmetry and heterogeneity of the clinical phenotype. Finally, the IC1 microdeletions are associated with a high incidence of Wilms' tumour, making their molecular diagnosis particularly important for genetic counselling and tumour surveillance at follow-up.