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Hepatoblastoma in a child with a paternally-inherited ABCC8 mutation and mosaic paternal uniparental disomy 11p causing focal congenital hyperinsulinism
European Journal of Medical Genetics 56 (2013) 114e117
Contents lists available at SciVerse ScienceDirect
European Journal of Medical Genetics journal homepage: http://www.elsevier.com/locate/ejmg
Short clinical report
Hepatoblastoma in a child with a paternally-inherited ABCC8 mutation and mosaic paternal uniparental disomy 11p causing focal congenital hyperinsulinism Elizabeth A. Calton a, c, *, I. Karen Temple c, d, Deborah J.G. Mackay c, d, Margaret Lever d, Sian Ellard e, Sarah E. Flanagan e, Justin H. Davies b, Khalid Hussain f, Juliet C. Gray a, c a
Department of Paediatric Oncology, University Hospital Southampton Foundation Trust, Southampton, UK Department of Paediatric Endocrinology, University Hospital Southampton Foundation Trust, Southampton, UK Faculty of Medicine, University of Southampton, Southampton, UK d Wessex Regional Genetics Service, Salisbury Hospital NHS Foundation Trust, Salisbury, UK e Institute of Biomedical and Clinical Science, Peninsula Medical School, University of Exeter, Exeter, UK f Great Ormond Street Hospital for Children, London, UK b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 4 July 2012 Accepted 9 December 2012 Available online 20 December 2012
Hepatoblastoma is a tumour of early childhood occurring in association with genetic syndromes including BeckwitheWiedemann Syndrome (BWS) which results from dominance of paternallyinherited genes on chromosome 11p15. We report a child without clinical BWS, neonatally diagnosed with focal congenital hyperinsulinism resulting from a paternally-inherited recessively-acting mutation of ABCC8 and pancreatic paternal uniparental disomy (UPD) for chromosome 11p15, who subsequently developed hepatoblastoma. Genetic testing showed UPD 11p15 in the pancreas and liver but not systemically, allowing the expression of mutated ABCC8 in both tissues. Infants with large or multifocal forms of focal congenital hyperinsulinism may be at risk of BWS-like tumours due to mosaic UPD despite negative whole-blood and buccal DNA testing and tumour surveillance should be considered for this minority. Ó 2013 Elsevier Masson SAS. All rights reserved.
Keywords: Congenital hyperinsulinaemic hypoglycaemia Hepatoblastoma BeckwitheWiedemann syndrome Uniparental disomy Chromosome 11p
1. Introduction Abnormalities of the growth-regulatory region of Chromosome 11p15 result in a variety of overgrowth (and, less commonly, undergrowth) syndromes, classically BeckwitheWiedemann Syndrome (BWS, OMIM #130650) [1]. This phenotypically-varied disorder presents with organomegaly, abdominal wall defects, ear and facial dysmorphism and neonatal hypoglycaemia, which may be persistent [2,3]. BWS cases exhibit a variety of abnormalities relating to chromosome 11p, usually uniparental disomy (UPD) for paternally-derived 11p15.3-5 (20%) or a failure of genomic imprinting at one of two sites (ICR1 and ICR2) in this region (70%), resulting in expression of only the paternally-derived genes [2].
* Corresponding author. Dept of Paediatric Oncology, Mailpoint 43, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, UK. Tel.: þ44 7813 184494; fax: þ44 23 8079 4962. E-mail address: [email protected] (E.A. Calton). 1769-7212/$ e see front matter Ó 2013 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.ejmg.2012.12.001
Otherwise rare tumours, most commonly nephroblastoma (OMIM #194070) and hepatoblastoma (OMIM #114550), affect up to 24% of BWS patients with UPD for 11p15 and regular surveillance with abdominal ultrasound and tumour markers is usually performed during early childhood although there remains minimal evidence for its efficacy [2,4,5]. Tumour predisposition in BWS is thought to arise from over-expression of the paternally-derived growth promoter IGF2 or under-expression of the tumour suppressors CDKN1C and H19 which are maternally derived. However, studies of hepatoblastoma in BWS have failed to demonstrate the predicted down-regulation of the tumour suppressors nor increased expression of IGF2, in contrast to BWS-associated nephroblastoma [6,7]. The precise genetic trigger for hepatoblastoma in this syndrome therefore remains unknown. 11p15 defects also cause abnormalities of glucose regulation, including neonatal diabetes (OMIM #616176) and congenital hyperinsulinaemic hypoglycaemia (CHI, OMIM #256450) [8,9]. CHI presents with persistent and profound hypoglycaemia, often associated with macrosomia and hepatomegaly but usually without other syndromic features [10]. The most common forms of
E.A. Calton et al. / European Journal of Medical Genetics 56 (2013) 114e117
115
Fig. 1. 18F-DOPA PET scan showing lesion in head of pancreas consistent with focal congenital hyperinsulinism.
CHI, affecting 50e60% of patients, are those affecting the two subunits of the membrane KATP channel, both coded for by genes sited on chromosome 11p15.1: KCNJ11 encodes the KIR6.2 potassium channel pore, whilst ABCC8 produces the SUR1 ATP binding cassette which controls channel opening and functions as a sulphonylurea receptor [11]. Most mutations prevent channel assembly and trafficking to the membrane; a minority cause failure of channel opening [11,12]. Mutations are usually recessively acting with homozygotes having diffuse abnormality of pancreatic b-cells. However, a histologically-distinct focal form may also be present where paternally-inherited mutations are expressed phenotypically due to uniparental disomy for the relevant loci on chromosome 11p15 [8,13,14]. Focal forms usually arise from a single genetic event late in embryonic development, producing an isolated UPD within a single small area of the pancreas, but UPD can also occur earlier leading to multiple or larger lesions [14]. Both diffuse and focal CHI require urgent medical or surgical management to prevent long-term hypoglycaemic damage, particularly to the central nervous system [12]. Rapid genetic testing for KATP channel mutations identifies cases with paternally inherited heterozygous mutations [15]. PET-CT scanning using 18F-dopa confirms the presence of a focal lesion and localises it prior to surgical removal [16]. For focal cases where the whole lesion is surgically resected, complications are limited to neurological deficits resulting from early hypoglycaemia [8]. There is no known association of either isolated CHI or its treatment with hepatoblastoma or other cancers.
caesarean section. Hepatosplenomegaly and a four-vessel umbilical cord were noted, without hemihypertrophy, nephromegaly, or other dysmorphism. Following delivery, profound hypoglycaemia (0.4 mmol/L) required glucose infusion up to 20 mg/kg/min; insulin was 57.3 mU/l and a diagnosis of congenital hyperinsulinaemic hypoglycaemia was made. 18F-DOPA-PET showed a 7 6 6 mm lesion within the pancreatic head (Fig. 1), consistent with focal CHI, and after failure of medical management he underwent curative surgical resection. At 20 months, the child presented with abdominal discomfort and intermittent fever. Both right and left upper abdominal quadrants were distended but jaundice was absent. Initial investigations showed microcytic anaemia (Hb 71 g/dL; MCV 64 fL) and marked neutrophilia (WCC 49.0$109/L; Neutrophils 34.2$109/L). Liver synthetic function was reduced (albumin 21 g/L and INR 1.7) although glucose and liver enzymes remained normal. Sepsis was presumed but blood, urine and CSF cultures were negative. CT imaging revealed a large hepatic mass (PRETEXT Stage 4 [17]) associated with a subcapsular haematoma. Multiple pulmonary nodules were also present. Serum alpha-fetoprotein was grossly elevated (328,000 u/l); biopsy confirmed metastatic embryonaltype hepatoblastoma with small foci histologically similar to hepatocellular carcinoma. Treatment with multi-agent chemotherapy was followed by complete resection of residual tumour after four cycles. Unfortunately, further lung metastases developed which were not controlled with additional chemotherapy. DNA was extracted from peripheral blood leucocytes, mouthbrush samples and lung biopsies by conventional methods. DNA from wax embedded resected pancreatic and hepatoblastoma samples was extracted using the Qiamp DNA Minikit (Qiagen Ltd, Crawley UK). Mutational analysis of KCNJ11 and ABCC8 genes (NM_000525.3 and NM_000352.2, incorporating the alternatively spliced residue in exon 17 (L78224)) was performed by Sanger sequencing according to standard methods. Methylation-specific
2. Clinical report A male infant was born to non-consanguineous Caucasian parents at 39 þ 5 weeks following an uncomplicated spontaneous pregnancy. Unexpected macrosomia (weight 5095 g, head circumference 38.4 cm; both >99th centile) led to an emergency
Table 1 Analysis of microsatellite inheritance in blood and pancreatic DNA from the proband (neonatal). Microsatellite
Cytoband
Position (Mb from p terminus)
Mother
Father
Proband (blood)
Interpretation
Proband (pancreas)
Interpretation
D11S2071 D11S2347 D11S419 D11S4160 ABCC8 D11S902
p15.5 P15.4 p15.2 p15.1 p15.1 p15.1
0.95 3.84 15.8 16.9 17.4e17.5 17.4
1.1 1.2 1.2 1.2
2.3 1.1 1.1 1.2
1.3 2.1 2.1 1.2
Biparental Excludes paternal UPD Excludes paternal UPD Uninformative
3 1 1 1
No No No No
1.2
3.4
2.3
Biparental
3
No maternal allele
maternal allele maternal allele maternal allele (presumed) maternal allele
Biparental: proband shows unambiguous inheritance of one allele from each parent. Excludes maternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of paternal origin. Excludes paternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of maternal origin. No maternal allele: there is no evidence of inheritance of a maternal allele in the proband; this is consistent with either paternal isodisomy or a deletion of the maternal allele.
116
E.A. Calton et al. / European Journal of Medical Genetics 56 (2013) 114e117
Table 2 Analysis of microsatellite inheritance in blood and hepatic DNA from the proband (2nd presentation). Microsatellite
Cytoband
Position (Mb from p terminus)
Mother
Father
Proband (blood)
Interpretation
Proband (liver)
Interpretation
D11S2071 D11S1477 D11S922 D11S4088 D11S1349 ABCC8 D11S4138 D11S904 D11S4137 Centromere D11S913 D11S937 D11S4176
p15.5 p15.5 p15.5 p15.5 p15.3 p15.1 p15.1 p14.2 p11.2
0.95 1.44 1.56 2.71 11.71 17.4e17.5 17.72 26.64 45.56
1.1 2.3 1.2 2.3 2.3
2.3 1.4 3.4 1.4 1.2
1.3 2.4 1.4 1.3 1.2
Biparental Biparental Biparental Biparental Excludes maternal UPD
3 4 4 1 1
No No No No No
1.3 1.3 3.3
2.3 2.3 1.2
1.2 1.3 1.3
Biparental Excludes maternal UPD Biparental
2 (2) 3 (3) 1 (1)
No maternal allele No maternal allele No maternal allele
q13.1 q14.2 q12
65.69 77.58 93.76
3.3 2.3 2.3
1.2 1.2 1.3
2.3 2.3 1.3
Biparental Excludes paternal UPD Excludes maternal UPD
2.3 2.3 1.3
Biparental Excludes paternal UPD Excludes maternal UPD
(3) (4) (4) (1) (1)
maternal maternal maternal maternal maternal
allele allele allele allele allele
Biparental: proband shows unambiguous inheritance of one allele from each parent. Excludes maternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of paternal origin. Excludes paternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of maternal origin. No maternal allele: there is no evidence of inheritance of a maternal allele in the proband; this is consistent with either paternal isodisomy or a deletion of the maternal allele.
multiplex ligation-dependent probe amplification (MS-MLPA) of chromosome 11p15 was performed using MRC Holland kit ME030B2 according to the manufacturer’s instructions, and analysed using Genemarker software (SoftGenetics, Pennsylvania). Microsatellite repeats between 11p11.2 and 11p15.5, and on 11q, were interrogated by standard methods (primer sequences and conditions available upon request). Neonatally, mutation analysis of DNA from peripheral blood leukocytes showed heterozygosity for a paternally inherited frameshift mutation (c.3512delT; p.L1171fs), in the ABCC8 gene sited on chromosome 11p15.1. Blood leukocyte DNA showed no abnormality of methylation at either ICR1 (H19-IGF2 imprinting centre) or ICR2 (KCNQ1OT1/KCNQ1 imprinting centre) or UPD of 11p15; systemic BeckwitheWiedemann syndrome was therefore not suspected. However, microsatellite analysis demonstrated maternal loss of heterozygosity (LOH) at 11p15.1 in the pancreatic lesion (Table 1). MS-MLPA confirmed paternal UPD as the cause of this result. Following re-presentation, further microsatellite and MLPA analysis showed paternal UPD for 11p11.2-15.5 in biopsied hepatoblastoma and lung metastasis, reflecting the previous findings in the pancreas. Repeat leucocyte and buccal DNA from the child and both parents confirmed normal results at 11p15, demonstrating a mosaic genotype (Table 2). A later sample of mixed tumour and normal liver showed no LOH at 11p15 suggesting intra-hepatic mosaicism.
head) and may have arisen from several confluent foci reflecting a rare causative event early in embryogenesis. Whilst our results cannot confirm that the pancreatic and hepatic UPD have a common origin, this remains the most likely explanation for their combined presence. Children with multiple somatic foci of UPD at 11p15 may therefore have a similar vulnerability to nephroblastoma or hepatoblastoma to those with BWS despite a lack of clinical manifestations of the syndrome. Testing of systemic samples such as blood and buccal cells may not reveal mosaic abnormality and be falsely reassuring in this minority. 4. Summary It is not current clinical practice to follow up children presenting with apparently isolated focal congenital hyperinsulinism, even those with very large lesions as in this case, for early childhood cancer. For those with atypical focal lesions, potentially caused by multiple foci of mosaic paternal uniparental disomy for chromosome 11p15, this case suggests that active monitoring during early life for BeckwitheWiedemann Syndrome associated tumours should be considered on the same basis as for infants with clinical feature of BWS. Conflict of interest The authors declare no conflicts of interest in respect of this work.
3. Discussion References Patients with mosaic UPD for 11p15 often express clinical signs of BeckwitheWiedemann Syndrome, in addition to persistent hyperinsulinism, if their isodisomy spans the imprinted genes on 11p15.5 controlling the somatic manifestations of this disorder [18]. However, recessively-acting mutations are known to lead to hyperinsulinism without clinically-evident BWS where abnormal tissue is confined to the pancreas [8]. These children test negative for BWS on blood samples and, as in this case, are not kept under surveillance for tumour development. In this case, whole blood and buccal DNA testing for BWS was negative but paternal uniparental disomy for 11p15 was present outside the pancreas in at least one other organ derived from the embryonic foregut. Although the original pancreatic lesion appeared isolated, it was large (affecting the whole pancreatic
[1] R.H. Scott, J. Douglas, L. Baskcomb, N. Huxter, K. Barker, S. Hanks, A. Craft, M. Gerrard, J.A. Kohler, G.A. Levitt, S. Picton, B. Pizer, M.D. Ronghe, D. Williams, J.A. Cook, P. Pujol, E.R. Maher, J.M. Birch, C.A. Stiller, K. Pritchard-Jones, N. Rahman, Constitutional 11p15 abnormalities, including heritable imprinting center mutations, cause nonsyndromic Wilms tumor, Nat. Genet. 40 (2008) 1329e1334. [2] T.Y. Tan, D.J. Amor, Tumour surveillance in BeckwitheWiedemann syndrome and hemihyperplasia: a critical review of the evidence and suggested guidelines for local practice, J. Paediatr. Child. Health 42 (2006) 486e490. [3] K. Hussain, K.E. Cosgrove, R.M. Shepherd, A. Luharia, V.V. Smith, S. Kassem, J.W. Gregory, A. Sivaprasadarao, H.T. Christesen, B.B. Jacobsen, K. Brusgaard, B. Glaser, E.A. Maher, K.J. Lindley, P. Hindmarsh, M. Dattani, M.J. Dunne, Hyperinsulinemic hypoglycemia in BeckwitheWiedemann syndrome due to defects in the function of pancreatic beta-cell adenosine triphosphate-sensitive potassium channels, J. Clin. Endocrinol. Metab. 90 (2005) 4376e4382. [4] A. Mussa, G.B. Ferrero, B. Ceoloni, E. Basso, N. Chiesa, A. De Crescenzo, E. Pepe, M. Silengo, L. de Sanctis, Neonatal hepatoblastoma in a newborn with severe
E.A. Calton et al. / European Journal of Medical Genetics 56 (2013) 114e117
[5]
[6]
[7]
[8] [9]
[10]
[11] [12]
phenotype of BeckwitheWiedemann syndrome, Eur. J. Pediatr. 170 (2011) 1407e1411. W.N. Cooper, A. Luharia, G.A. Evans, H. Raza, A.C. Haire, R. Grundy, S.C. Bowdin, A. Riccio, G. Sebastio, J. Bliek, P.N. Schofield, W. Reik, F. Macdonald, E.R. Maher, Molecular subtypes and phenotypic expression of BeckwitheWiedemann syndrome, Eur. J. Hum. Genet. 13 (2005) 1025e1032. J.M. Schnater, S.E. Kohler, W.H. Lamers, D. von Schweinitz, D.C. Aronson, Where do we stand with hepatoblastoma? A review, Cancer 98 (2003) 668e678. M. Steenman, A. Westerveld, M. Mannens, Genetics of BeckwitheWiedemann syndrome-associated tumors: common genetic pathways, Genes Chromosomes Cancer 28 (2000) 1e13. R.R. Kapoor, S.E. Flanagan, C. James, J. Shield, S. Ellard, K. Hussain, Hyperinsulinaemic hypoglycaemia, Arch. Dis. Child. 94 (2009) 450e457. E.L. Edghill, S.E. Flanagan, S. Ellard, Permanent neonatal diabetes due to activating mutations in ABCC8 and KCNJ11, Rev. Endocr. Metab. Disord. 11 (2010) 193e198. J.B. Arnoux, P. de Lonlay, M.J. Ribeiro, K. Hussain, O. Blankenstein, K. Mohnike, V. Valayannopoulos, J.J. Robert, J. Rahier, C. Sempoux, C. Bellanne, V. Verkarre, Y. Aigrain, F. Jaubert, F. Brunelle, C. Nihoul-Fekete, Congenital hyperinsulinism, Early Hum. Dev. 86 (2010) 287e294. C. James, R.R. Kapoor, D. Ismail, K. Hussain, The genetic basis of congenital hyperinsulinism, J. Med. Genet. 46 (2009) 289e299. I. Giurgea, C. Bellanne-Chantelot, M. Ribeiro, L. Hubert, C. Sempoux, J.J. Robert, O. Blankenstein, K. Hussain, F. Brunelle, C. Nihoul-Fekete, J. Rahier, F. Jaubert,
[13]
[14]
[15]
[16]
[17]
[18]
117
P. de Lonlay, Molecular mechanisms of neonatal hyperinsulinism, Horm. Res. 66 (2006) 289e296. V. Verkarre, J.C. Fournet, P. de Lonlay, M.S. Gross-Morand, M. Devillers, J. Rahier, F. Brunelle, J.J. Robert, C. Nihoul-Fekete, J.M. Saudubray, C. Junien, Paternal mutation of the sulfonylurea receptor (SUR1) gene and maternal loss of 11p15 imprinted genes lead to persistent hyperinsulinism in focal adenomatous hyperplasia, J. Clin. Invest. 102 (1998) 1286e1291. K. Hussain, S.E. Flanagan, V.V. Smith, M. Ashworth, M. Day, A. Pierro, S. Ellard, An ABCC8 gene mutation and mosaic uniparental isodisomy resulting in atypical diffuse congenital hyperinsulinism, Diabetes 57 (2008) 259e263. I. Banerjee, M. Skae, S.E. Flanagan, L. Rigby, L. Patel, M. Didi, J. Blair, S. Ehtisham, S. Ellard, K.E. Cosgrove, M.J. Dunne, P.E. Clayton, The contribution of rapid KATP channel gene mutation analysis to the clinical management of children with congenital hyperinsulinism, Eur. J. Endocrinol. 164 (2011) 733e740. K. Mohnike, O. Blankenstein, H. Minn, W. Mohnike, F. Fuchtner, T. Otonkoski, [18F]-DOPA positron emission tomography for preoperative localization in congenital hyperinsulinism, Horm. Res. 70 (2008) 65e72. D.C. Aronson, Predictive value of the pretreatment extent of disease system in hepatoblastoma: results from the International Society of Pediatric Oncology Liver Tumor Study Group SIOPEL-1 study, J. Clin. Oncol. 23 (2005) 1245e1252. A.C. Smith, C. Shuman, D. Chitayat, L. Steele, P.N. Ray, J. Bourgeois, R. Weksberg, Severe presentation of BeckwitheWiedemann syndrome associated with high levels of constitutional paternal uniparental disomy for chromosome 11p15, Am. J. Med. Genet. A 143A (2007) 3010e3015.
Contents lists available at SciVerse ScienceDirect
European Journal of Medical Genetics journal homepage: http://www.elsevier.com/locate/ejmg
Short clinical report
Hepatoblastoma in a child with a paternally-inherited ABCC8 mutation and mosaic paternal uniparental disomy 11p causing focal congenital hyperinsulinism Elizabeth A. Calton a, c, *, I. Karen Temple c, d, Deborah J.G. Mackay c, d, Margaret Lever d, Sian Ellard e, Sarah E. Flanagan e, Justin H. Davies b, Khalid Hussain f, Juliet C. Gray a, c a
Department of Paediatric Oncology, University Hospital Southampton Foundation Trust, Southampton, UK Department of Paediatric Endocrinology, University Hospital Southampton Foundation Trust, Southampton, UK Faculty of Medicine, University of Southampton, Southampton, UK d Wessex Regional Genetics Service, Salisbury Hospital NHS Foundation Trust, Salisbury, UK e Institute of Biomedical and Clinical Science, Peninsula Medical School, University of Exeter, Exeter, UK f Great Ormond Street Hospital for Children, London, UK b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 4 July 2012 Accepted 9 December 2012 Available online 20 December 2012
Hepatoblastoma is a tumour of early childhood occurring in association with genetic syndromes including BeckwitheWiedemann Syndrome (BWS) which results from dominance of paternallyinherited genes on chromosome 11p15. We report a child without clinical BWS, neonatally diagnosed with focal congenital hyperinsulinism resulting from a paternally-inherited recessively-acting mutation of ABCC8 and pancreatic paternal uniparental disomy (UPD) for chromosome 11p15, who subsequently developed hepatoblastoma. Genetic testing showed UPD 11p15 in the pancreas and liver but not systemically, allowing the expression of mutated ABCC8 in both tissues. Infants with large or multifocal forms of focal congenital hyperinsulinism may be at risk of BWS-like tumours due to mosaic UPD despite negative whole-blood and buccal DNA testing and tumour surveillance should be considered for this minority. Ó 2013 Elsevier Masson SAS. All rights reserved.
Keywords: Congenital hyperinsulinaemic hypoglycaemia Hepatoblastoma BeckwitheWiedemann syndrome Uniparental disomy Chromosome 11p
1. Introduction Abnormalities of the growth-regulatory region of Chromosome 11p15 result in a variety of overgrowth (and, less commonly, undergrowth) syndromes, classically BeckwitheWiedemann Syndrome (BWS, OMIM #130650) [1]. This phenotypically-varied disorder presents with organomegaly, abdominal wall defects, ear and facial dysmorphism and neonatal hypoglycaemia, which may be persistent [2,3]. BWS cases exhibit a variety of abnormalities relating to chromosome 11p, usually uniparental disomy (UPD) for paternally-derived 11p15.3-5 (20%) or a failure of genomic imprinting at one of two sites (ICR1 and ICR2) in this region (70%), resulting in expression of only the paternally-derived genes [2].
* Corresponding author. Dept of Paediatric Oncology, Mailpoint 43, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, UK. Tel.: þ44 7813 184494; fax: þ44 23 8079 4962. E-mail address: [email protected] (E.A. Calton). 1769-7212/$ e see front matter Ó 2013 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.ejmg.2012.12.001
Otherwise rare tumours, most commonly nephroblastoma (OMIM #194070) and hepatoblastoma (OMIM #114550), affect up to 24% of BWS patients with UPD for 11p15 and regular surveillance with abdominal ultrasound and tumour markers is usually performed during early childhood although there remains minimal evidence for its efficacy [2,4,5]. Tumour predisposition in BWS is thought to arise from over-expression of the paternally-derived growth promoter IGF2 or under-expression of the tumour suppressors CDKN1C and H19 which are maternally derived. However, studies of hepatoblastoma in BWS have failed to demonstrate the predicted down-regulation of the tumour suppressors nor increased expression of IGF2, in contrast to BWS-associated nephroblastoma [6,7]. The precise genetic trigger for hepatoblastoma in this syndrome therefore remains unknown. 11p15 defects also cause abnormalities of glucose regulation, including neonatal diabetes (OMIM #616176) and congenital hyperinsulinaemic hypoglycaemia (CHI, OMIM #256450) [8,9]. CHI presents with persistent and profound hypoglycaemia, often associated with macrosomia and hepatomegaly but usually without other syndromic features [10]. The most common forms of
E.A. Calton et al. / European Journal of Medical Genetics 56 (2013) 114e117
115
Fig. 1. 18F-DOPA PET scan showing lesion in head of pancreas consistent with focal congenital hyperinsulinism.
CHI, affecting 50e60% of patients, are those affecting the two subunits of the membrane KATP channel, both coded for by genes sited on chromosome 11p15.1: KCNJ11 encodes the KIR6.2 potassium channel pore, whilst ABCC8 produces the SUR1 ATP binding cassette which controls channel opening and functions as a sulphonylurea receptor [11]. Most mutations prevent channel assembly and trafficking to the membrane; a minority cause failure of channel opening [11,12]. Mutations are usually recessively acting with homozygotes having diffuse abnormality of pancreatic b-cells. However, a histologically-distinct focal form may also be present where paternally-inherited mutations are expressed phenotypically due to uniparental disomy for the relevant loci on chromosome 11p15 [8,13,14]. Focal forms usually arise from a single genetic event late in embryonic development, producing an isolated UPD within a single small area of the pancreas, but UPD can also occur earlier leading to multiple or larger lesions [14]. Both diffuse and focal CHI require urgent medical or surgical management to prevent long-term hypoglycaemic damage, particularly to the central nervous system [12]. Rapid genetic testing for KATP channel mutations identifies cases with paternally inherited heterozygous mutations [15]. PET-CT scanning using 18F-dopa confirms the presence of a focal lesion and localises it prior to surgical removal [16]. For focal cases where the whole lesion is surgically resected, complications are limited to neurological deficits resulting from early hypoglycaemia [8]. There is no known association of either isolated CHI or its treatment with hepatoblastoma or other cancers.
caesarean section. Hepatosplenomegaly and a four-vessel umbilical cord were noted, without hemihypertrophy, nephromegaly, or other dysmorphism. Following delivery, profound hypoglycaemia (0.4 mmol/L) required glucose infusion up to 20 mg/kg/min; insulin was 57.3 mU/l and a diagnosis of congenital hyperinsulinaemic hypoglycaemia was made. 18F-DOPA-PET showed a 7 6 6 mm lesion within the pancreatic head (Fig. 1), consistent with focal CHI, and after failure of medical management he underwent curative surgical resection. At 20 months, the child presented with abdominal discomfort and intermittent fever. Both right and left upper abdominal quadrants were distended but jaundice was absent. Initial investigations showed microcytic anaemia (Hb 71 g/dL; MCV 64 fL) and marked neutrophilia (WCC 49.0$109/L; Neutrophils 34.2$109/L). Liver synthetic function was reduced (albumin 21 g/L and INR 1.7) although glucose and liver enzymes remained normal. Sepsis was presumed but blood, urine and CSF cultures were negative. CT imaging revealed a large hepatic mass (PRETEXT Stage 4 [17]) associated with a subcapsular haematoma. Multiple pulmonary nodules were also present. Serum alpha-fetoprotein was grossly elevated (328,000 u/l); biopsy confirmed metastatic embryonaltype hepatoblastoma with small foci histologically similar to hepatocellular carcinoma. Treatment with multi-agent chemotherapy was followed by complete resection of residual tumour after four cycles. Unfortunately, further lung metastases developed which were not controlled with additional chemotherapy. DNA was extracted from peripheral blood leucocytes, mouthbrush samples and lung biopsies by conventional methods. DNA from wax embedded resected pancreatic and hepatoblastoma samples was extracted using the Qiamp DNA Minikit (Qiagen Ltd, Crawley UK). Mutational analysis of KCNJ11 and ABCC8 genes (NM_000525.3 and NM_000352.2, incorporating the alternatively spliced residue in exon 17 (L78224)) was performed by Sanger sequencing according to standard methods. Methylation-specific
2. Clinical report A male infant was born to non-consanguineous Caucasian parents at 39 þ 5 weeks following an uncomplicated spontaneous pregnancy. Unexpected macrosomia (weight 5095 g, head circumference 38.4 cm; both >99th centile) led to an emergency
Table 1 Analysis of microsatellite inheritance in blood and pancreatic DNA from the proband (neonatal). Microsatellite
Cytoband
Position (Mb from p terminus)
Mother
Father
Proband (blood)
Interpretation
Proband (pancreas)
Interpretation
D11S2071 D11S2347 D11S419 D11S4160 ABCC8 D11S902
p15.5 P15.4 p15.2 p15.1 p15.1 p15.1
0.95 3.84 15.8 16.9 17.4e17.5 17.4
1.1 1.2 1.2 1.2
2.3 1.1 1.1 1.2
1.3 2.1 2.1 1.2
Biparental Excludes paternal UPD Excludes paternal UPD Uninformative
3 1 1 1
No No No No
1.2
3.4
2.3
Biparental
3
No maternal allele
maternal allele maternal allele maternal allele (presumed) maternal allele
Biparental: proband shows unambiguous inheritance of one allele from each parent. Excludes maternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of paternal origin. Excludes paternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of maternal origin. No maternal allele: there is no evidence of inheritance of a maternal allele in the proband; this is consistent with either paternal isodisomy or a deletion of the maternal allele.
116
E.A. Calton et al. / European Journal of Medical Genetics 56 (2013) 114e117
Table 2 Analysis of microsatellite inheritance in blood and hepatic DNA from the proband (2nd presentation). Microsatellite
Cytoband
Position (Mb from p terminus)
Mother
Father
Proband (blood)
Interpretation
Proband (liver)
Interpretation
D11S2071 D11S1477 D11S922 D11S4088 D11S1349 ABCC8 D11S4138 D11S904 D11S4137 Centromere D11S913 D11S937 D11S4176
p15.5 p15.5 p15.5 p15.5 p15.3 p15.1 p15.1 p14.2 p11.2
0.95 1.44 1.56 2.71 11.71 17.4e17.5 17.72 26.64 45.56
1.1 2.3 1.2 2.3 2.3
2.3 1.4 3.4 1.4 1.2
1.3 2.4 1.4 1.3 1.2
Biparental Biparental Biparental Biparental Excludes maternal UPD
3 4 4 1 1
No No No No No
1.3 1.3 3.3
2.3 2.3 1.2
1.2 1.3 1.3
Biparental Excludes maternal UPD Biparental
2 (2) 3 (3) 1 (1)
No maternal allele No maternal allele No maternal allele
q13.1 q14.2 q12
65.69 77.58 93.76
3.3 2.3 2.3
1.2 1.2 1.3
2.3 2.3 1.3
Biparental Excludes paternal UPD Excludes maternal UPD
2.3 2.3 1.3
Biparental Excludes paternal UPD Excludes maternal UPD
(3) (4) (4) (1) (1)
maternal maternal maternal maternal maternal
allele allele allele allele allele
Biparental: proband shows unambiguous inheritance of one allele from each parent. Excludes maternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of paternal origin. Excludes paternal UPD: the inheritance of alleles in the proband is not unambiguously biparental, but is consistent either with biparental inheritance or uniparental heterodisomy of maternal origin. No maternal allele: there is no evidence of inheritance of a maternal allele in the proband; this is consistent with either paternal isodisomy or a deletion of the maternal allele.
multiplex ligation-dependent probe amplification (MS-MLPA) of chromosome 11p15 was performed using MRC Holland kit ME030B2 according to the manufacturer’s instructions, and analysed using Genemarker software (SoftGenetics, Pennsylvania). Microsatellite repeats between 11p11.2 and 11p15.5, and on 11q, were interrogated by standard methods (primer sequences and conditions available upon request). Neonatally, mutation analysis of DNA from peripheral blood leukocytes showed heterozygosity for a paternally inherited frameshift mutation (c.3512delT; p.L1171fs), in the ABCC8 gene sited on chromosome 11p15.1. Blood leukocyte DNA showed no abnormality of methylation at either ICR1 (H19-IGF2 imprinting centre) or ICR2 (KCNQ1OT1/KCNQ1 imprinting centre) or UPD of 11p15; systemic BeckwitheWiedemann syndrome was therefore not suspected. However, microsatellite analysis demonstrated maternal loss of heterozygosity (LOH) at 11p15.1 in the pancreatic lesion (Table 1). MS-MLPA confirmed paternal UPD as the cause of this result. Following re-presentation, further microsatellite and MLPA analysis showed paternal UPD for 11p11.2-15.5 in biopsied hepatoblastoma and lung metastasis, reflecting the previous findings in the pancreas. Repeat leucocyte and buccal DNA from the child and both parents confirmed normal results at 11p15, demonstrating a mosaic genotype (Table 2). A later sample of mixed tumour and normal liver showed no LOH at 11p15 suggesting intra-hepatic mosaicism.
head) and may have arisen from several confluent foci reflecting a rare causative event early in embryogenesis. Whilst our results cannot confirm that the pancreatic and hepatic UPD have a common origin, this remains the most likely explanation for their combined presence. Children with multiple somatic foci of UPD at 11p15 may therefore have a similar vulnerability to nephroblastoma or hepatoblastoma to those with BWS despite a lack of clinical manifestations of the syndrome. Testing of systemic samples such as blood and buccal cells may not reveal mosaic abnormality and be falsely reassuring in this minority. 4. Summary It is not current clinical practice to follow up children presenting with apparently isolated focal congenital hyperinsulinism, even those with very large lesions as in this case, for early childhood cancer. For those with atypical focal lesions, potentially caused by multiple foci of mosaic paternal uniparental disomy for chromosome 11p15, this case suggests that active monitoring during early life for BeckwitheWiedemann Syndrome associated tumours should be considered on the same basis as for infants with clinical feature of BWS. Conflict of interest The authors declare no conflicts of interest in respect of this work.
3. Discussion References Patients with mosaic UPD for 11p15 often express clinical signs of BeckwitheWiedemann Syndrome, in addition to persistent hyperinsulinism, if their isodisomy spans the imprinted genes on 11p15.5 controlling the somatic manifestations of this disorder [18]. However, recessively-acting mutations are known to lead to hyperinsulinism without clinically-evident BWS where abnormal tissue is confined to the pancreas [8]. These children test negative for BWS on blood samples and, as in this case, are not kept under surveillance for tumour development. In this case, whole blood and buccal DNA testing for BWS was negative but paternal uniparental disomy for 11p15 was present outside the pancreas in at least one other organ derived from the embryonic foregut. Although the original pancreatic lesion appeared isolated, it was large (affecting the whole pancreatic
[1] R.H. Scott, J. Douglas, L. Baskcomb, N. Huxter, K. Barker, S. Hanks, A. Craft, M. Gerrard, J.A. Kohler, G.A. Levitt, S. Picton, B. Pizer, M.D. Ronghe, D. Williams, J.A. Cook, P. Pujol, E.R. Maher, J.M. Birch, C.A. Stiller, K. Pritchard-Jones, N. Rahman, Constitutional 11p15 abnormalities, including heritable imprinting center mutations, cause nonsyndromic Wilms tumor, Nat. Genet. 40 (2008) 1329e1334. [2] T.Y. Tan, D.J. Amor, Tumour surveillance in BeckwitheWiedemann syndrome and hemihyperplasia: a critical review of the evidence and suggested guidelines for local practice, J. Paediatr. Child. Health 42 (2006) 486e490. [3] K. Hussain, K.E. Cosgrove, R.M. Shepherd, A. Luharia, V.V. Smith, S. Kassem, J.W. Gregory, A. Sivaprasadarao, H.T. Christesen, B.B. Jacobsen, K. Brusgaard, B. Glaser, E.A. Maher, K.J. Lindley, P. Hindmarsh, M. Dattani, M.J. Dunne, Hyperinsulinemic hypoglycemia in BeckwitheWiedemann syndrome due to defects in the function of pancreatic beta-cell adenosine triphosphate-sensitive potassium channels, J. Clin. Endocrinol. Metab. 90 (2005) 4376e4382. [4] A. Mussa, G.B. Ferrero, B. Ceoloni, E. Basso, N. Chiesa, A. De Crescenzo, E. Pepe, M. Silengo, L. de Sanctis, Neonatal hepatoblastoma in a newborn with severe
E.A. Calton et al. / European Journal of Medical Genetics 56 (2013) 114e117
[5]
[6]
[7]
[8] [9]
[10]
[11] [12]
phenotype of BeckwitheWiedemann syndrome, Eur. J. Pediatr. 170 (2011) 1407e1411. W.N. Cooper, A. Luharia, G.A. Evans, H. Raza, A.C. Haire, R. Grundy, S.C. Bowdin, A. Riccio, G. Sebastio, J. Bliek, P.N. Schofield, W. Reik, F. Macdonald, E.R. Maher, Molecular subtypes and phenotypic expression of BeckwitheWiedemann syndrome, Eur. J. Hum. Genet. 13 (2005) 1025e1032. J.M. Schnater, S.E. Kohler, W.H. Lamers, D. von Schweinitz, D.C. Aronson, Where do we stand with hepatoblastoma? A review, Cancer 98 (2003) 668e678. M. Steenman, A. Westerveld, M. Mannens, Genetics of BeckwitheWiedemann syndrome-associated tumors: common genetic pathways, Genes Chromosomes Cancer 28 (2000) 1e13. R.R. Kapoor, S.E. Flanagan, C. James, J. Shield, S. Ellard, K. Hussain, Hyperinsulinaemic hypoglycaemia, Arch. Dis. Child. 94 (2009) 450e457. E.L. Edghill, S.E. Flanagan, S. Ellard, Permanent neonatal diabetes due to activating mutations in ABCC8 and KCNJ11, Rev. Endocr. Metab. Disord. 11 (2010) 193e198. J.B. Arnoux, P. de Lonlay, M.J. Ribeiro, K. Hussain, O. Blankenstein, K. Mohnike, V. Valayannopoulos, J.J. Robert, J. Rahier, C. Sempoux, C. Bellanne, V. Verkarre, Y. Aigrain, F. Jaubert, F. Brunelle, C. Nihoul-Fekete, Congenital hyperinsulinism, Early Hum. Dev. 86 (2010) 287e294. C. James, R.R. Kapoor, D. Ismail, K. Hussain, The genetic basis of congenital hyperinsulinism, J. Med. Genet. 46 (2009) 289e299. I. Giurgea, C. Bellanne-Chantelot, M. Ribeiro, L. Hubert, C. Sempoux, J.J. Robert, O. Blankenstein, K. Hussain, F. Brunelle, C. Nihoul-Fekete, J. Rahier, F. Jaubert,
[13]
[14]
[15]
[16]
[17]
[18]
117
P. de Lonlay, Molecular mechanisms of neonatal hyperinsulinism, Horm. Res. 66 (2006) 289e296. V. Verkarre, J.C. Fournet, P. de Lonlay, M.S. Gross-Morand, M. Devillers, J. Rahier, F. Brunelle, J.J. Robert, C. Nihoul-Fekete, J.M. Saudubray, C. Junien, Paternal mutation of the sulfonylurea receptor (SUR1) gene and maternal loss of 11p15 imprinted genes lead to persistent hyperinsulinism in focal adenomatous hyperplasia, J. Clin. Invest. 102 (1998) 1286e1291. K. Hussain, S.E. Flanagan, V.V. Smith, M. Ashworth, M. Day, A. Pierro, S. Ellard, An ABCC8 gene mutation and mosaic uniparental isodisomy resulting in atypical diffuse congenital hyperinsulinism, Diabetes 57 (2008) 259e263. I. Banerjee, M. Skae, S.E. Flanagan, L. Rigby, L. Patel, M. Didi, J. Blair, S. Ehtisham, S. Ellard, K.E. Cosgrove, M.J. Dunne, P.E. Clayton, The contribution of rapid KATP channel gene mutation analysis to the clinical management of children with congenital hyperinsulinism, Eur. J. Endocrinol. 164 (2011) 733e740. K. Mohnike, O. Blankenstein, H. Minn, W. Mohnike, F. Fuchtner, T. Otonkoski, [18F]-DOPA positron emission tomography for preoperative localization in congenital hyperinsulinism, Horm. Res. 70 (2008) 65e72. D.C. Aronson, Predictive value of the pretreatment extent of disease system in hepatoblastoma: results from the International Society of Pediatric Oncology Liver Tumor Study Group SIOPEL-1 study, J. Clin. Oncol. 23 (2005) 1245e1252. A.C. Smith, C. Shuman, D. Chitayat, L. Steele, P.N. Ray, J. Bourgeois, R. Weksberg, Severe presentation of BeckwitheWiedemann syndrome associated with high levels of constitutional paternal uniparental disomy for chromosome 11p15, Am. J. Med. Genet. A 143A (2007) 3010e3015.