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Human Genetics ISSN 0340-6717 Hum GenetDOI 10.1007/s00439-011-1020-y
The role of the TCF4 gene in thephenotype of individuals with 18qsegmental deletions
Minire Hasi, Bridgette Soileau, CourtneySebold, Annice Hill, Daniel E. Hale,Louise O’Donnell & Jannine D. Cody
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ORIGINAL INVESTIGATION
The role of the TCF4 gene in the phenotype of individuals with 18qsegmental deletions
Minire Hasi • Bridgette Soileau • Courtney Sebold •
Annice Hill • Daniel E. Hale • Louise O’Donnell •
Jannine D. Cody
Received: 15 February 2011 / Accepted: 25 May 2011
� Springer-Verlag 2011
Abstract The goal of this study is to define the effects
of TCF4 hemizygosity in the context of a larger seg-
mental deletion of chromosome 18q. Our cohort included
37 individuals with deletions of 18q. Twenty-seven had
deletions including TCF4 (TCF4?/-); nine had deletions
that did not include TCF4 (TCF4?/?); and one individual
had a microdeletion that included only the TCF4 gene.
We compared phenotypic data from the participants’
medical records, survey responses, and in-person evalu-
ations. Features unique to the TCF4?/- individuals
included abnormal corpus callosum, short neck, small
penis, accessory and wide-spaced nipples, broad or
clubbed fingers, and sacral dimple. The developmental
data revealed that TCF4?/? individuals were only mod-
erately developmentally delayed while TCF4?/-
individuals failed to reach developmental milestones
beyond those typically acquired by 12 months of age.
TCF4 hemizygosity also conferred an increased risk of
early death principally due to aspiration-related compli-
cations. Hemizygosity for TCF4 confers a significant
impact primarily with regard to cognitive and motor
development, resulting in a very different prognosis for
individuals hemizygous for TCF4 when compared to
individuals hemizygous for other regions of distal 18q.
Introduction
It has recently been shown that hemizygosity of the TCF4
gene causes Pitt–Hopkins syndrome (MIM ID #610042)
through a haploinsufficiency mechanism (Zweier et al.
2007). The TCF4 gene is located at 18q21.1 and is,
therefore, also hemizygous in some individuals with larger
segmental deletions of 18q. The goal of this study is to
define the effect of TCF4 hemizygosity in individuals with
18q deletions.
The constellation of phenotypic features known as
Pitt–Hopkins syndrome is characterized by severe intel-
lectual disability, wide mouth with fleshy lips, a beaked
nose, and intermittent hyperventilation followed by apnea
(Pitt and Hopkins 1978). Since it was first described, the
phenotype has been shown to have three different
genetic causes. It can be caused dominantly by hemi-
zygosity of or inactivating mutations in the TCF4 gene,
or recessively by mutations in the CNTNAP2 gene on
chromosome 7q35 or the NRXN1 gene on chromosome
2p16.3 (Zweier et al. 2009). Because this phenotype has
multiple underlying molecular mechanisms, the phrase
‘‘Pitt–Hopkins’’ refers only to the clinically-defined
syndrome.
Web Resources Online Mendelian Inheritance in Man (OMIM),
http://www.ncbi.nlm.nih.gov/Omim/.
M. Hasi � B. Soileau � C. Sebold � A. Hill �D. E. Hale � L. O’Donnell � J. D. Cody (&)
Department of Pediatrics, UT Health Science Center,
7703 Floyd Curl Dive, San Antonio, TX 78229, USA
e-mail: [email protected]
D. E. Hale � J. D. Cody
CHRISTUS Santa Rosa Children’s Hospital,
San Antonio, TX, USA
L. O’Donnell
Department of Psychiatry, UT Health Science Center
at San Antonio, San Antonio, USA
J. D. Cody
The Chromosome 18 Registry and Research Society,
San Antonio, TX, USA
123
Hum Genet
DOI 10.1007/s00439-011-1020-y
Author's personal copy
The identification of the genetic bases of Pitt–Hopkins
has allowed a fuller appreciation of the range of phenotypic
features associated with mutations or deletions of these
genes. It has recently been realized that 2% of individuals
with phenotypic Angelman syndrome actually had TCF4
aberrations (Takano et al. 2010). In addition, Rosenfeld
et al. (2009) identified seven cases with TCF4 deletions
which were referred for chromosomal microarray analysis
due to intellectual disability. In this genotypically ascer-
tained group, a reevaluation of the phenotypic effects of
TCF4 hemizygosity was undertaken. Of these patients,
only 3 of 7 had a breathing abnormality and none had
seizures, indicating that the penetrance of these features is
significantly less than 100%. In their review of published
cases likely caused by haploinsufficiency of TCF4
(N = 36), they found that the Pitt–Hopkins facial appear-
ance was present in 91% of cases. Other features seen in
more than half of the subjects included: severe psycho-
motor delay (97%), hypotonia (88%), happy disposition
(85%) (though no elaboration of this behavior was pro-
vided), single palmar crease (69%), microcephaly (64%),
constipation (58%), and brain abnormalities (56%).
Interestingly, variations in all three genes responsible
for Pitt–Hopkins syndrome are associated with schizo-
phrenia and autism. Hemizygous deletions of NRXN1 and
CNTNAP2 as well as SNPs in both genes are associated
with an increased risk of schizophrenia, epilepsy, and
autism spectrum disorder (Kirov et al. 2009; Friedman
et al. 2008) and SNPs of TCF4 have been associated with a
slightly increased risk of schizophrenia (Stefansson et al.
2009). In addition, individuals with 18q deletions including
TCF4, NETO1 and FBXO15 are more likely to exhibit
autistic-like behavior (O’Donnell et al. 2010). These data
point to a potential functional relationship between the
products of these three genes as well as a causal relation-
ship between autism and schizophrenia.
The TCF4 gene produces a basic helix-loop-helix (bHLH)
transcription factor which acts through binding to E-box
consensus sequences in the promoter regions of the target
genes. It belongs to a family of proneural bHLH transcription
factors controlling differentiation of neuronal subtypes.
The temporal and spatial differentiation of the numerous
neural cell types is controlled by a relatively small number
of transcription factors acting as homo- and hetero-dimers.
The hetero-dimerization of transcription factors produces a
greater diversity of regulatory combinations, thereby a
greater diversity and specificity of gene expression (Guille-
mot 2007). It is likely that the TCF4 gene product interacts
with numerous other transcription factors. In the mouse, Tcf4
dimerizes with Math1, another bHLH transcription factor.
Interestingly, mice hemizygous for Math1 die shortly after
birth from central apnea (Rose et al. 2009).
One of the overall goals of our research group is to
determine which genes on 18q contribute to the 18q-
phenotype through haploinsufficiency. This information
will eventually allow the molecular karyotype to be pre-
dictive with regard to the phenotype. The aim of this study
was to determine the extent to which hemizygosity of
TCF4 contributes to the physical and behavioral phenotype
in people with segmental 18q deletions.
Methods
Subject recruitment
Potential participant families learn about the research study
from a variety of sources. Primarily, however, families are
referred to the Research Center from the Chromosome 18
Registry & Research Society. Eligibility for the study
requires a cytogenetic or molecular diagnosis of an 18q
deletion. This study was approved by the Institutional
Review Board of the University of Texas Health Science
Center at San Antonio. All families were and continue to
be involved in the informed consent process, which is
appropriately documented.
Phenotypic assessment
For all families enrolling at the Chromosome 18 Clinical
Research Center, phenotypic data are compiled from three
sources. First, upon enrollment, families provide the
Research Center with extensive medical records to doc-
ument the participant’s medical and developmental his-
tories. These data are entered into a relational database
which is updated annually with the most recent medical
and developmental information obtained from the fami-
lies, providing longitudinal data on each of the study
participants. Second, all families are solicited annually by
mail to complete psychological surveys. Parents are asked
to complete the following questionnaires and return them
by mail: Behavior Assessment System for Children-Sec-
ond Edition (BASC-2; Reynolds and Kamphaus 2004)
which provides information regarding the presence of
behavior problems and emotional disturbance; the Vine-
land Adaptive Behavior Scales-Second Edition (Sparrow
et al. 2005) which asks parents to rate communication,
daily living and socialization skills and the Gilliam Aut-
ism Rating Scale-First Edition (GARS; Gilliam 1995) or
Second Edition (GARS-2; Gilliam 2006) which provides
an overall probability of autism rating.
The third data source is the comprehensive clinical
evaluation at the Chromosome 18 Clinical Research Center
as previously described (Cody et al. 2009).
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The evaluation completed at the Research Center
includes a comprehensive neuropsychological evaluation
comprised multiple measures. To obtain a measurement of
estimated cognitive functioning, an individually adminis-
tered measure of ability is given. If the participant is able to
understand the task demands to the extent that it is possible
to obtain a reliable and valid estimate of ability, then
measures based on the chronological age are given: Bayley
Scales of Infant and Toddler Development-Third Edition
(Bayley 2006); Differential Abilities Scales-Second
Edition (Elliot 2007) or the Wechsler Adult Intelligence
Scales-Third Edition (Wechsler 1997). If it is not possible
to evaluate the intellectual functioning using tasks and
activities based on their chronological age, it is necessary
to employ standardized measures which are routinely used
to assess the cognition, language, and motor abilities of
infants and toddlers. Because some participants are sig-
nificantly older than the normative comparison group for
these evaluations no standard comparison was possible.
Therefore, estimates of cognitive functioning are generated
from age equivalent scores.
To gain additional information about the group of par-
ticipants described in this project, it was necessary to
re-contact participating families to fill in gaps in the data
and to address new questions. All families participating in
this study were contacted by telephone to obtain additional
information about the participant’s respiratory history,
including abnormal breathing patterns.
Genotypic assessment
The DNA of all participants was evaluated using custom
designed oligonucleotide microarray comparative genomic
hybridization as previously described (Heard et al. 2009).
All participants in the study cohort assessed here had a
hemizygous region of 18q without other major chromo-
somal copy number changes.
Results
To determine the contribution of TCF4 hemizygosity to the
overall phenotype of 18q-, we compared the phenotypes
of those with and without hemizygosity of TCF4. We have
a cohort of over 200 individuals with hemizygosity for a
portion of chromosome 18q, each with a unique hemizy-
gous region (Heard et al. 2009). Within this cohort, there
are 27 individuals with a deletion that includes TCF4
(TCF4?/-). As shown in Fig. 1 and pictures in Fig. 2 (a–
m), this is a very heterogeneous group including 13 people
with terminal deletions between 24.67 and 30.71 Mb in
size (Fig. 1, Panel b), 8 people with small interstitial
deletions (between 5.6 and 16.88 Mb in size) and 6 with
large interstitial deletions (between 24.02 and 27.58 Mb in
size) (Fig. 1, Panel c).
Within our cohort, there are 132 individuals with ter-
minal deletions of 18q with breakpoints distal to the TCF4
gene (TCF4?/?). The comparison group was selected from
these individuals. We selected the TCF4?/? individuals
with the specific aim of creating a comparison group of
similar size to the TCF4?/- group. Because we had cog-
nitive data on 8 of the TCF4?/- individuals and adaptive
behavior data on 11, we selected nine TCF4?/? individuals
with breakpoints as close as possible to TCF4 to serve as a
control group. Parenthetically, the next smallest deletion
contained several more genes than other members of the
control group. The photographs of six of these individuals
are shown in Fig. 2n–s.
In this study, the TCF4?/? and TCF4?/- groups were
compared with regard to both physical and behavioral
phenotypes. The TCF4?/- group included 27 individuals:
12 males and 15 females. Since this is a longitudinal study,
the information was gathered over a period of time.
However, the age range at the most recent assessment was
10 months to 24 years 7 months, with an average age of
10 years 10 months. The TCF4?/? group of nine individ-
uals included four males and five females with an average
age range from 10 months to 28 years 7 months with an
average age of 13 years.
We aimed to (1) identify which features are found only
in the TCF4?/- group, and (2) determine the incidence of
features associated with the Pitt–Hopkins phenotype in
both the TCF4?/- group and the TCF4?/? group. The
physical features that distinguish the TCF4?/- group from
the TCF4?/? group are listed in Table 1 (Zweier et al.
2007, 2008; Pitt and Hopkins 1978; Rosenfeld et al. 2009;
Amiel et al. 2007; Andrieux et al. 2008; Brockschmidt
et al. 2007; de Pontual et al. 2009; Giurgea et al. 2008;
Kalscheuer et al. 2008; Peippo et al. 2006; Singh 1993;
Van Balkom et al. 1998). Only abnormalities of the corpus
callosum (64%) were found in more than half of the
TCF4?/- group. Other features were found in fewer than
35% of participants with TCF4 hemizygosity, yet were
unique to this group.
We then compared the non-unique phenotypic features
between the two groups. The features previously described
as associated with Pitt–Hopkins syndrome are indicated in
italics (Rosenfeld et al. 2009). These findings are shown in
Table 2 and reveal that the majority are non-specific find-
ings often associated with many different conditions
(Zweier et al. 2007, 2008; Pitt and Hopkins 1978; Rosen-
feld et al; Amiel et al. 2007; Andrieux et al. 2008;
Brockschmidt et al. 2007; de Pontual et al. 2009; Giurgea
et al. 2008; Kalscheuer et al. 2008; Peippo et al. 2006;
Singh 1993; Van Balkom et al. 1998; Taddeucci et al.
2010).
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To isolate the effect of TCF4 hemizygosity on cognitive
functioning, we wanted to compare the TCF4?/- group
with terminal deletions (N = 13) to the control group of
nine TCF4?/? individuals who also had terminal deletions.
However, we had cognitive data from in-person assess-
ments on only 8 of the 13 individuals in the TCF4?/-
group. Table 3 presents the estimated intellectual abilities
of these two groups by age. As Table 3 indicates, the
estimated intellectual abilities for the TCF4?/? group range
from mild intellectual disability to low average cognitive
functioning. This is quite mild in comparison to the eight
persons in the TCF4?/-group. In this group, cognitive and
motor development is very significantly delayed with
standard scores within the profound to severe range of
intellectual disability. In fact, the individuals in the
TCF4?/- group were unable to do even the easiest items of
each intellectual assessment instrument designed to assess
typical children under 1 year of age. Therefore, the most
informative indication of their cognitive function was to
report their age equivalent; the age at which the skills are
acquired in a typically developing child. These pronounced
deficits were present in children under 5 years of age and
continue to be present across the lifespan into child and
young adulthood. In general, the cognitive and motor
growth pattern is essentially flat across the lifespan
meaning that they did not acquire additional skills beyond
that of a 12 months old.
We also compared the language development of these
same two groups of individuals. All persons in the
TCF4?/- group are nonverbal with receptive language
limited to reactions to sounds in the environment, calming
when spoken to and recognition of caregiver’s voice with
Fig. 1 Panel a Chromosome 18 ideogram. The box indicates the
region of the chromosome shown in panels b–d. Panels b–d Display
the aCGH data from study participants with distal 18q deletions. The
light bars indicated the presence of the intact chromosome. The darkband at the end of the light bar indicated the breakpoint region. The
participant’s study number is to the left of their data. Panel
b Individuals with terminal deletions of 18q, who have one copy of
TCF4. Panel c Individuals with interstitial deletions of 18q who have
one copy of TCF4. Panel d The nine individuals in our study cohort
with terminal deletions and breakpoints closest to but not including
TCF4. These individuals phenotypic data were used as the compar-
ison group
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increased motor movement (wiggling). Expressive lan-
guage consisted of undifferentiated throaty or nasal sounds.
In contrast, in the TCF4?/? group, six of the nine persons
communicate verbally; one child uses sign language to
communicate; and two individuals were under the age of
two at the time of testing and were not yet speaking. These
two toddlers do however communicate through the use of
engaged eye contact, facial expressions, gestures and sub-
vocalizations.
Parents in both terminal deletion groups completed an
adaptive behavior questionnaire which compared the
communication, daily living, and socialization skill
development of their children with a normative group of
same-age peers. We had data on 11 of the 13 individuals in
the terminal deletion TCF4?/- group using this instrument
(Table 4). Parental ratings of both groups across all
domains were congruent with the intellectual assessment
results obtained through one-on-one assessment. The
adaptive behavior functioning of the TCF4?/- group was
rated as severely impaired while the behavioral functioning
of the TCF4?/? group was rated as falling within the mild
range of impairment.
Parents also completed the BASC-2 which provides an
evaluation of problem behaviors which fall into three
general areas: externalizing problems (hyperactivity,
aggressiveness, conduct problems); internalizing problems
(anxiety, depression, somatization) and those in the
behavior index [atypicality (disconnection from reality),
withdrawal and attention problems]. Only problems with
attention were rated by parents in both groups as cause for
concern (average T-Score = 63.18 for the TCF4?/- group
and average T-Score = 60.55 for the TCF4?/? group).
Average parental ratings across all of the other domains
were within normal or typical limits compared with same
age peers.
To determine the presence of behaviors consistent with
autism, parents evaluated their children’s communication
and social skill functioning along with the presence of
stereotyped or perseverative behaviors using one of the
Gilliam Autism Rating Scales (GARS or GARS-2). The
Fig. 2 Individuals with terminal deletions of 18q, who have one copy
of TCF4. a 3 years 9 months, b 6 years, c 6 years 8 months,
d 8 years 4 months, e 4 months 29 days, f 9 months, g 14 months
22 days. Individuals with interstitial deletions of 18q who have one
copy of TCF4. h 19 years 10 months, i 4 years, j 1 year 5 months,
k 4 years 2 months, l 2 years 8 months, m 1 year 10 months.
Individuals with terminal deletions and breakpoints closest to but
not including TCF4. n 25 years 9 months, o 14 years, p 7 years
5 months, q 8 years 9 months, r 6 years 6 months, s 1 year 2 months.
One individual with a small interstitial deletion that includes only the
TCF4 gene. t 12 years 8 months
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GARS is a screening tool and should not be used alone to
make a definitive diagnosis of autism. Therefore, scores
from this instrument are cataloged as to the probability of
having an autistic diagnosis. Parental ratings of children in
the TCF4?/- group indicated a very likely or high proba-
bility of autism being part of the diagnostic picture while
the ratings of parents of children in the TCF4?/? group
indicated that it was a possibility.
Within the group of TCF4?/-, children who had been
evaluated at the Research Center, there is great genotypic
variability between the individuals with regard to the
number of other genes involved in the deletion. Although
Table 1 Unique features of
those with TCF4 hemizygosity
Features in bold italics are those
in the literature as TCF4phenotypic componentsa Zweier et al. (2007, 2008);
Pitt and Hopkins (1978);
Rosenfeld et al. (2009); Amiel
et al. (2007); Andrieux et al.
(2008); Brockschmidt et al.
(2007); de Pontual et al. (2009);
Giurgea et al. (2008);
Kalscheuer et al. (2008); Peippo
et al. (2006); Singh (1993); Van
Balkom et al. (1998)
TCF4 phenotypic componentsa Hemizygous for TCF4
Number %
Abnormal corpus callosum 16/25 64
Atrial septal defect 7/20 35
Sacral dimple 7/23 30
Clubbed or broad fingers 3/11 27
Short neck 5/19 26
Hypertonia 7/27 26
Genital abnormalities-small penis 3/12 25
Camptodactyly of the fingers 5/22 23
Wide spaced nipples 5/22 23
Toe-2nd and 4th overlapping 3rd toe bilaterally 4/21 19
Premature death (\22 y/o) 5/27 18
Overfolding of the ears 4/22 18
Short philtrum 4/23 17
Malrotation of intestine 4/25 16
Cortical visual impairment 4/26 15
Absence or flattening of the superior fork of the antihelix 3/22 13
Optic atrophy 3/26 12
Accessory nipple 2/22 9
Wolff–Parkinson–White syndrome 2/25 8
Kyphosis 1/22 4
Table 2 Features NOT unique
to TCF4 hemizygosity
Features in bold italics are those
in the literature as TCF4phenotypic componentsa Zweier et al. (2007, 2008);
Pitt and Hopkins (1978);
Rosenfeld et al. (2009); Amiel
et al. (2007); Andrieux et al.
(2008); Brockschmidt et al.
(2007); de Pontual et al. (2009);
Giurgea et al. (2008);
Kalscheuer et al. (2008); Peippo
et al. (2006); Singh (1993); Van
Balkom et al. (1998); Taddeucci
et al. (2010)
TCF4 phenotypic componentsa Hemizygous for
TCF4Nine largest terminal deletions with two
copies of TCF4
Number % Number %
Hypotonia 26/26 100 9/9 100
Microcephaly 16/26 62 3/9 33
Postnatal growth retardation 15/27 55 1/9 11
Single palmar crease 12/22 54 1/9 11
Seizures 14/27 52 4/9 44
Myopia 14/27 52 3/9 33
Intra uterine growth retardation 13/25 52 1/9 11
Constipation 11/25 44 2/9 22
Central apnea 9/25 36 2/9 22
Drooling 7/25 28 3/9 33
Genital abnormalities-cryptorchidism 3/12 25 1/4 25
Strabismus 6/27 22 2/9 22
Scoliosis 2/14 14 1/9 11
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the behavioral and cognitive data presented in Tables 3 and
4 are from only those individuals with terminal deletions,
we wished to evaluate whether other regions of hemizy-
gosity of 18q had an impact on the effect of TCF4 hemi-
zygosity. For this analysis, we now included the cognitive
data from both the interstitial as well as the terminal
deletions in the TCF4?/- group. Again, these data were not
available on every individual whose genotype data are
shown in Fig. 1 and phenotype data shown in Tables 1 and
2. Because there are no other genes on 18q specifically
identified as haploinsufficient, the comparison between the
TCF4?/- subgroups was made based on grouping them by
the size of their deletion. We created three sub-groups:
those with terminal deletions (N = 13, 8 with cognitive
data), those with large interstitial deletions (N = 8, 5 with
cognitive data) and those with small interstitial deletions
(N = 6, 4 with cognitive data). The behavioral perfor-
mance of these three groups was compared to each other.
In addition, we have one participant, age 12 years and
8 months who has a small interstitial deletion of only the
TCF4 gene whose data have not been included elsewhere
in our analysis. Again, age equivalent scores were gener-
ated for the three groups because the use of standard peer-
based assessment measures was not possible due to the
participants’ very low cognitive and motor functioning. As
Table 5 indicates, the cognitive functioning of the three
groups is not significantly different. All three groups of
people hemizygous for the TCF4 gene have significantly
delayed cognitive and motor functioning. This suggests
that the size of the hemizygous region has little to no effect
on the developmental impact of the TCF4 gene. In essence,
the effect of TCF4 hemizygosity is so profound that, with
regard to development, children with large regions of
hemizygosity including many other genes are not more
developmentally delayed than children with hemizygosity
for the TCF4 gene alone.
One of the more striking findings was the realization
that, within our large cohort of people with simple terminal
18q deletions (N = 132), the majority of individuals who
died before the age of 18 had a deletion that included the
TCF4 gene. Figure 3 illustrates this point. The figure shows
either the current age or the age of death for the two
Table 3 Comparison of intellectual abilities
18q-, TCF4?/?
Intellectual
abilities
N = 9
CA less than
5 years
N = 3
CA between
6–12 years
N = 4
CA 13 years
and older
N = 2
Overall IQ and overall
range of scores
Full scale IQ 55a 63a 62a 61a
(50–82)
Verbal IQ 57a 66a 59a 63a
(50–81)
Nonverbal IQ 55a 75a 70a 69a
(50–90)
18q-, TCF4?/-
Intellectual abilities
N = 8
CA less than 5 years
N = 3
CA between 6–12 years
N = 4
CA 13 years and older
N = 1
Cognitive abilities AE = 1.3 months of age AE = 7 months of age AE = 6 months of age
Motor abilities AE = 2 months of age AE = 6 months of age AE = 6 months of age
CA chronological age, AE age equivalenta Average standard scores with a mean of 100 and a standard deviation of 15
Table 4 Average vineland
adaptive behavior scales-second
edition parental ratings
a Average standard scores with
a mean of 100 and a standard
deviation of 15
Type Communication Daily living
skills
Socialization Overall adaptive
behavior
TCF4?/? 60.22a 57.44a 61.22a 56.77a
N = 9
Terminal deletion, TCF4?/- 36.38a 36.84a 40.15a 36.46a
N = 11
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subgroups within our entire cohort; those with one copy of
TCF4 and those with two copies of TCF4. We investigated
the cause of death by reviewing medical records and
interviewing the parents. While a variety of reasons were
given for the cause of death, almost all of the children who
died had a history of multiple pneumonias, primarily
thought to be due to chronic aspiration, as is common in
such severely delayed children. This may have been
complicated by the breathing abnormalities that are com-
mon in the affected individuals.
Discussion
The analysis revealed three key observations. First, the
features unique to those with TCF4 hemizygosity were
abnormal corpus callosum, small penis, accessory nipples,
broad or clubbed fingers, sacral dimple, short neck and
wide spaced nipples. The potentially most clinically sig-
nificant of these physical findings is an abnormally thin or
absent corpus callosum. It would be reasonable to expect
that such an abnormality would be associated with an
inability to walk. However, none of the individuals with
TCF4 hemizygosity in our cohort were able to walk
regardless of the morphology of their corpus callosum.
This should not imply that all individuals with TCF4
deletions are never able to walk. There is anecdotal evi-
dence that some children with TCF4 are able to walk;
however, none of the individuals enrolled in this study had
attained that milestone.
Second, it could be anticipated that individuals with
larger deletions that include TCF4 might be more impaired
physically and mentally than those with smaller deletions.
However, we did not find this to be true. The presence or
absence of TCF4 seems to be more important in predicting
severity than the size of the deletion. We are fortunate to
have in this sample an individual with TCF4 hemizygosity
alone. As Table 5 illustrates, this person has significant
cognitive and motor delay. Given the very small number of
persons in each of our three sub-groups missing the TCF4
gene, it is not possible to determine if the differences in age
equivalent scores among these three deletion types are
statistically significant. Functionally, however, the cogni-
tive and motor delays across all groups are significant and
appear to be lifelong. In fact, in the cohort we evaluated,
TCF4 hemizygosity resulted in a developmental ceiling of
12 months irrespective of chronologic age, which ranged
from 10 to 238 months.
Third, the analysis of the ages and the age at death of
those with TCF4 hemizygosity and those with deletions of
chromosome 18 not including the TCF4 gene showed that
TCF4 hemizygosity conferred an increased risk of early
death. The cause of death was in most cases related to the
consequences of chronic aspiration.
The long-term goal of the Research Center is to deter-
mine which genes are responsible for which aspects of the
phenotype of 18q-. In this study, we analyzed the effect of
TCF4 hemizygosity to determine which components of the
Table 5 Average age equivalents of the three groups missing the TCF4 gene
Terminal deletion
N = 8
Large interstitial deletion
N = 5
Small interstitial deletion
N = 4
Deletion of only TCF4
N = 1
Cognitive abilities AE = 3.8 months of age AE = 2.4 months of age AE = 5 months of age AE = 11 months of age
Motor abilities AE = 3.4 months of age AE = 2.6 months of age AE = 6.25 months of age AE = 10 months of age
Fig. 3 Current age and age at death. The open circles indicate current
age and the black diamonds indicate the age at death. TCF4?/?; data
from 132 individuals with simple distal 18q deletions. Average
current age of those with two copies of TCF4 is 17 years. The two
individuals in this group who died were a female age 20 years,
6 months and a male 19 years, 9 months. TCF4?/- data from 22
subjects average current age is 11.5 years. Average age at death was
12 years old. The five participants who died included two males and
three females; ages 22 months, 6 years 10 months, 13 years, 20 years
11 months, and 22 years
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18q- phenotype can be attributed to the loss of one copy
of the gene. We compared the TCF4?/- group with the
TCF4?/? group using data from medical records, parental
questionnaires and in-person physical and behavioral
evaluations. Comparison to the literature was somewhat
hampered by the dearth of detailed phenotypic information
in previously reported cases of Pitt–Hopkins. Much of the
syndrome description is limited to dysmorphology, much
of which is imprecise, (e.g., ‘‘microcephaly’’), and medical
record abstraction. In particular, the behavioral and
developmental assessments are very limited in scope. This
made comparisons to the literature difficult. There is an
urgent need, now that the molecular genetics are known, to
perform a comprehensive multidisciplinary assessment of a
cohort of individuals with hemizygosity of or loss of
functions mutations in the TCF4 gene alone.
It is interesting to note that, in our population, the only
Pitt–Hopkins phenotypic components that were unique to
those with TCF4 hemizygosity were thin or absent corpus
callosum, small penis, accessory nipples, broad or clubbed
fingers, sacral dimple, short neck and wide spaced nip-
ples. Results of our analysis indicated that many of
the features of Pitt–Hopkins were present in both the
TCF4?/- as well as the TCF4?/? populations. Two of the
reported cardinal features of Pitt–Hopkins syndrome are
irregular breathing and seizures. The breathing abnor-
mality phenotype is poorly documented in the literature,
so we were not able to distinguish between documented
central apnea, episodes of heavy breathing and hyper-
ventilation. Interestingly, in our population this feature
was not unique to those with TCF4 hemizygosity. Like-
wise, seizures were not unique to the TCF4?/- group.
This is in part due to the fact that many of the features
such as hypotonia, single palmar crease, microcephaly,
and seizures are genetically heterogeneous and therefore
non-specific findings.
Alterations in the TCF4 gene have been implicated in
schizophrenia, and schizophrenia associated genes have
been linked to myelin-related pathways (Rietkerk et al.
2009). Since a key gene important in the compaction and
function of myelin (MBP) is located near the end of 18q,
and individuals with deletions of a critical region that
includes this gene have dysmyelination of the brain (Cody
et al. 2009), it might be postulated that hemizygosity of
both TCF4 and MBP would exacerbate their individual
effects. However, we did not see significant differences
between the TCF4 hemizygous participants whose 18q
deletion included the myelin basic protein gene (MBP)
(terminal deletions) and those that did not (interstitial
deletions) as shown in Table 5.
These data help us to make recommendations for this
unique group of individuals with 18q deletions. The cause
of death data highlights the need for aggressive detection
and intervention to prevent aspiration. Our data also
highlight the deleterious and chronic impact that hemizy-
gosity of the TCF4 gene has on cognitive and behavioral
development. In our study, all individuals hemizygous for
the TCF4 gene regardless of their age were similar to
typically developing babies less than 12 months old. It is
critical that parents plan for the long-term 24-h care their
children will need and refocus their developmental
expectations. It is important to provide a nurturing envi-
ronment that is rich in sensory stimulation and is one where
the child can feel comfortable and secure.
Of note, we are now faced with the challenge of
devising a meaningful nomenclature for 18q- that conveys
both the information about the genotype and as well as its
implications for phenotype. In this study, we used the
mouse nomenclature as a guide, since our goal is to merely
indicate gene copy number. We are indicating the diploid
state with regard to the TCF4 gene as TCF4?/? and the
hemizygous state as TCF4?/-. However, as we are able
to classify more genes as being either haplosufficient or
haploinsufficient, it will become a challenge for the
molecular cytogeneticist to write a clinically meaningful
karyotype. We do not envision a diagnostic code for
someone with a segmental deletion to include a long list of
all the hemizygous genes, but rather an edited list of those
genes that are haploinsufficient, i.e. have clinical signifi-
cance. Ultimately such a genotype would imply a particular
phenotype and thereby direct a plan of medical surveillance
and therapy.
Lastly, these data reinforce the need to eliminate the
word ‘‘syndrome’’ when referring to 18q- for two reasons.
First, rather than being defined by a constellation of fea-
tures, this condition is defined by a genotype, as implied by
the fact that it is named after the type of chromosome
aberration. Second, this particular chromosome abnormal-
ity is uniquely heterogeneous. No two unrelated individuals
have the same exact region of hemizygosity. Therefore, we
could identify numerous pairs of individuals who both have
18q- yet have no hemizygous genes in common (Heard
et al. 2009). The term ‘‘syndrome’’, which implies a col-
lection of similar phenotypic findings attributable to the
same genetic cause, is thus not appropriate. Rather, the data
presented here begin to define the molecular basis of 18q-,
highlighting the role of genomic heterogeneity in creating
phenotypic heterogeneity. Thus, the word ‘‘syndrome’’ is
no longer appropriate.
Acknowledgments The authors would like to first thank the fami-
lies that participated in this study for their willingness to share their
knowledge and for answering numerous questionnaires and emails.
This work was funded by the MacDonald family, The Chromosome
18 Registry & Research Society, the Institute for the Integration of
Medicine and Science (UL 1RR025767; National Center for Research
Resources) and CHRISTUS Santa Rosa Children’s Hospital.
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