IMed Genet 1998;35:165-168

Prenatal diagnosis by FISH of a 22ql 1 deletion intwo families

Marie-France Portnoi, Nicole Joye, Marie Gonzales, Suzanne Demczuk, Laurent Fermont,Gilles Gaillard, Guy Bercau, Genevieve Morlier, Jean-Louis Taillemite

Service d'EmbryologiePathologique et deCytogenetique, HopitalSaint-Antoine, 75012Paris, FranceM-F PortnoYN JoyeM GonzalesG MorlierJ-L Taillemite

Cytogenetics, PrenatalDiagnosis, MontrealChildren's Hospital,Montreal, Quebec,CanadaS Demczuk

Unite d'ExplorationsCardiologiques,Institut dePuericulture de Paris,75014 Paris, FranceL Fermont

Service de Gynecologieet Obstetrique, H6pitalSaint-Antoine, 75012Paris, FranceG Gaillard

Service de Gynecologieet Obstetrique, H6pitalNotre-Dame deBon-Secours, 75014Paris, FranceG Bercau

Correspondence to:Dr Portnoi.

Received 8 April 1997Revised version accepted forpublication 30 September1997

AbstractWe report on prenatal diagnosis by FISHof a sporadic 22qll deletion associatedwith DiGeorge syndrome (DGS) in twofetuses after an obstetric ultrasonographicexamination detected cardiac anomalies,an interrupted aortic arch in case 1 andtetralogy of Fallot in case 2. The parentsdecided to terminate the pregnancies. Atnecropsy, fetal examination showed char-acteristic facial dysmorphism associatedwith congenital malformations, confirm-ing full DGS in both fetuses. In addition tothe 22ql 1 deletion, trisomy X was found inthe second fetus and a reciprocal balancedtranslocation t(l 1 ;22) (q23;ql 1) was foundin the clinically normal father of case 1.These findings highlight the importanceof performing traditional cytogeneticanalysis and FISH in pregnancies with ahigh risk of having a deletion.(7Med Genet 1998;35:165-168)

Keywords: DiGeorge syndrome; chromosome 22ql 1;FISH; microdeletion

DiGeorge syndrome (DGS), an aetiologicallyheterogeneous developmental field defect ofthe third and fourth pharyngeal pouches, ischaracterised by hypoplasia or aplasia of thethymus and parathyroid glands, a conotruncalheart defect, and varying craniofacialdysmorphism.' DGS is frequently associatedwith a chromosome 22 abnormality. Unbal-anced translocations which result in mono-somy of 22pter-22ql 1 and interstitial deletionshave mapped the DGS region to 22ql 1.2.2 3Molecular studies using probes for variousloci in the 22ql 1 region have detected sub-microscopic deletions in more than 90%of DGS cases without visible cytogeneticabnormalities.4 6

Deletions within 22q1 1 are associated with awide variety of birth defects embraced by theacronym CATCH 22,7 including Sprintzensyndrome (velocardiofacial syndrome),conotruncal congenital heart disease, and DGSat the more severe end of the clinical spectrum.This haploinsufficiency of 22ql 1 is a relativelyfrequent cause of birth defect (1/5000 livebirths).8 Fluorescence in situ hybridisation(FISH) using unique sequence DNA probes isan efficient, quick, and direct method fordetection of 22q 1 microdeletions.We report on prenatal diagnosis by FISH of

a 22ql 1 deletion associated with DGS in twofamilies. Both of these cases are sporadic.Moreover, in the first case, a reciprocal

balanced translocation t(I 1;22)(q23;ql 1) wasshown in the father's karyotype; in the secondcase trisomy X was associated with a 22ql 1deletion in the fetus.

Materials and methodsKARYOTYPING AND FISHFetal blood samples were obtained for karyo-type analysis and FISH. Cells were harvestedfrom cultures of phytohaemagglutinin stimu-lated lymphocytes and spread onto slides forthe production of G banded or R banded chro-mosomes.FISH of metaphase chromosomes using dig-

oxigenin labelled cosmid probes D22S75(N25, ONCOR) from the DGS chromosomeregion was carried out basically according toPinkel et al.' This probe was premixed with acontrol cosmid (D22S39) in 22q13.3 facilitat-ing chromosome identification.

Case reportsCASE 1A 31 year old, nulliparous woman, who hadhad a previous miscarriage at 8 weeks of gesta-tion, was seen for fetal echocardiography at 23weeks of gestation after an obstetric ultrasono-graphic examination detected cardiac anoma-lies and absence of the thymic shadow. Fetalechocardiography showed an interrupted aor-tic arch (IAA) associated with a membranousventricular septal defect. The possibility ofDGS was discussed because of its strongassociation with IAA and absence of thethymus. A fetal blood sample was obtained forkaryotype determination. A 46,XX karyotypewith a 22ql 1 deletion detected by FISH wasfound, consistent with the DG phenotype andprovided a prenatal diagnosis of DGS (fig 1).The serum calcium level was determined andshowed hypocalcaemia (66 mg/l, normal fetal

Figure 1 Partial metaphase spread showing lack ofsignalon one of the chromosomes 22. The deleted chromosome 22with distal q arm marker present, but without DiGeorgeprobe signal is indicated by an arrowhead.

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Figure 4 Partial metaphase spread of the father of case 1after FISH showing a fluorescent signal on bothchromosome 22 homologues at 22q11.2 with probe D22S75(ONCOR) and at 11q23 on der(l 1) and on normalchromosome 22 with probe D22S39.

Figure 2 Fetus 1.

A

Fr

Figure 3 Fetus 2.

levels 90.2 + 8 mg/i). T cell subpopulationstudies were not performed. The parents wereinformed about the risks and decided to termi-nate the pregnancy.At necropsy, fetal examination showed facial

dysmorphism with hypertelorism, a squarenasal tip, a small, recessed chin, and a smallhypoplastic thymus (fig 2). Cardiac examin-ation confirmed the ultrasonographic findings.The aortic arch ended after the left commoncarotid artery, consistent with type B interrup-tion, and was associated with a ventricular sep-tal defect.

Neither parent had any evidence of a 22ql 1deletion by FISH. The mother had normalchromosomes and the father was found to havea balanced reciprocal translocation involvingchromosome 22, t(l 1;22) (q23;ql 1).

CASE 2Fetal blood was obtained for karyotype analysisfrom a fetus whose mother and father were 27and 40 years of age, respectively. Prenatalultrasound examination at 23 weeks showedtetralogy of Fallot (TF).The fetus was found to have a 47,XXX

karyotype and a 22qll deletion detected byFISH. Neither the serum calcium level nor thelymphocyte population was evaluated in thiscase. The TF associated with a 47,XXX karyo-type and a 22ql 1 deletion had obviousimplications for genetic counselling and riskassessment. Following counselling the preg-nancy was terminated.At necropsy, the fetus had facial dysmorphic

features including a broad nose, downwardslanting palpebral fissures with hypertelorism,a small mouth, microstomia, micrognathia,rounded, posteriorly rotated ears with absentear lobes, and facial hypertrichosis (fig 3). TFwas associated with a complete absence of thethymus, vertebral anomalies, and a left talipesequinovarus deformity.The parental karyotypes were both normal.

Deletion 22q1 1 was excluded by FISH.

DiscussionVery few data are available concerning theprognosis for fetuses diagnosed prenatally ashaving a 22ql 1 deletion. To the best of ourknowledge only three cases of prenatally

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167Prenatal diagnosis by FISH of a 22ql1 deletion

diagnosed DGS with a 22ql 1 deletion havebeen published.Van Hemel et al' described a familial case

illustrating the variable clinical expression ofthe chromosome 22ql 1 deletion. A 22ql 1deletion was found in a child who died twoweeks after birth with symptoms of full DGSand truncus arteriosus. This deletion wasdetected in the physically normal father whohad mild learning disabilities and a tendency todepression, as well as in a subsequent preg-nancy. Ultrasound studies did not show cardiacor other anomalies in the latter fetus. At birththe boy developed hypocalcaemia and had amoderate T cell deficit. Echocardiographyshowed a right sided aortic arch withoutintracardiac anomalies.Puder et al" reported a case of prenatally

diagnosed IAA, which led to the detection ofsegmental monosomy of chromosome 22qllby FISH in the fetus and in his mother, whohad a small ventricular septal defect.The first case of prenatal detection of a fetus

with an interrupted aortic arch and a 22qlldeletion, in the absence of a family history, wasreported by Davidson et al. 12 The mother wasalso found to carry the deletion; she had nocardiac abnormality but was mildly retarded.The pregnancy continued and the prenataldiagnosis was helpful for planning postnatalcare. In this family and in others, the deletionwas transmitted from the mother to her moreseverely affected child. The range of pheno-types associated with the 22ql 1 deletion com-plicates genetic counselling. Moreover, so farthere has been no experience with the progenyof severely affected subjects who nowadayssurvive after heart surgery.Both ofour cases were referred for cordocen-

tesis and fetal blood karyotype analysis in the23rd week of pregnancy because of detectionby ultrasound examination of a cardiacanomaly, an IAA associated with thymic aplasiain the first case and TF in the second.IAA, truncus arteriosus, unusual cardiovas-

cular lesions, and TF are congenital conotrun-cal heart diseases commonly found in DGS."The results of recent molecular studies inpatients with TF, including isolated TF andsyndromic cases, suggest that isolated TF is notassociated with a 22ql 1 deletion, whilehemizygosity for 22ql 1 is present in patientswith TF associated with additional cardiovas-cular or non-cardiac anomalies.'4 '5 Therefore,detection of these cardiac diseases duringpregnancy should prompt the investigation ofcharacteristic abnormalities associated withDGS. These associated anomalies are, how-ever, difficult to detect by prenatal diagnosis.The discovery during pregnancy of a

conotruncal heart defect associated with a22q 1 deletion might indicate a severe form ofthis syndrome, which is known to show greatphenotypic variability. The most severely af-fected patients have serious, life threateningheart defects and die neonatally. The leastseverely affected patients have only mild facialanomalies and some developmental delay, butno heart defects.'6 However, most of the

patients with DGS who have survived infancyhave been mildly to moderately retarded.'The presence of a deletion associated with

conotruncal heart disease is an important pointwhich has obvious implications for geneticcounselling. In both of our cases, necropsyconfirmed that the fetuses were severelyaffected and had multiple congenital anoma-lies. This study, the first to our knowledge thatdescribes fetal examination of DGS with a22ql 1 deletion, has shown that facial dysmor-phism is present and recognisable in the fetusand is very similar to that of the newborn.

Furthermore, in our report, karyotypeanalysis showed a second chromosomalabnormality. In the first case, the father wasfound to have a balanced translocationt(1 1;22)(q23;ql 1). FISH analysis showedD22S75 to be present on the derivative 22translocation chromosome and to be locatedcentromeric to the translocation breakpoint in22ql1 (fig 4). The presence of a 22ql 1deletion in the offspring of t(l 1;22) carriers hasnot been reported previously. An attractivehypothesis is that this recurrent translocationin the father may have played a role in theexistence of the deletion in the fetus. Structuralcharacteristics of the DNA in the 22ql 1region, especially the presence of low copyrepeat elements, would favour interstitial dele-tion with more or less precise breakpointsites.'6 17 However, the role of chance cannot beexcluded. Published data have shown a higherfrequency of deletions of maternal origin(70%).8 In the present case, determination ofthe parental origin of the deleted chromosome22 in the fetus was attempted, but was notinformative with the polymorphic marker used.Further endeavours to obtain other polymor-phic markers mapping to the DGS deletedregion and to continue the investigation intothis family are in progress.When neither parent has a deletion, it is

expected that there will be a low risk of havinga further child with a 22ql 1 deletion sincegonadal mosaicism cannot be ruled out. In thisfamily, an early first trimester prenatal diagno-sis is relevant because of the risk of inheritingan unbalanced translocation derivative in theoffspring.

In the second case a triple X karyotype wasfound and the association of triple X with DGShas never previously been reported either.However, trisomy X is a relatively commonaneuploidy and we suggest that this associationis likely to be a chance event. The finding ofthis anomaly might be an indication for prena-tal diagnosis in a subsequent pregnancy.These findings highlight the importance of

performing traditional cytogenetic analysis andFISH for suspected DGS during pregnancywhen ultrasound studies show a conotruncalcardiac defect. In these cases prenatal diagnosisof a molecular deletion in the DGS criticalregion of chromosome 22 suggests the exist-ence of associated anomalies and risks ofdevelopmental delay. However, counsellingremains difficult in view of the clinical variabil-

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ity described in DGS, where the phenotypecannot be accurately predicted from the geno-type.

We are grateful to F Langlet and C Souleyan for their technicalassistance. We thank Dr B Lecolier for the fetal serum analysisand Dr C Horn for critical reviewing of the manuscript.

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