American Journal of Medical Genetics 137A:47–51 (2005)
22q11.2 Duplication Syndrome:
Two New Familial Cases With Some Overlapping
Features With DiGeorge/Velocardiofacial Syndromes
Marie-France Portnoı ¨,1* Fanny Lebas,2Nicolas Gruchy,1Azarnouche Ardalan,1
Vale ´rie Biran-Mucignat,2Vale ´rie Malan,1Lina Finkel,1Gilles Roger,3Sarah Ducrocq,2
Francis Gold,2Jean-Louis Taillemite,1and Sandrine Marlin4
1Laboratoire de Cytoge ´ne ´tique, Hopital Saint-Antoine AP-HP, Paris, France
2Service de Ne ´onatologie, Hopital Armand-Tousseau AP-HP, Paris, France
3Service d’Oto-Rhino-Laryngologie, Hopital Armand-Tousseau AP-HP, Paris, France
4Unite ´ de Ge ´ne ´tique me ´dicale, Hopital Armand-Tousseau AP-HP, Paris, France
Twenty-one patients, including our two cases,
with variable clinical phenotype, ranging from
mild learning disability to severe congenital
DiGeorge/velocardiofacial syndromes (DG/VCFS),
have been shown to have a chromosome duplica-
tion 22q11 of the region that is deleted in patients
with DG/VCFS. The reported cases have been
have escaped identification and been missed by
routine cytogenetic analysis. Here we report on
two inherited cases, referred to us, to rule out
22q11 microdeletion diagnosis of VCFS. The first
patient was a 2-month-old girl, who presented
with cleft palate, minor dysmorphic features
including short palpebral fissures, widely spaced
eyes, long fingers, and hearing loss. Her affected
mother had mild mental retardation and learning
disabilities. The second patient was a 7½-year-old
boy with velopharyngeal insufficiency and mild
developmental delay. He had a left preauricular
tag, bifida uvula, bilateral fifth finger clinodac-
tyly, and bilateral cryptorchidism. His facial
features appeared mildly dysmorphic withhyper-
telorism, large nose, and micro/retrognathia. The
affected father had mild mental retardation and
had similar facial features. FISH analysis of
interphase cells showed three TUPLE1-probe
signals with two chromosome-specific identifica-
tion probes in each cell. FISH analysis did not
show the duplication on the initial testing of
metaphase chromosomes. On review, band q11.2
was brighter on one chromosome 22 in some
metaphase spreads. The paucity of reported cases
of 22q11.2 microduplication likely reflects a com-
of diagnosis by FISH analysis on metaphase
spreads. These findings illustrate the importance
of scanning interphase nuclei when performing
FISH analysis for any of the genomic disorders.
? 2005 Wiley-Liss, Inc.
KEY WORDS:22q11.2 duplication; interphase
FISH; chromosome 22; genomic
The chromosome 22q11.2 has long been implicated in
(DG/VCFS), der(22) and cat-eye syndrome (CES), which are
associated with either decreased or increased gene dosage
[Edelmann et al., 1999b; Shaikh et al., 2000]. CES and der(22)
syndrome are rare disorders characterized by duplications
(tetrasomy and trisomy, respectively) of part of 22q11.2
[McDermid and Morrow, 2002]. The microdeletions of
22q11.2 that are associated with DG/VCFS occur more often
in the general population, with an estimated frequency of
1/4,000 live births [Lindsay, 2001; Yamagishi, 2002; Botto
et al., 2003].
These different congenital anomaly disorders share a
physical region of overlap containing copies of chromosome
22-specific low-copy repeats (LCRs) [McDermid and Morrow,
2002]. LCR22s are presumed to predispose to homologous
recombination events and to mediate nonallelic homologous
recombinations that result in rearrangements of 22q11.2.
Meiotic inter- or intra-chromosomal nonallelic homologous
recombination events, due to unequal crossovers between
LCR22s, might generate the common 3 Mb deletion in DG/
VCFS and the reciprocal 3 Mb interstitial duplication
[Edelmann et al., 1999a,b; Shaikh et al., 2000, 2001].
Twenty-one patients with variable clinical phenotype,
ranging from mild learning disability to severe congenital
malformations or some overlapping features with DG/VCFS,
have beenshown to harbor a chromosome duplication 22q11 of
the genomic region that is deleted in patients with DG/VCFS.
The microduplication of 22q11.2, establishing the comple-
mentary genomic disorder of the 22q11.2 deletion syndrome,
appears to represent a new syndrome [Ensenauer et al., 2003;
Hassed et al., 2004a]. All cases have been identified primarily
by interphase FISH and could escape identification and are
22q11.2, with some overlapping features with DG/VCF
syndrome, determined by FISH studies.
The patient was born at term to a 23-year-old G2P1 mother
and to a 39-year-old father. Her birthweight was 2,920 g,
*Correspondence to: Marie-France Portnoı ¨, Laboratoire de
Cytoge ´ne ´tique, Ho ˆpital Saint-Antoine 75012 Paris, France.
Received 9 December 2004; Accepted 30 May 2005
? 2005 Wiley-Liss, Inc.
length 50 cm, and head circumference 35 cm. At age 2 months,
because of the presence of a posterior cleft palate and minor
facial anomalies, she was referred to us to rule out DG/VCFS.
The facial anomalies consisted of widely spaced eyes, short
palpebral fissures, short and beaked nose, micrognathia,
dysplastic ears with preauricular pit on the right (Fig. 1A, B).
Arachnodactyly of fingers and toes was noted. In addition, she
ultrasound examinations, she had no detectable renal and
cardiac anomalies. At age 6 months, she was treated for
urinary tract infection. Her weight, height, and OFC were
She had shown significant hypotonia. She was unable to
maintain a sitting position due to poor trunk control.
Her father was normal. The mother had mild mental
retardation with learning disabilities and behavioral pro-
blems. Her facial features appear mildly dysmorphic and she
had brachymesophalangia (Fig. 1C). She had recurrent
urinary tract infections, but her kidneys had not been
examined by imaging. She underwent surgical treatment for
bilateral clubfoot. There were cognitive deficits in her family
history. Her parents were not available for testing.
The patient was a 7½-year-old boy who was referred for
genetic evaluation ofvelopharyngeal insufficiency withhyper-
nasalspeech. Hewasborn attermbycesarean sectionbecause
of maternal preeclampsia to a 34-year-old G1P1 mother and
35-year-old father, after a pregnancy complicated by hydram-
nios. His birth measurements were within the normal range.
The mother had amniocentesis at 32 weeks of gestation.
Chromosome analysis showed a normal karyotype, 46,XY.
FISH was not used onprenatal chromosome study. During the
first months of life, he had feeding problems and recurrent
gastroesophageal reflux. At 4 months of age, he was hospita-
lized for severe laryngomalacia requiring surgical treatment.
He had a left preauricular tag, bifida uvula, bilateral fifth
finger clinodactyly, large toes and bilateral 2, 3, 4 partial
cutaneous syndactyly, bilateral cryptorchidism, short stature,
and absence of 12th rib. High resolution chromosome analysis
showed a normal karyotype.
by 20 months. At age 7½ years, he measured 114 cm in height
(2 SD below the mean) and had an OFC of 52 cm. His facial
features appeared mildly dysmorphic with hypertelorism,
short palpebral fissures, large nose with prominent nasal
retrognathia (Fig. 1D,E). There was a significant delay in
language skills and mild developmental delay. He had been
anomalies on clinical and ultrasound examinations.
Both parents had learning disabilities. The father’s facial
features appear mildly dysmorphic (Fig. 1F). There was no
family history of miscarriages or congenital anomalies.
Blood samples from the patients and their parents were
processed by standard chromosome and FISH procedures.
FISH analysis was performed using DNA fluorescent probes
for the DG/VCFS critical region, N25 (CLTD) (Q-Biogene,
Illkirch, France) and TUPLE1 (Vysis, Downers Grove, IL), at
22q11.2, and a control probe at 22q13.3.
In order to establish the size of the duplication in each
patient, FISH analysis was performed using four bacterial
artificial chromosomes (BACs) clones, b476c20, 519d21,
b135h6, CTA-526G4, each clone in the vicinity of LCRs
spanning 22q11.2, centromeric and telomeric to TUPLE 1, as
previously described [Ensenauer et al., 2003]. Stab cultures of
the BACs were received from the Wellcome Trust Sanger
DNA were performed using the QIAGEN plasmid Maxi Kit,
according to the manufacturer’s instructions. Nick translation
Meylan, France). The digoxigenin-labeled DNA probes were
applied to interphase and metaphase cells from the patients.
A, B: Patient 1 at age 2 months. Arrow points to preauricular pit. C: Mother of patient 1. D, E: Patient 2 at age 7½ years. F: Father of patient 2.
48Portnoı ¨ et al.
all 100 cell nuclei analyzed (Fig. 2C in color online). Initial
FISH analysis of metaphase cells did not reveal the duplica-
tion. Further analysis showed that band q11.2 was brighter on
one chromosome 22 in some metaphases (Fig. 2B). In both
cases, the affected mother and the affected father were found
by FISH to have the same duplication as their own child.
Analysis with FISH probes from BACs showed three signals
(duplication) for two adjacent clones, 519d21 and b135h6
(Fig. 2D,E), and normal signal pattern (two signals) for the
more proximal clone, 476c20, and for the more distant, CTA-
526G4 (Fig. 2F). The breakpoints mapped to the proximal and
distal LCRs implicated in the 3 Mb DG/VCFS deletion,
suggesting that the patients had a 3 Mb-sized duplication.
Like other microdeletion syndromes with a paired condition
due to duplication of the same genomic region, the 22q11.2
deletion syndrome has a complementary 22q11.2 duplication
syndrome [Lupski, 1998; Potocki et al., 2000; Shaw et al.,
To our knowledge, the first case of microduplication 22q11.2
wasdescribedbyEdelmann etal. [1999b].Thepatient wasa4-
year-old girl with a narrow face, downslanting palpebral
fissures, velopharyngeal insufficiency, and a significant delay
presented with failure to thrive, marked hypotonia, and
seizure-like episodes. The normal mother and grandmother
were found to carry the duplication. Both individuals had a
history of preauricular ear pits.
A systematic search for microduplication 22q11.2 was
undertaken by Ensenauer et al. , in a population of
650 patients referred for deletion analysis for DG/VCFS. They
found 10 unrelated patients (1.5% of patients) with micro-
duplication 22q11.2, defining a new 22q11.2 microduplication
syndrome. The clinical phenotype of patients in this series
appears variable, ranging from mild learning disability to
severe congenital malformations leading to early death. Two
as palatal clefting, velopharyngeal insufficiency, or thymus
aplasia are features also observed in patients with DG/VCFS.
However, the patients have other distinctive characteristics,
including widely spaced eyes, superior placement of eyebrows,
downslanting palpebral fissures with or without ptosis, mild
micro/retrognathia and a long, narrow face, were observed.
One patient presented with preauricular tags and pits, which
have been reported to be the most consistent finding in CES.
There is some partial overlap of manifestations between dup
22q11.2 and CES [Rosias et al., 2001; Ensenauer et al., 2003].
Hassed et al. [2004a] report a family with a paternal
duplication 22q11.2 and relatively mild phenotype suggestive
of VCFS. The patient, a 5-month-old girl, had cleft palate,
failure to thrive, gastro-esophageal reflux, hydronephrosis,
and mild developmental delay. She had low-set posteriorly
rotated ears, down-slanting palpebral fissures, micrognathia,
learning disabilities. The father and both siblings had similar
facial features. All three had the same duplication.
Likewise, our two cases were ascertained as suspected DG/
VCFS cases because of palatal clefting/insufficiency with
Recently, new cases of duplication 22q11.2 have been
described briefly in abstracts. Two de novo and one mater-
nally-inherited 22q11.2 duplications have been reported in
patients with hypernasal speech, neuropsychological pro-
blems, learning disability, and subtle dysmorphic features
[Beiraghi et al., 2004]. Hassed et al. [2004b] report another
family case. The patient was an 18-year-old female with aortic
hypernasal speech and features of Asperger syndrome. At
19 years of age, the brother had cerebral vascular accidents.
The father and both siblings had speech problems and were
mildly dysmorphic with hypertelorism and downslanting
palpebral fissures. All three were found to have the same
larger-sized signal compared with the normal chromosome 22 (small arrow). C: Interphase cells showing duplication of TUPLE1 (three red signals, long
to TUPLE1. F: Interphase cells showing normal signal pattern (two signals) for BAC CTA-526G4, telomeric to BAC b135h6.
A: Partial karyotype. Arrows point to G-banded chromosome 22 pair from one patient. The dup(22)(q11.2) is not visible. B: FISH metaphase
22q11.2 Duplication Syndrome49
TABLE I. Phenotypic Findings Associated With 22q11.2 Microduplications
Number of patients
4 (3 Related)
Long narrow face
Superior placement of
Minor ear anomalies
With cleft palate
Seizure or abnormal EEG
Two prospective studies have also been reported in
abstracts. One was undertaken on 200 cases referred for DG/
VCFS [Lamb et al., 2004]. One of the 200 patients (0.5%) was
identified with a microduplication 22q11.2. The patient was a
22-month-old male with tetralogy of Fallot, moderate hearing
loss, global developmental delay, and facial dysmorphic
features. Another prospective study was undertaken by using
a real-time PCR array to measure relative dosage of two genes
at the chromosome 22q11.2 locus, in 98 females with
intellectual disability [Somerville et al., 2004]. Two of these
98 cases (2%) were found to have a 22q11.2 duplication. One
patient had heart and hearing defects, in addition to a cogni-
tive deficit. The other patient was a mother of 2 male children
who were found to carry the 22q11 duplication. One had a
significant cognitive deficit, and the other had marked
In previous reports and in our cases, use of interphase FISH
analysis was primarily responsible for the diagnosis. Inter-
phase FISH is warranted in identifying these cases, because
the ability to see duplication on metaphase cells in 22q11.2,
both by cytogenetic analysis and FISH, is limited. These
findings illustrate the importance of scanning interphase
nuclei when performing FISH analysis for any of the genomic
disorders [Lupski, 1998].
The sizeofthe duplication, determined byFISH probesfrom
BACs and PACs, mostly corresponds to the same 3 Mb region
that is deleted in DG/VCFS patients, ranging from 3 (most
common) to 4 and 6 Mb. The duplication breakpoints occur in
the same proximal and distal LCRs as for the DG/VCFS
patients with the common deletion [Edelmann et al., 1999a;
Ensenauer et al., 2003].
Thus, a total of 21 unrelated patients, including our two
cases, with a microduplication 22q11.2, have now been
reported (Table I). Nine cases have been found to be familial.
Not all parents were available for testing. Intrafamilial
variability of the condition was observed. The rarity of cases
with duplication 22q11.2 compared to those of 22q11.2 micro-
deletion may be ascribed to the limits of microscopic meta-
phase chromosome analysis, as well as the mild nonspecific
associated clinical findings. Ascertainment bias may con-
tribute to underestimation of the real incidence of these
However, the 22q11.2 duplication syndrome may not be so
benign a condition and has been reported in individuals with a
range of features including palate, urogenital abnormalities,
hearts defects, hearing loss, cognitive, behavioral defects,
and psychiatric abnormalities. Expression of these features
has been variable, with a wide range of severity [Ensenauer
et al., 2003]. The most consistent findings have been some
degree of intellectual disability, neuropsychological problems,
and speech disorder, sharing features with 22q11-deletion
NOTE ADDED IN PROOF
Since the original submission of our manuscript, two cases
of patients with this syndrome, and the first patient wtih a
22q11.2 triplication were reported [Yobb et al., 2005]. This
additional report further supports that the phenotype of these
patients with microduplication is extremely diverse.
We thank Isabelle Vollot and Catherine Jacquet for their
expert technical help. We also thank David Marsh for careful
linguistic text revision.
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22q11.2 Duplication Syndrome51