18.03.12 BWS/SRS

Question 1

What is imprinting?

Answer 1

1. A developmental process which sometimes leads to the exclusive expression of specific genes from only one parent.
2. The gene is imprinted with the parent of origin (POI) during gametogenesis. Involves epigenetic modification of DNA e.g. methylation, histone modification, noncoding RNAs.
3. The genes in imprinted areas are expressed monoallelically depending on the parent of origin. This monoallelic expression can vary between organs, during developmental stages and disease.
4. Inheritance of both maternal and paternal genes is therefore required for normal development to proceed.

Question 2

Which imprinting locus is associated with BWS ans SRS?

Answer 2

Aberrant genomic imprinting of the 11p15 region, one of the major imprinting clusters, is important in both BWS (BWS; MIM# 130650) and RSS (SRS; MIM# 180860) syndromes.

Question 3

Briefly describe the 11p15 imprinting cluster.

Answer 3

This cluster is organized in two neighboring imprinted domains each controlled by its own imprinting centre, IC1 and IC2, which regulate the expression of imprinted genes in cis over large distances and show differential methylation of the parental alleles.

Question 4

Which genes are present in the 11p15 domain 1?

Answer 4

1. IGF2 - a fetal growth factor.
2. H19 - a biologically active non-translated mRNA that may function as a tumor suppressor.
3. IC1 – (also called H19DMR, Differentially Methylated Region) regulates IGF2 and H19. Found methylated on the paternal allele, causing H19 to expressed from maternal chromosome and IGF2 to be expressed from the paternal chromosome only.
4. CTCF - binds to unmethylated IC1 causing H19 to be expressed. H19 interacts with the enhancers of IGF2 located downstream of H19. By this mechanism, H19 is able to prevent the interaction of IGF2 and its enhancers, thus blocking IGF2 expression

Question 5

Which genes are present in the 11p15 domain 2?

Answer 5

1. KCNQ1 - voltage gated potassium channel subunit. Mutations cause heart diseases e.g. LongQT, Jerrett and Lange-Nielsen syndrome. Bi-allelic expression in heart.
2. KCNQ1OT1- a non-coding RNA with antisense transcription to KCNQ1. Spans intron 10-intron 11 of KCNQ1.
3. IC2 - (also called KvDMR) maternally methylated CpG-island and includes the promoter of KCNQ1OT1. IC2 regulate in cis the maternally expressed genes of domain 2
4. CDKN1C - cyclin dependent kinase inhibitor, part of the Cip gene family of cell cycle regulators. It acts by arresting the cell cycle in G1 by binding to G1 cyclin-CDK complexes.

Question 6

What is the incidence of Beckwith-Weidemann syndrome? How is this syndrome diagnosed?

Answer 6

Paediatric overgrowth disorder with estimated incidence of 1 in 13,700.

Variable phenotypic expression with >30 clinical features. Diagnosed if 3 major or 2 major and one minor feature is present

Question 7

Give 5 major clinical features of BWS.

Answer 7

1. Positive family history of BWS
2. Macrosomia (excessive birth weight) (traditionally defined as height and weight >97th %)
3. Anterior linear ear lobe creases/posterior helical ear pits
4. Macroglossia (large tongue)
5. Omphalocele (abdominal wall does not fuse)(also called exomphalos), umbilical hernia
6. Visceromegaly involving one or more intra-abdominal organs including liver, spleen, kidneys, adrenal glands, and pancreas
7. Embryonal tumor (e.g., Wilms tumor, hepatoblastoma, rhabdomyosarcoma) in childhood
8. Hemihyperplasia defined as asymmetric overgrowth of region(s) of the body
9. Adrenocortical cytomegaly
10. Renal abnormalities including structural abnormalities, nephromegaly, nephrocalcinosis
11. Cleft palate (rare)

Question 8

Give 3 minor clinical features of BWS.

Answer 8

1. Polyhydramnios
2. Prematurity
3. Neonatal hypoglycemia
4. Facial nevus flammeus (stork bite)
5. Hemangioma
6. Characteristic facies, including midfacial hypoplasia and infraorbital creases
7. Cardiomegaly/structural cardiac anomalies/rarely cardiomyopathy
8. Diastasis recti - where the right and left sides of the rectus abdominus spread apart at the body’s midline.
9. Advanced bone age
10. Monozygotic twinning. Monozygous twins with BWS are usually female and discordant; however, both male and female monozygous twins concordant for BWS have been reported, as well as monozygous male twins discordant for BWS

Question 9

What proportion of BWS cases are sporadic?

Answer 9

~85% of BWS cases are sporadic in nature, the remaining 15% demonstrate an autosomal dominant familial inheritance

Question 10

What are the causes of sporadic BWS?

Answer 10

1. IC2 hypomethylation 50-60% (mosaic): due to loss (complete or partial) of maternal methylation at IC2
2. Paternal UPD ~20%
3. CDKN1C point mutation 5-10%
4. IC1 hypermethylation 2-7%
5. Translocations/inversions (<1%) Duplications tend to be show paternal inheritance. Inversions and translocations involve the maternal allele and almost always disrupt KCNQ1
6. Other: other genomic loci may be involved. E.g. NALP2 (chr 19)- linked to rare forms of familial BWS, and ZFP57 (chr 6) shown to work in trans and modulate imprinting at IC2.

Question 11

What is the effect of paternal UPD 11p15.5?

Answer 11

BWS

Epigenetic alterations that involve hypermethylation of IC1 and hypomethylation of IC2 indicate paternal UPD.

All cases have UPD for a segment including 11p15.5

Majority of cases are mosaic for paternal disomy which is very often very difficult to diagnose.

Question 12

What is the effect of CDKN1C point mutations?

Answer 12

5-10% of sporadic BWS
40% of BWS cases with a positive family history –shows dominant transmission).

Some variation in symptoms presented with CDKN1C mutations (Polydactyly, genital abnormalities, extra nipple, and cleft palate are more frequently observed). PHLDA2 and SLC22A18 also show preferential maternal expression in the fetus - thought to be involved in dominant BWS in some way

Question 13

What is the effect of IC1 hypermethylation?

Answer 13

2-7% sporadic BWS (mosaic):

IC1 gain of maternal methylation causing H19-dependent IGF2 biallelic expression.

In many cases of BWS biallelic IGF2 expression is accompanied by monoallelic H19 expression (H19-independent biallelic expression) but its significance in BWS is not completely understood.

These cases show normal methylation and expression of H19 from the maternal allele with biallelic IGF2 expression.

Question 14

What are the testing strategies for BWS if there is no known family history and/or cleft palate of the condition?

Answer 14

1. MLPA or other test for methylation and copy number change at 11p15.5 and karyotype
2. CDKN1C testing for cases with no 11p15.5 structural abnormality detected or methylation abnormality identified, If positive = screen family
3. Karyotype of CNV +ve for 11p15.5 abnormality = likely heritable translcation, duplication, inversion = screen family
4. If gain of H19 methylation observed = BWS aetiology likely heritable = screen family.
5. If loss of KvDMR methylation = likely sporadic
6. If gain of H19 and loss of KvDMR methylation present = consistent with UPD. Confirm by microsatellite analysis (sporadic).

Question 15

What are the testing strategies for BWS if there is a known family history and/or cleft palate of the condition?

Answer 15

First line test is screening for CDKN1C mutations following by methylation analysis if negative.

Question 16

Describe MS-MLPA for use in BWS.

Answer 16

(MS-MLPA): robust method for detecting majority of epigenetic and genetic changes associated with BWS. It detects microdeletions/microduplications, alterations in gene dosage, and DNA methylation including UPD. However in case of very low level UPD11 it’s best to perform microsatellite analysis especially in hemihypertrophy cases.

Question 17

Give a differential diagnosis for BWS.

Answer 17

Other overgrowth syndromes

1) Simpson-Golabi-Behmel, Perlman
2) Costello
3) Sotos
4) Weaver syndromes.

Question 18

Give an overview of Silver-Russel syndrome and how it is diagnosed.

Answer 18

Genetically and clinically heterogeneous condition characterised by intrauterine and postnatal growth retardation, dysmorphic facial features (a characteristic small, triangular face, and body asymmetry (SRS, MIM 180860).

Frequency of SRS is currently unknown, but probably underdiagnosed due to broad range of features (prevalence according to GeneReviews=1 in 100,000).

Diagnosis of SRS if have 3 major criteria or 2 major and 1 minor:

Question 19

Give three major clinical features of SRS.

Answer 19

1. Intrauterine growth retardation/small for gestational age (<10th percentile)
2. Postnatal growth with height/length <3rd percentile
3. Normal head circumference (3rd-97th percentile)
4. Limb, body, and/or facial asymmetry

Question 20

Give three minor clinical features of SRS.

Answer 20

1. Short (arm) span with normal upper- to lower-segment ratio
2. Fifth finger clinodactyly
3. Triangular facies
4. Frontal bossing/prominent forehead

Question 21

Give three supportive clinical features of SRS.

Answer 21

1. Café au lait spots or skin pigmentary changes
2. Genitourinary anomalies (cryptorchidism, hypospadias)
3. Motor, speech, and/or cognitive delays
4. Feeding disorder
5. Hypoglycemia

Question 22

What are the two known genetic causes of SRS?

Answer 22

1. chromosome 11p15.5-related

2. chr 7-related:

Question 23

Describe 11p15.5-related SRS.

Answer 23

Chr 11p15.5-related SRS: associated with hypomethylation of IC1 causing biallelic H19 expression and biallelic silencing of IGF2 resulting in growth restriction.

Question 24

What are the different mechanisms of 11p15.5-related SRS?

Answer 24

1) IC1 hypomethylation 30-50%: most common cause, loss of methylation on paternal chr 11p15.
2) maternal duplication of 11p15 region 1-2%
3) matUPD11: mosaic maternal uniparental disomy – single cases reported

In most cases both IGF2 and H19 are hypomethylated because IC1 regulates methylation of both genes. A small no. SRS patients have selective hypomethylation of only H19 or only IGF2.

Recently, deletions of the shared IGF2/H19 enhancer region have been reported. Such deletions could explain the selective hypomethylation of IGF2 as reported by Gronskov et al. (~1%)

As hypomethylation at paternal IC1 is a postzygotic event, most SRS patients are mosaic for abnormal methylation patterns.

A maternally inherited duplication of IC2 has been identified in one individual with SRS

Question 25

Describe chr 7-related SRS.

Answer 25

matUPD 7 account for 7-10% of SRS. Isodisomy and heterodisomy observed. Mosaicism found. Segmental UPD also reported.

Rare reports of chr. 7 abnormalities e.g. deletion of long arm of chr. 7.

Question 26

Other than 11p15.5 and chr 7, what other locus has been associated with SRS?

Answer 26

submicroscopic chromosomal imbalances 1-2%

A small no. have abnormal hypermethylation of IGF2R (the gene encoding the IGF2 receptor on chromosome 6q25-q27) with normal methylation of H19. SRS may result from reduction of IGF2R, which clears IGF2 from circulation.

Question 27

Give a differential diagnosis of SRS.

Answer 27

e.g. chromosomal abnormalities e.g. Deletions of Yq, 12p14.

DNA repair disorders e.g. Fanconi anaemia, Bloom syndrome – IUGR

3-M syndrome, SHORT syndrome, Temple syndrome, 15q26 microdeletion syndrome: see pre- and postnatal growth retardation

Question 28

Describe the genetic testing strategy for SRS.

Answer 28

1) Test methylation and CNV at 11p15.5 with MS-MLPA

2) If aberrant;
a) ICR1 hypomethylation (~40%) MS-assay for further imprinted loci - abberrant methylation at additional loci (75)
b) UPD(11p15.5)mat - confirm by microsatellite analysis
c) 11p15.5 dup (1-2%) - molecular karyotyping (FISH, MLPA) for confirmation

3) If normal methylation - test for UPD(7) mat - confirm by MSa. If negative - molecular karyotyping for submicroscopic dup/dels

Question 29

What proportion of SRS patients will not have a detectable genetic abnormality?

Answer 29

48%

Question 30

Describe the role of multi-locus methylation defects in SRS and BWS patients .

Answer 30

Recently significant number of SRS and BWS patients has also been reported with aberrant methylation at multiple imprinted loci (multilocus methylation defects; MLMD).

These patients demonstrated not only the disease-specific IC1 and IC2 hypomethylation but also other imprinting domains in the genome showed an abnormal methylation pattern.

About 9.5% of RSS patients with LOM (Loss Of Methylation) at IC1 11p15 has other imprinted loci and about

22% of BWS patients with ICR2 11p15 LOM also has LOM at least at one other locus.

No significant differences in clinical features were observed between RSS and BWS with multilocus and monolocus LOM patients.