Improved molecular detection of mosaicism in Beckwith-Wiedemann Syndrome

Baker, Samuel White; Duffy, Kelly A.; Richards‐Yutz, Jennifer; Deardorff, Matthew A.; Kalish, Jennifer M.; Ganguly, Arupa

doi:10.1136/jmedgenet-2019-106498

Cited by 24 publications

(44 citation statements)

References 36 publications

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“…18 Despite use of this sensitive assay, our data indicate individuals who receive a prenatal diagnosis of BWS have significantly greater magnitude methylation level changes relative to patients diagnosed after birth. 18 In fact, just 1/21 (4.8%) patients who received a molecular diagnosis on the basis of methylation testing had a diagnostic methylation level change of <20% relative to normal 50% values. These results suggest that low level changes in methylation levels may be uncommon among patients diagnosed prenatally.…”

Section: Discussionmentioning

confidence: 66%

“…Measurement of methylation levels at IC1 and IC2 via allele-specific methylated multiplex quantitative real-time PCR (ASMM-qRT-PCR) and calculation of the proportion of cells with altered methylation levels were performed as previously described. 18,19 818 -BAKER ET AL.…”

Section: Measurement and Reporting Of Methylation Levels At Ic1 And Ic2mentioning

confidence: 99%

“…Sanger sequencing of CDKN1C was performed as previously described. 18 Variants identified in CDKN1C were annotated using the NM_000076.2 RefSeq transcript. Variant classification was performed in accordance with the GDL variant classification protocol, which is based on the ACMG recommended guidelines.…”

Section: Cdkn1c Variant Characterization Classification and Reportingmentioning

confidence: 99%

“…Methylation levels at IC1 and IC2 were measured in samples from 91 patients using an ASMM-qRT-PCR-based assay that can reliably detect methylation level differences of 3% relative to normal 50% values. 18,19 This sensitive assay enabled identification of methylation defects as well as measurement of the methylation defect magnitude, that is a methylation level of 20% at IC2 represents a methylation level difference of 30% relative to normal 50% values. The median magnitude of methylation level change identified in prenatal patients who received a molecular diagnosis was 38.4%, a value significantly greater than the mean methylation level difference of 20.0% observed in >900 patients who received a molecular diagnosis from postnatal testing (p < 0.001) (Figure 1).…”

Section: Methylation Findings In Prenatal Cases Of Bwsmentioning

confidence: 99%

“…the affected individuals were mosaic for methylation level defects in the tested tissue), methylation level changes of <20% relative to normal 50% levels were significantly less common among prenatal patients, with such mosaic methylation levels identified in only 1/21 (4.3%) prenatal patients (p = 0.0003). 18 Postnatal testing of blood was performed for 10/60 (16.7%) patients who received prenatal methylation testing of amniocytes.…”

Section: Methylation Findings In Prenatal Cases Of Bwsmentioning

confidence: 99%

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Prenatal molecular testing and diagnosis of Beckwith‐Wiedemann syndrome

et al. 2021

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Objective: The objective of this study was to describe molecular findings and phenotypic features among individuals referred for prenatal Beckwith-Wiedemann syndrome (BWS) testing.Methods: Molecular diagnostic testing was performed using a sensitive quantitative real-time PCR-based assay capable of detecting mosaic methylation to the level of 3% at IC1 and IC2. Sanger sequencing of CDKN1C was performed in cases with normal methylation.Results: Of the 94 patients tested, a molecular diagnosis was identified for 25.5% of cases; 70.9% of diagnosed cases had loss of methylation at IC2, 4.2% had gain of methylation at IC1, 12.5% had paternal uniparental isodisomy, and 12.5% had CDKN1C loss-of-function variants. Methylation level changes in prenatal cases were significantly greater than changes identified in cases tested after birth. Cases with a prenatal molecular diagnosis had a significantly greater number of BWS-associated phenotypic features. The presence of either macroglossia or placentomegaly was most predictive of a BWS diagnosis. Conclusion:Our results support the consensus statement advocating BWS molecular testing for all patients with one or more BWS-associated prenatal features and suggest that low-level mosaic methylation changes may be uncommon among prenatal BWS diagnoses. Key PointsWhat is already known about this topic? � Prenatal features and molecular findings associated with prenatal diagnosis of BWS have been previously described in multiple publications. What does this study add?� This study demonstrates the use of an assay with improved mosaic detection for BWS in a prenatal cohort of patients.� We report which prenatal features are predictive of a diagnosis in our cohort. | INTRODUCTIONEpigenetic and genetic alterations at chromosome 11p15.5 cause Beckwith-Wiedemann Syndrome (BWS [MIM: 130650]), a congenital overgrowth disorder associated with lateralized overgrowth, omphalocele, macroglossia, hyperinsulinism, and embryonal tumor risk. 1 Epimutations within the KCNQ1OT1:TSS (IC2) differentially methylated region (DMR) and H19/IGF2:IG (IC1) DMR imprinting control regions on chromosome 11p15.5 are the most common causes of BWS. 1 Methylation levels at IC1 and IC2 regulate

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Section: Discussionmentioning

confidence: 66%

Section: Measurement and Reporting Of Methylation Levels At Ic1 And Ic2mentioning

confidence: 99%

Section: Cdkn1c Variant Characterization Classification and Reportingmentioning

confidence: 99%

Section: Methylation Findings In Prenatal Cases Of Bwsmentioning

confidence: 99%

Section: Methylation Findings In Prenatal Cases Of Bwsmentioning

confidence: 99%

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Prenatal molecular testing and diagnosis of Beckwith‐Wiedemann syndrome

et al. 2021

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Familial Beckwith‐Wiedemann syndrome in a multigenerational family: Forty years of careful phenotyping

Best

Duffy

George

et al. 2022

American J of Med Genetics Pt A

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Beckwith‐Wiedemann Spectrum (BWSp) is an overgrowth and cancer predisposition disorder characterized by a wide spectrum of phenotypic manifestations including macroglossia, abdominal wall defects, neonatal hypoglycemia, and predisposition to embryonal tumors. In 1981, Best and Hoekstra reported four patients with BWSp in a single family which suggested autosomal dominant inheritance, but standard clinical testing for BWSp was not available during this time. Meticulous phenotyping of this family has occurred over the past 40 years of follow‐up with additional family members being identified and samples collected for genetic testing. Genetic testing revealed a pathogenic mutation in CDKN1C, consistent with the most common cause of familial BWSp. CDKN1C mutations account for just 5% of sporadic cases of BWSp. Here, we report the variable presentation of BWSp across the individuals affected by the CDKN1C mutation and other extended family members spanning multiple generations, all examined by the same physician. Additional phenotypes thought to be atypical in patients with BWSp were reported which included cardiac abnormalities. The incidence of tumors was documented in extended family members and included rhabdomyosarcoma, astrocytoma, and thyroid carcinoma, which have previously been reported in patients with BWSp. These observations suggest that in addition to the inheritance of the CDKN1C variant, there are modifying factors in this family driving the phenotypic spectrum observed. Alternative theories are suggested to explain the etiology of clinical variability including diffused mosaicism, anticipation, and the presence of additional variants tracking in the family. This study highlights the necessity of long‐term follow‐up in patients with BWSp and consideration of individual familial characteristics in the context of phenotype and/or (epi)genotype associations.

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

Schlaich,

Hubens,

Eggermann

2023

Molec Gen & Gen Med

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BackgroundBeckwith‐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.MethodsWe 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.ResultsMS‐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.ConclusionWe 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.

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Improved molecular detection of mosaicism in Beckwith-Wiedemann Syndrome

Cited by 24 publications

References 36 publications

Prenatal molecular testing and diagnosis of Beckwith‐Wiedemann syndrome

Prenatal molecular testing and diagnosis of Beckwith‐Wiedemann syndrome

Familial Beckwith‐Wiedemann syndrome in a multigenerational family: Forty years of careful phenotyping

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

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