JMed Genet 1990; 27: 729-737

Original articles

HEALTH EDUCATION LIBRARYBOOTH MEMORIAL MED-CALCENTERFLSHINGv3 N. Ya g

Alagille syndrome and deletion of 20pF Anad, J Burn, D Matthews, I Cross, B C C Davison, R Mueller, M Sands, D M Lillington,E Eastham

AbstractWe add five cases of 20p deletion to the 10 casesalready published. Four had craniofacial, vertebral,ocular, and cardiovascular features of Alagiliesyndrome, which adds weight to the assignment ofthis disorder to the short arm of chromosome 20.Included in our series is the first report of familialtransmission of a 20p deletion.

In 1%9, Alagille et all described a syndrome charac-terised by intrahepatic biliary ductal hypoplasia,peripheral pulmonary artery stenosis, a typical facieswith deep set eyes and a bossed forehead, posteriorembryotoxon of the eye, and vertebral defects. Avariety of less common features included renaldefects, short stature, and developmental delay.Several reports of affected subjects in more than onegeneration favoured an autosomal dominant trait withvariable expression.24 In 1986, Byrne et a15 reported20p deletion in an infant with features of Alagillesyndrome and drew attention to similar clinicalfeatures in two other reported cases with this deletion.In 1989, Schnittger et a16 reported a 19 year old girl

Division of Human Genetics, University of Newcastleupon Tyne, 19 Claremont Place, Newcastle upon TyneNE2 4AA.

F Anad, J Burn, I Cross

Department of Child Health, University of Newcastleupon Tyne.D Matthews, E Eastham

East Anglian Regional Genetics Service, Addenbrooke'sHospital, Cambridge.B C C Davison

Department of Clinical Genetics, The General Infirmary,Leeds.R Mueller

Cytogenetics Unit, Norfolk and Norwich Hospital,Norwich.M Sands, D M LillingtonCorrespondence to Dr Burn.

Received for publication 15 June 1990.Accepted for publication 26 June 1990.

with Alagille syndrome associated with interstitialdeletion of 20pl .23-p12. 1. More recently, Zhang eta17 and Legius et al8 reported a re-examination of thechromosomes of two boys with Alagille syndromewhich showed a deletion of 20p with the breakpointsat pll.13 and pl1.2, respectively.We report five further cases of 20p- to add to the

total of 10 already published. Details of four publishedcases-8 taken together with our cases confirm theassignment of Alagille syndrome to 20p and expandthe phenotype associated with deletion of this region.

Case reportsCASE 1

The proband was born at 39 weeks' gestation after anormal pregnancy. She was the first and only child ofa young, unrelated couple of white European origin.Her birthweight was 2350 g (less than the 10thcentile) and she was noted to be dysmorphic frombirth with a prominent forehead and deep set eyes.Difficulty in feeding and frequent vomiting led toinitial failure to thrive. She developed jaundice on herthird day of life and required phototherapy for fourdays with a maximum serum bilirubin level of 222mmol/l. Apart from this short lived neonatal jaundice,she had never developed liver problems and review at13 years showed normal liver function test with abilirubin of 6 mmol/l. Serum cholesterol was normalat 5*5 mmol/l but triglyceride levels were raised at 3-4mmol/l (normal range 0-1 to 2-1). Renal glycosuria,mild to moderate uraemia (blood urea of 16-9mmol/l), and a generalised aminoaciduria were notedin infancy; these resolved spontaneously. At the age of5 months she was found to be hypercalcaemic. Thisresponded to a low calcium diet for some time but shecontinued to fail to thrive. Intravenous pyelographyshowed a duplex right kidney with bilateral mild tomoderate vesicoureteric reflux. Renal biopsy per-formed at the age of 10 months showed sclerosedglomeruli, tubular atrophy, and calcification. Longterm prophylactic antibiotics were introduced. Evi-dence of impaired renal function persisted. At the ageof 13 years her creatinine was 150 tmol/l and ureawas 8 1 mmol/l. It is anticipated that she will requiredialysis or a transplant during adolescence.

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Cardiac catheterisation was performed at 18 monthsto investigate a heart murmur. This showed a mildbilateral pulmonary branch stenosis.Growth and development were impaired. She was

of short stature throughout childhood. Thelarchebegan at 9 years and menarche at II years. By 13 yearsshe had reached a height of 136 cm (below 3rdcentile). A moderate degree of mental handicapresulted in attendance at a school for children withspecial educational needs and only partial inde-

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pendence in day to day activities at 13 years. A partialleft sensorineural hearing loss was detected.A convergent strabismus was corrected at 11 years.

Ophthalmological review at 13 years confirmed thepresence of bilateral posterior embryotoxon.Her dysmorphic features were compatible with

those seen in Alagille syndrome (fig IA and B). Otherphysical features were a short neck, small ears,crowded teeth, small hands and feet, and mildhypoplasia of all finger and toenails.

Figure I (A) Facial appearance ofcase I in infancy and (B) at 13years. Note thefrontal bossing with a concave nasal bridge,upward slanting palpebralftssures, and a short philtrum. (C) The de novo deletion (20) (p1l.23-p13) in case 1.

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Alagille !yndrome and deletion of20p

Radiological examination showed butterfly-likedeformity affecting multiple thoracic vertebrae with acomplete sagittal cleft of the body of T4.

Cytogenetic studiesCytogenetic studies showed deletion of the short armof chromosome 20 with a breakpoint in band20plI.23 (46,XX,del(20XpIl.23-pI3) (fig IC). Paren-tal chromosomes were normal.

CASE 2This female infant was born at term by normal vaginaldelivery after a normal pregnancy weighing 3310 g.The mother at that time was 29 years of age and father32 years of age and they were unrelated and of whiteEuropean origin. A male sib born in 1983 was normal.At birth the proband was noted to have a left sided

cleft lip, a high arched palate, small, low set ears,overfolded helices, a flat nasal bridge, hypertelorism,epicanthic folds, widely separated skull sutures with alarge anterior fontanelle, a short neck, and generalisedhypotonia. During the first week of her life anejection systolic murmur was noted in the pulmonaryarea radiating to the back. ECG showed right ventri-cularhypertrophy and features ofthe Wolff-Parkinson-White syndrome. Echocardiogram showed a leftsuperior vena cava joining into the coronary sinus anda normal pulmonary valve and main pulmonaryartery. These findings were felt to be consistent with amild to moderate peripheral pulmonary artery stenosis.Skeletal survey showed spina bifida occulta of the firstcervical vertebra and hypoplastic acetabula.From birth she exhibited poor sucking with un-

coordinated swallowing and had several episodes ofaspiration with poor weight gain. This initial failure tothrive persisted throughout life and despite severalhospital admissions and attempts to increase calorieintake she remained below the 3rd centile.At the age of 3 years 6 months she developed mild

jaundice. Investigations showed slightly raised liverenzyme levels but there was no evidence of chronichepatitis or autoimmune disease. The jaundicegradually resolved and on review at the age of 5 yearsher total serum bilirubin was 6 mmol/l. However,both serum alkaline phosphatase and gamma-glutamyltranspeptidase were raised (666 IU/l and 87 IU/1,respectively). The rest of the liver enzymes and serumlipids were within normal limits.

Examination at 18 months and 5 years of ageshowed a short, thin child wearing hearing aids whohad the following facial features: successfully repairedleft cleft lip, long philtrum, moderate hypertelorism,a depressed nasal bridge, frontal bossing, and down-ward slanting palpebral fissures. She had a short chestwith pectus carinatum. Cardiovascular examinationindicated persistence of the pulmonary systolic

murmur. Ophthalmological examination showed noevidence of eye defects. The facial appearance was nottypical of Alagille syndrome (permission to publishphotograph withheld).

Developmental history indicated moderate globaldelay. Her milestones had been delayed to sittingunsupported at 18 months and walking independentlyat 3 years. She began to vocalise at 3 years 7 months.Significant hearing impairment had been noted andbilateral hearing aids provided. She has been placed ina school for the educationally subnormal. At the ageof 5 years she could say several words and was able tojoin two or three words together in sentences, thoughher speech was difficult to understand. She washypotonic but not weak.

Cytogenetic studiesStudies in the neonatal period showed a deletion ofthe short arm of chromosome 20. Subsequent analysisshowed the breakpoint to be at p12 (46,XX,del(20)(pl2-pl3) (fig 2). The maternal karyotype wasnormal, but the father and brother were not availablefor testing.

CASE 3This female child was the first offspring of unrelatedparents of white European origin, aged 28 and 27years. She was born on 10.5.88 after a normal deliveryat 39 weeks' gestation, birthweight 3090 g. Thepregnancy was complicated by an episode of gastro-enteritis but was otherwise normal. She was admittedto special care because of failure to suck adequatelyand mild hypoglycaemia, but was discharged at 3days. She was readmitted at 4 days of age withirritability, jaundice, and feeding problems. On day 6abnormal liver function tests but a normal liverultrasound were reported. She was discharged at 17days and readmitted at 6 weeks with hepatomegaly

2222 1

122 221 121 1

1 2 =1-22 2222

12 2133 2

1 31 3213-311332 '2213333

Figure 2 Cytogenetic abnormality in case 2:46,XX,del(2OXpI2-p13).

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and splenomegaly, possibly owing to neonatal hepa-titis. Persistent jaundice was noted and a liver biopsyshowed chronic persistent hepatitis. An echocardio-gram showed patent ductus arteriosus with possibleperipheral pulmonary artery stenosis. Eye examinationindicated anterior segment changes of posteriorembryotoxon. X ray investigation showed a normalspine and normal ribs.

Examination at 6 months of age showed a smallinfant with both height and weight on the 3rd centile.There was no evidence of jaundice. Frontal bossing,antimongoloid slant, and a prominent nasal root were

Figure 3(A) Facial appearance of case 3 showing deep set

eyes and a short philtrum. (B) The de novo interstitial delet'wn(2OXpI.2-12) in case 3.

evident (fig 3). The head circumference was 40-8 cmwith an inner canthal distance of 2-6 cm, inter-pupillary distance of 4-5 cm, and an outer canthaldistance of 7-1 cm. These measurements representmild hypertelorism in a child otherwise on the 3rdcentile for physical growth. Repeat examination at 14months showed a head circumference of 42-5 cm anda weight and height still on the 3rd centile. Develop-ment was grossly normal.

Cytogenetic studiesThe karyotype showed an interstitial deletion in theshort arm of chromosome 20 with breakpoints withinbands pll.2 and p12, 46,XX,del(20)(pll.2pl2). Theabnormal chromosome is shown in fig 3B. Parentalchromosomes were normal.

CASE 4This 3 year old boy (fig 4A) presented at six weeks forfurther investigation of prolonged conjugated hyper-bilirubinaemia, renal dysplasia, congenital heartdisease, abnormal facies, and poor weight gain. Hewas born at 37 weeks' gestation by forceps deliveryafter an uneventful pregnancy complicated only bythe finding of intrauterine growth retardation and adilated right renal tract on ultrasound scan. Theinfant's immediate neonatal period was complicatedby a septicaemic illness and acute renal failure. Heresponded to antibiotics and conservative manage-ment of his renal failure. The infant was first noted tobe jaundiced on day 2. The jaundice persisted and, onfurther investigation, was found to be obstructive innature.On examination, he was a small, jaundiced infant

weighing 2-65 kg. He had abnormal facies with a highforehead, small, pointed chin, and an iris colobomaon the right. He had a long systolic murmur at thepulmonary area radiating to all parts of the lung fieldswith a soft pulmonary component to the secondsound. He had a 2 cm palpable, smooth, non-tenderliver. There were no other abnormalities.

Investigations confirmed a conjugated hyper-bilirubinaemia with abnormal liver function tests, anda raised creatinine of 70 imol/l. Abdominal ultrasoundscan showed a normal liver and gallbladder andbilateral small dysplastic kidneys with dilatation ofthe right collecting system. A liver biopsy showedbiliary duct hypoplasia. An electrocardiogram showedright ventricular strain and echocardiographyindicated pulmonary stenosis and pulmonary arterystenosis. Ophthalmological assessment confirmed thepresence of unilateral anterior segment dysmorphismbut no other abnormalities. All other investigationswere normal.A diagnosis of Alagille syndrome was made on the

basis of biliary duct hypoplasia, abnormal facies,

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Alagille syndrome and deletion of20p

Figure 4 (A) Facial appearance ofcases 4 and 5. (B) Cytogeneticabnormality showing an interstitialdeletion present in mother and son:del(2OXpIl.2-p12.2).t B

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congenital heart disease, and growth retardation.There was no evidence of vertebral arch defects or

hypogonadism.The infant was treated initially with modified feeds,

vitamin supplements, cholestyramine, and pheno-barbitone. He remained well although his growth was

poor, persistently well below the 3rd centile butparallel to it. He showed evidence of developmentaldelay.The child was readmitted at 17 months of age with

poor weight gain and diarrhoea, for which no explana-tion could be found. It has been noted that some

patients with Alagille syndrome have pancreaticinsufficiency and the child was therefore startedempirically on pancreatic enzymes. He respondedwell with good weight gain and the diarrhoea ceased.Despite continual surveillance for signs of rickets, thechild presented at 2 years 3 months with clinicalrickets confirmed biochemically and radiologically.Rapid healing occurred with treatment with oralcalcium supplements and intramuscular injections ofcholecalciferol.

At the age of 2 years 10 months the boy presentedwith a rash on his hands which had been worseningover the past two weeks. Examination by a derma-

Table I Principal features of Alagille syndrome.

Major featuresChronic cholestasis 91Characteristic facies 95Cardiac abnormalities (68/80) 85

(Peripheral pulmonary artery stenosis56/80) 70

Vertebral abnormality (butterfly-like) 87Posterior embryotoxon* 88

Mior featuresMental retardation 16Growth retardation 50Other skeletal abnormality (vertebral)

23/43 53Glomerular renal involvement (mesangiolipidosis)

17/23 74

*Embryotoxon: a congenital condition of the eye in which themargin of the cornea is opaque. The condition resembles that of arcussenilis and is sometimes referred to as arcus juvenilis.

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Anad, Burn, Matthews, Cross, Davison, Mueller, Sands, Lillington, Eastham

tologist showed areas of erythema, vesicles, crustedlesions with blood staining, and considerable scarring.The lesions were thought to be compatible withporphyria cutanea tarda or variegate porphyria.Porphyria screening was therefore carried out whichshowed normal urine and stool porphyrins, butmassively raised blood protoporphyrins making adiagnosis of erythropoietic protoporphyria.

Cytogenetic studiesChromosome analysis showed an interstitial deletionof chromosome 20 (pll.23-pl2.2) (fig 4B).

CASE 5A review of the parents of case 4 showed that hismother also had the typical facies of Alagille syndrome(fig 4A) with a high forehead and pointed chin. Inaddition, like her son, she had iris colobomatabilaterally. Further enquiries showed that she hadnever been jaundiced as an infant but was beingfollowed up because of right ventricular outflow tractobstruction. She had been reviewed by an ophthal-mologist as a child and found to have both bilateralanterior segment dysmorphism and posterior embryo-toxon. She was also oflimited intelligence. A diagnosisof Alagille syndrome without hepatic involvement wastherefore made. The child's father was entirelynormal. Both parents had normal porphyrin levels.The mother's chromosomes showed the same patternas that described in her son (fig 4B). The father hadnormal chromosomes.

DiscussionTable 1 contains a recent review of the clinicalfeatures of 80 cases of Alagille syndrome9 and table 2those of all cases of deletion of chromosome 20pincluding our five cases. Cases 1, 3, 4, and 5 in thepresent report show clinical features which would becompatible with a diagnosis of Alagille syndrome.Case 2 has evidence of the typical heart defect, growthand mental retardation, vertebral abnormalities, andsome liver function derangement though the cranio-facial features shared less in common with Alagillesyndrome. This may be the result of a somewhat moredistal breakpoint involving 20p12 and amore extensiveloss of material from the short arm. The significanceof the erythropoietic protoporphyria in case 4 remainsto be elucidated.These cases together with the reports by Schnittger

et al,6 Zhang et al,7 and Legius et al8 confirm theproposal by Byrne et aP that Alagille syndromeresults from a genetic defect on the short arm ofchromosome 20 and we concur with the proposal bySchnittger et a16 that the characteristic features result

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FigureS Diagrammatic representation of 15 cases of20pdeletion. Note that cases I and 2from the present report are notshown as extending to the telomere. This is based on preliminarymolecular genetic studies which have shown the presence oftwocopies ofprobesfrom the telomeric region (I Hansmann, personalcommunication). The area ofminimal overlap (AMO) oncytogenetic grounds is pll.23-p12.2.

from a deletion involving the region p1 1.23-pl2.2 (fig5). Molecular genetic studies are in progress to refinethe area of minimum overlap further and to establishwhether this syndrome constitutes a contiguous genedefect. Linkage studies in familial cases of Alagillesyndrome are now indicated.

The following clinicians have been involved in thecare and investigation of the reported cases: Case 1 DrM Oo, Dr M Coulthard; case 2 Dr F N Porter, DrKuzemko, Dr H L Smith; case 3 Dr R Beach, Dr CBennett, Dr C Ball, Dr G Mieli-Vergani. Linda Burnprepared the manuscript.

I Alagilie D, Habib EC, Thomassin N. L'atresie des voies biliairesintrahepatiques avec voies biliaires extrahepatiques permeables chezl'enfant. Paris: Editions Medicales Flammarion, 1969:301-18.

2 Watson GH, Miller V. Arteriohepatic dysplasia: familial pul-monary arterial stenosis with neonatal liver disease. Arch DisChild 1973;48:459-66.

3 Mueller RF, Pagon RA, Pepin MG, et al. Arteriohepaticdysplasia: phenotypic features and family studies. Clin Genet1984;25:323-31.

4 Shulman SA, Hyams JS, Gunta R, Greenstein RM, Cassidy SB.Arteriohepatic dysplasia (Alagille syndrome): extreme variabilityamong affected family members. Am J Med Genet 1984;19:325-32.

5 Byrne JLB, Harrod MJE, Friedman JM, Howard-Peebles PN.Del(20p) with manifestations of arteriohepatic dysplasia. Am JMed Genet 1986;24:673-8.

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Alagille syndrome and deletion of20p

6 Schnittger S, Hofers C, Heidemann P, Beermann F, HansmannI. Molecular and cytogenetic analysis of an interstitial 20pdeletion associated with syndromatic intrahepatic ductularhypoplasia (Alagille syndrome). Hum Genet 1989;83:239-44.

7 Zhang F, Deleuze J, Aurias A, et al. Interstitial deletion of theshort arm of chromosome 20 in arteriohepatic dysplasia(Alagille syndrome). Pediatr 1990;116:73-7.

8 Legius F, Fryns JP, Eyskens B, et al. Alagille syndrome (arterio-hepatic dysplasia) and del(2OXp 1.2). AmJ Med Genet 1990;35:532-5.

9 Alagille D, Estrada A, Hadchonel M, Gautier M, Odievre M,Dommergues JP. Syndromic paucity of interlobular bile ducts(Alagille syndrome or arteriohepatic dysplasia): review of 80cases. Pediatr 1987;110:195-200.

10 Kalousek K, Therien S. Deletion of the short arm of chromosome20. Hum Genet 1976;34:89-92.

11 Kogame K, Fukuhara T, Maeda A, Kudo Y. A partial short armdeletion of chromosome 20(46,XY,del(20XplI). Jpn HumGenet 1978;23:153-60.

12 Garcia-Cruz D, Rivera H, Barajas LO, et al. Monosomy 20p dueto a de novo del (20) (pI2.2). Clinical and radiologicaldelineation of the syndrome. Ann Genet (Paris) 1985;28:231-4.

13 Vianna-Morgante AM, Richieri-Costa A, Rosenberg C. Deletionof the short arm of chromosome 20. Clin Genet 1987;31:406-9.

14 Silengo MC, Bell GL, Biagioli M, Franceschini P. Partial deletionof the short arm of chromosome 20:46,XX,del(20Xpl 1)/46,XXmosaicism. Clin Genet 1988;33:108-10.

15 Kiss P, Osztovics M. Deletion of the short arm of chromosome20. Clin Genet 1988;33:140-1.

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