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Genetics and inheritance issues in congenital heart disease
van Engelen, K.
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van Engelen, K. (2013). Genetics and inheritance issues in congenital heart disease
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Download date: 18 Jun 2017
Chapter 1
Introduction and outline of this thesis
1
10
Introduction
Congenital heart disease (CHD) is among the most common birth defects, occurring in approximately 8 per 1,000 live births.1 CHD comprises a wide range of cardiovascular malformations,
from complex and critical defects presenting prenatally or in the newborn period, to mild defects
that are not detected until adulthood. It leads to significant morbidity and mortality in children as
well as adults. Due to improvements in cardiac surgery and medical care, the population of adult
CHD patients is growing.2-4 Nowadays approximately 90% of CHD patients reach adulthood.
Most CHD occurs sporadically and in a non-syndromic fashion. In a subgroup of CHD a genetic
origin can be demonstrated, which include chromosomal abnormalities, copy number variations
(CNVs) and single gene defects. In the remaining cases there is significant heritability, which is
currently largely unexplained. The majority of non-syndromic CHD is historically believed to be
multifactorial in origin, i.e. multiple (unknown) genetic and environmental factors contributing to
the CHD. In the last decades, many research projects have focused on the genetic causes of
non-syndromic CHD and although significant progress has been made, the contributing genetic
defects remain largely unknown. The traditional approaches for gene discovery, including positional
cloning strategies, are hampered by the rarity of extended CHD families with clear Mendelian
inheritance patterns, incomplete penetrance and variable expression of the genetic defects. Studies
in animal models have elucidated many fundamental pathways involved in cardiac development
leading to the identification of candidate genes. Although several genes involved in human CHD
have been identified by screening these candidate genes in human patients, such candidate gene
studies are hypothesis driven - genes that seem to be less obvious in heart development at first
thus remain undiscovered. In addition, the non-coding regions of the genome have not been studied
extensively. Moreover, due to the heterogeneous genetic and clinical nature of CHD, large cohorts
of homogeneous CHD were difficult to realize in the past, hampering candidate gene studies or
case-control studies of common variants leading to CHD susceptibility.
Despite these pitfalls, there has been progress in the identification of genes and signaling pathways that are involved in cardiovascular development. Due to new and evolving genetic analysis
techniques, knowledge is currently expanding at an increasing pace. This introductory chapter
provides an overview of non-syndromic and syndromic causes of CHD. In addition, the implications of CHD genetics for patients and family members are discussed.
Non-syndromic CHD
In the majority of patients, CHD occurs in a non-syndromic fashion. It happens mostly sporadically,
although in some families a monogenetic inheritance pattern is present. A genetic cause can be
demonstrated in only a small subset of individuals and families with non-syndromic CHD. In some
patients, high-penetrance mutations in one of several genes that are known to be involved in heart
development can be identified (see below). In addition, more recently de novo CNVs have been
shown to contribute to several types of non-syndromic CHD, including tetralogy of Fallot (TOF),
left-sided heart defects (bicuspid aortic valve (BAV), aortic coarctation) and several other types of
11
1
1
CHD.5-7 In some of these CNVs known CHD genes are located; e.g. GATA4, which is located on
8p23.1, a region in which recurrent CNVs have been identified in CHD cases.6 It was estimated
that 5% to over 10% of sporadic, non-syndromic CHD occurs because of a rare CNV. Penetrance
is often incomplete.
Single gene defects and CNVs are found in only a minority of non-syndromic CHD, however.
The majority of non-syndromic CHD is historically thought to be due to multifactorial inheritance.
This implies that there is a cumulative effect of multiple variations in many different genes, each
of which contributes only for a small part to a person’s susceptibility to CHD. These genetic
susceptibility factors interact with each other as well as with environmental risk factors, which, if a
certain threshold is reached, together lead to CHD. Several of such susceptibility variants with low
penetrance have been identified through small-scale case-control studies, and recently the first
genome-wide association studies (GWAS) in CHD have shown associations for some CHD types.
Cordell et al. demonstrated two loci on 12q24 and 13q32 to be associated with TOF.8 In addition, a
recent GWAS study of cases with VSD, ASDII, transposition of the great arteries (TGA), conotruncal
malformations, and left-sided heart defects showed an association of single nucleotide polymorphisms in a region on chromosome 4p16 for ASD, but no significant associations were found for
the other phenotypes studied.9 A GWAS study in a Han Chinese population identified two risk loci
for CHD (including septal defects but also other CHD types) on 1p12 and 4q31.10 Odds ratios for
CHD risk in these studies ranged from 1.2 to 1.4. It seems remarkable that no consistent loci were
identified in these GWAS studies, however, differences in population ethnicity, patient characteristics and study design may explain these differences. Moreover, subcohorts with specific CHD
types might have been too small to detect associations.
Environmental factors that have been implied in CHD include maternal disease (e.g. diabetes
mellitus, hypercholesterolemia, hyperphenylalaninemia), infectious agents (maternal Rubella),
several medications (ACE inhibitors, retinoids) and substance abuse (alcohol, cocaine).11 Use
of folic acid during pregnancy has been hypothesized to be protective against CHD in the child,
and folate deficiency is suspected to be a CHD risk factor, however evidence for this association
remains inconclusive.11 Some common variants in genes encoding enzymes involved in folic acid
metabolism were shown to be ‘susceptibility alleles’ for CHD.12,13 A specific variation that has
been investigated extensively, is the common variant c.667C>T of the MTHFR gene, which
in homozygous state is known to cause lower levels of plasma folate. Interestingly, the largest
genetic study and meta-analysis performed so far showed no evidence for a relationship between
several subtypes of CHD and the MTHFR c.677C/T genotype (in CHD patients as well as mothers
of CHD patients).14
Non-syndromic monogenetic CHD
The proportion of non-syndromic CHD that has a monogenetic cause is not precisely known, but
it is presumed to be small. Only few families demonstrate CHD with a clear Mendelian inheritance
pattern. In these families, mostly autosomal dominant inheritance is seen. In some CHD families
12
as well as sporadic CHD, a pathogenic mutation underlying the defect can be identified. In these
cases, there usually is extensive genetic as well as clinical heterogeneity; i.e. a particular type of
CHD can be caused by mutations in different genes mutations, and mutations in one gene can
lead to distinct types and severity of CHD, even within a family. Moreover, penetrance is often incomplete.
The first reported single-gene mutation in humans with non-syndromic CHD was in the NKX2-5
gene, encoding a homeobox transcription factor.15 Since then, mutations in a substantial number
of other genes have been identified, mostly by positional cloning or candidate gene approaches.
Mutations have been found in several steps of the multiple pathways that contribute to heart
development, including genes coding for extracellular receptor ligands, membrane receptors and
transcription factors. Additionally, mutations in sarcomere protein genes and histone-modifying
genes have been identified in patients with (familial) CHD.16 Table 1 provides an overview of
a selection of the more well known genes involved in human CHD. A subset of these will be discussed in more detail below.
NKX2-5
NKX2-5 belongs to the NK-2 family of homeodomain-containing transcription factors, which are
conserved from flies to humans.17 Its role as a transcription regulator during early embryonic heart
developmental has been known for many years. Mice haplo-insufficient for nkx2.5 show abnormalities of the (atrial) septum and valve development as well as hypoplasia of the cardiac conduction system, especially the AV-node.18,19 Analogue to these abnormalities in animal studies,
in humans most NKX2-5 mutations have been reported in patients with (familial) atrial septal
defects (ASD) and conduction disorders, mainly atrioventricular block.15,20,21 Although NKX2-5
mutations can also lead to other types of CHD, the proportion of patients carrying such a mutation is lower in these groups.20,21 The overall mutation detection rate in sporadic CHD is reported
to be 2%.21 The mutations that have been identified are spread among the entire coding region of
the gene, without genotype-phenotype correlation. Most mutations (were predicted to) lead to a
truncated protein (haplo-insufficiency) or impaired protein function.
NOTCH1
NOTCH1 encodes a large, single-pass transmembrane receptor that functions in a highly conserved
cell-to-cell signaling system involved in multiple developmental processes.22,23 In mammals, there
are four Notch receptors (1-4), which interact with their ligands (Delta-like 1, 3 and 4, and Jagged
1 and 2) that are expressed on the surface of adjacent cells.23 Notch1-/- mice were shown to
have impaired trabeculation and hypoplastic cardiac cushions. In 2005, Garg et al. performed a
genome-wide linkage scan in a large family with bicuspid aortic valve, severe aortic calcification
and other CHD, including TOF. A single locus was identified on chromosome 9q34-35 and subsequently NOTCH1 was identified as the causal gene.24 Following this paper, mutations in NOTCH1
were also identified in patients with other left ventricular outflow tract obstruction, including hypoplastic
left heart syndrome and aortic coarctation.25 Functional studies of the mutations identified in
13
1
14
Activin receptor 1
Activin receptor 2B
2q24.1
3p22.2
ACVR1 (ALK2)
ACVR2B
Membrane receptors
Zinc finger protein of cerebellum 3
Xq26.3
T-BOX 5 transcription factor
12q24.1
TBX5
ZIC3
T-BOX 1 transcription factor
22q11.2
TBX1
T-BOX 20 transcription factor
Homeobox containing transcription factor
8p21.2
NKX2-6
Transcription factor AP-2
beta
Homeobox containing transcription factor
5q35.1
NKX2-5
7p14.2
Mediator complex subunit
13-like
12q24.2
MED13L
6p12.3
GATA-binding protein 6
18q11.2
GATA6
TBX20
GATA-binding protein 4
8p23.1
GATA4
TFAP2B
Friend of GATA2
Forkhead activin signal
transducer 1
8q23.1
8q24.3
FOG2 (ZFPM2)
CBP/p300 interacting transactivator with glu/asp rich
c-terminal domain
6q24.1
CITED2
FOXH1
Ankyrin repeat domaincontaining protein 1
Protein
10q23.3
Locus
ANKRD1
Transcription factors and regulators
Gene
Table 1. Genes in human non-syndromic CHD
Heterotaxy, TGA, PS, DORV
AVSD
TGA, heterotaxy, DORV, ASD, PS
PDA
ASD, VSD, CoA, PDA, MS
ASD, VSD, AVSD
IAA, VSD, TOF
PTA
TGA, heterotaxy, TOF
TGA
ASD, TOF, AVSD, DORV, PS, PA/
VSD, VSD
ASD, VSD, PS, TOF, HRV, PAPVR,
TGA, PTA
TOF, VSD, TGA
TOF, DORV
ASD, VSD
TAPVR
CHD
Char syndrome
Holt-Oram syndrome
22q11.2 deletion syndrome
Associated syndrome
102,105
104
88,102,103
36
99-101
97,98
34,35,96
95
88,92
94
92,93
32,70,91,92
88-90
87
86
85
References
1
15
Cryptic protein
Cysteine-rich protein with
EGF like domains 1
NOTCH1
Teratocarcinoma-derived
growth factor 1
2q21.1
3p25.3
9q34.3
3p21.3
CFC1
CRELD1
NOTCH1
TDGF1
Myosin heavy chain 6
Alpha myosin heavy chain
Beta myosin heavy chain
Myosin binding protein C3
14q11.2
14q11.2
16p13.11
11p11.2
MYH6
MYH7
MYH11
MYBPC3
ASD
PDA
EA, ASD, VSD, CoA
ASD, VSD, PS, TA, TGA
SVAS, PAS, AS, PS
ASD, VSD
Heterotaxy, HLH
BAV, subvalvular AS
BAV, CoA, AS
TVS, BAV, CoA, AS, VSD, PDA
TGA, heterotaxy, PA, TOF, AVSD
Heterotaxy, AVSD, CoA
TOF, PS
TOF, TGA, heterotaxy, AVSD
TOF, VSD
BAV, AS, HLH, CoA, VSD, TOF
AVSD, TGA, heterotaxy, CoA
TOF, TGA, heterotaxy, AVSD, BAV,
AVSD, PTA
AVSD, ASD, PDA, PAPVR + PAH
Williams syndrome
Oculodentodigital dysplasia
Alagille syndrome
122,129
128
126,127
29,31,32
58,124,125
122,123
120,121
119
118
116,117
88,115
114
27
112,113
89,111
24,25
54,109,110
89,91,107,108
106
CHD, congenital heart disease; TAPVR, total anomalous pulmonary venous return; AVSD, atrioventricular septal defect; BAV, bicuspid aortic valve; CoA, aortic coarctation; HLH,
hypoplastic left heart; ASD, atrial septal defect; VSD, ventricular septal defect; DORV, double outlet right ventricle; TGA, transposition of the great arteries; HRV, hypoplastic right ventricle; PAPVR, partially anomalous pulmonary venous return; PTA, persistent truncus arteriosus; PAH, pulmonary arterial hypertension; TVS, tricuspid valve stenosis; AS, aortic stenosis; IAA, interrupted aortic arch; TOF, tetralogy of Fallot; PA, pulmonary atresia; SVAS, supravalvar aortic stenosis; PAS, pulmonary artery stenosis; PS, pulmonary stenosis; PPH,
peripheral pulmonary hypoplasia; PDA, patent ductus arteriosus; EA, Ebstein anomaly; AD, autosomal dominant.
Alpha-cardiac actin 1
Elastin
15q14
7q11.23
ACTC1
Connexin 43
TAK1-binding protein 2
6q25.1
6q22.31
MAD-related protein 6
ELN
Structural molecules
GJA1
Gap junctions
TAB2
SMAD6
15q22.31
Vascular endothelial growth
factor
6p21.1
VEGF
Signaling molecules
Left-right determination factor
NODAL
1q42.12
10q22.1
LEFTY2
Jagged-1
10p12.2
JAG1
NODAL
Growth differentiation factor 1
19p13.11
GDF1
Extracellular ligands
Bone morphogenetic protein
receptor
2q33.1
BMPR2
1
1
human patients with CHD showed a reduction of the amount of receptor at the cell surface
as well as reduced binding of ligands to the receptor.25,26 Mutations in other components of the
Notch signaling pathway can also lead to syndromic and non-syndromic CHD. Mutations in JAG1,
encoding one of the Notch receptor ligands, cause Alagille syndrome characterized by liver
disease, dysmorphic features, vertebral abnormalities and CHD, specifically TOF. JAG1 mutations have also been identified in patients with non-syndromic CHD.27 In some patients with Alagille
syndrome, mutations in NOTCH2 are found.28
MYH6
In addition to mutations in transcription factors and receptor/ligand molecules, mutations in structural proteins can also lead to CHD. An example is the alpha-myosin heavy chain, a cardiac
sarcomeric protein, which is encoded by MYH6. It is expressed at high levels in the developing
atria, and studies in animal models showed that deficiency of MYH6 during heart development
leads to disturbed cardiogenesis and subsequent heart malformations.29,30 In 2005, a missense
mutation in MYH6 was shown to cause an autosomal dominant form of ASD with incomplete
penetrance.29 More recently, mutations were also identified in small proportions of patients with
other CHD, among which tricuspid atresia, VSD and pulmonary stenosis.31,32 These mutations
were shown to lead to disturbance of the assembly of normal myofibrils.31 Like mutations in other
sarcomeric protein genes, including MYH7 and MYBPC3, mutations in MYH6 have also been
identified in patients with hypertrophic and dilated cardiomyopathy.33
Syndromic CHD
There are numerous syndromes in which CHD may occur, and many of these are associated with
specific CHD types. These syndromes can be caused by chromosomal abnormalities, including
aneuploidies and structural aberrations. Moreover, mutations in genes involved in pathways that
are important in the development of multiple organ systems can lead to syndromic CHD. Interestingly, mutations in some of the genes involved in syndromic CHD have also been identified in
non-syndromic CHD patients, including TBX1, TFAP2B and JAG1.27,34-36 Table 2 provides an overview of some well-known syndromic forms of CHD. A few of these are discussed in more detail below.
Turner syndrome
Turner syndrome is caused by complete or partial monosomy for the X chromosome in all or
part of the cells. It occurs in 1 in 2500 female live births. However, the majority of
Turner syndrome conceptions do not survive until term. The main clinical features include fetal
lymphedema, webbed neck, short stature, gonadal dysgenesis and usually a normal intelligence,
with nonverbal learning disability. CHD is present in 20-40% of patients.37,38 However, in Turner
syndrome fetuses with cystic hygroma, who include the more severely affected patients with a
high mortality in utero, the prevalence of CHD is much higher. CHD in Turner syndrome mostly
comprises left-sided heart defects, including bicuspid aortic valve and aortic coarctation. Moreover,
16
patients are at increased risk of aortic aneurysm and dissection. An association between karyotype and CHD has been reported; frequency of CHD is higher in patients with 45,X than in those
with structural abnormalities of the X-chromosome (including X-chromosome deletions, ring chromosome X and isochromosome X), though a higher prevalence of bicuspid aortic valve in patients
with ring chromosome X was also reported.37,39 The genetic mechanisms that are implicated in
CHD in Turner syndrome are unknown.
22q11.2 microdeletion syndrome
The incidence of the 22q11.2 microdeletion syndrome is estimated to be at least one in 4,000,
making it the most common microdeletion syndrome in humans.40,41 Most patients have a specific
3 Mb deletion, but atypical deletions are also found. In past and present times, several diagnostic
terms have been assigned to the combination of features associated with microdeletion 22q11.2,
including velocardiofacial syndrome, DiGeorge syndrome and CATCH22. These terms represent
varying manifestations of the same genetic entity. The features associated with 22q11.2
microdeletion syndrome include CHD, cleft palate, velopharyngeal insufficiency with hypernasal
speech, hypocalcaemia, psychiatric disturbances, (mild) dysmorphic facial features and mild to
moderate mental retardation (reviewed in Kobrynski and Sullivan41). The presence and severity of
clinical features is highly variable, however. This can make it difficult to recognize the syndrome in
mildly affected patients, especially in adults.42,43 Three quarter of patients have CHD which typically
constitute conotruncal malformations such as interrupted aortic arch type B, truncus arteriosus
communis, TOF and pulmonary atresia (PA) with ventricular septal defect (VSD).42,44,45 The
frequency of 22q11.2 deletion in children with these particular heart defects ranges from about
10% (TOF) to up to 50% (interrupted aortic arch type B), and screening for the deletion is therefore
warranted in all patients with these CHD.
In over 90% of patients the deletion has arisen de novo, whereas in the remaining patients it
is inherited from one of the parents.46 Individuals with 22q11.2 microdeletion syndrome have a
50% chance of transmitting the deletion to their offspring. The 22q11.2 locus encompasses the
TBX1 gene, encoding a transcription factor of the T-box family. Mice with one or two disrupted
copies of TBX1 were shown to have CHD that are similar to those in 22q11.2 microdeletion
syndrome, implying that haplo-insufficiency of TBX1 is a major cause of heart defects in patients
with a 22q11.2 deletion.47 Indeed, mutations in TBX1 have also been identified in human patients
with CHD, with and without the other abnormalities seen in 22q11.2 microdeletion syndrome.34,48
More recently, CRKL was also identified as a possible CHD candidate gene residing in
the 22q11 locus.49,50
17
1
18
Monosomy X
Trisomy 13
Trisomy 18
Turner syndrome
Patau syndrome
Edward syndrome
AD (mostly de novo)
Deletion
7q11.23 (ELN)
Deletion 4p16.3
Deletion 5p15.2
Deletion 1p36
Williams syndrome
Wolf-Hirschhorn
syndrome
Cri-Du-Chat syndrome
1p36 Deletion
syndrome
AD (de novo)
AD (de novo)
AD (de novo)
AD (de novo >90%)
Deletion
22q11.2 (TBX1)
Sporadic/chromosomal
Sporadic/chromosomal
Sporadic/chromosomal
Sporadic/chromosomal
Inheritance pattern
22q11.2 Microdeletion syndrome
Structural chromosomal abnormalities
Trisomy 21
Genetic basis
Down syndrome
Aneuploidies
Syndrome
Table 2. Syndromic forms of CHD
PDA, NCCM
VSD, ASD, TOF
ASD, PS, VSD, TOF
SVAS, PAS
IAA type B, TOF, PA/VSD, aortic
arch abnormalities
40,41,130
55,131
132
133,134
135
Intellectual disability, multiple arterial stenosis, outgoing personality, distinct ‘elfin’
face, hypercalcemia
Intellectual disability, growth retardation,
central nervous system abnormalities,
skeletal abnormalities, ‘Greek helmet’ face
Severe intellectual disability, high-pitched
cry, microcephaly, micrognathia, facial
dysmorphism
Intellectual disability, microcephaly, eye
abnormalities, central nervous system
abnormalitues, distinct facial features
78
Growth retardation, clenched fingers,
distinct facial features, central nervous
system abnormalites
ASD, VSD, polyvalvular disease
Intellectual disability, velopharyngeal cleft/
insufficiency, psychiatric disorders, thymic
hypoplasia, hypocalcemia, mild facial
dysmorphism
78
Growth retardation, orofacial cleft, microphtalmia, central nervous system abnormalities, polydactyly
ASD, VSD, polyvalvular disease
78
Lymphedema, short stature, gonadal
dysgenesis, learning disability
BAV, CoA, HLH, ascending aorta
aneurysm
78
References
Intellectual disability, distinct facial features, hearing loss, early onset dementia
Extracardiac manifestations
AVSD
Cardiac disease
1
AD
AD
AD
AD
Single gene
defect -JAG1,
NOTCH2
Single gene
defect -CHD7
Single gene
defect -TFAP2B
Single gene
defect -MLL2
Alagille syndrome
CHARGE syndrome
Char syndrome
Kabuki syndrome
CoA, VSD, ASD
PDA
TOF, ASD, VSD
PPH, PS, TOF
ASD, VSD, conduction disorders
valvular PS, HCM
140
141,142
Distinct facial feaures, skeletal anomalies,
fetal finger pads, intellectual disability,
growth deficiency
138,139
Coloboma, choanal atresia, retarded
growth and development, genital abnormalities, ear anomalies
Distinct facial features, fifth finger anomalies
137
62,65
136
Intrahepatic bile duct paucity, skeletal abnormlities, posterior embryotoxon, distinct
facial features
Radial ray defects upper limbs
Short stature, distinct facial features,
pectus deformity, webbed neck
CHD, congenital heart disease; AVSD, atrioventricular septal defect; BAV, bicuspid aortic valve; CoA, aortic coarctation; HLH, hypoplastic left heart; ASD, atrial septal defect;
VSD, ventricular septal defect; IAA, interrupted aortic arch; TOF, tetralogy of Fallot; PA, pulmonary atresia; SVAS, supravalvar aortic stenosis; PAS, pulmonary artery
stenosis; PS, pulmonary stenosis; NCCM, noncompaction cardiomyopathy; HCM, hypertrophic cardiomyopathy; PPH, peripheral pulmonary hypoplasia; PDA, patent
ductus arteriosus; AD, autosomal dominant.
AD
Single gene
defect - TBX5
Holt-Oram syndrome
AD
Single gene
defect PTPN11, KRAS,
SOS1, RAF1,
BRAF, MEK1,
HRAS, NRAS,
SCHOC2, CBL
Noonan syndrome and other
syndromes with
disrupted RASsignaling
Monogenetic disorders
1
19
1
Down syndrome
Down syndrome, caused by an extra copy of chromosome 21, is the most common chromosomal
abnormality among live born infants and is the most frequent cause of intellectual disability. In over
95% of patients, there is a free copy of chromosome 21 in all cells and in most of the
remaining patients, a Robertsonian translocation involving chromosome 21 and another acrocentric chromosome leads to the Down syndrome phenotype. The syndrome is characterized by welldefined phenotypic features including characteristic facial features, growth retardation, hypotonia
and congenital malformations. About 50% of patients have CHD, most frequently atrioventricular
septal defect (AVSD) and VSD.51,52 A CHD “critical region” on chromosome 21 as well as several
candidate genes have been proposed, but the genetic basis and pathogenesis of CHD in Down
syndrome remain largely unknown.52 Mutations in ALK2 and CRELD1 have been identified in
some Down syndrome patients with AVSD.53,54
Williams syndrome
Williams syndrome is caused by a heterozygous deletion of circa 1.5 - 2 Mb on chromosome
7q11.23.55,56 Apart from CHD, which is present in 80% of patients, clinical features include intellectual
disability, characteristic facial features (‘elfin facies’), hypercalcemia, connective tissue abnormalities
and a friendly and outgoing personality. The most typical CHD in Williams syndrome comprise
supravalvar aortic stenosis (SVAS) and peripheral pulmonary artery stenosis, although other CHD
have also been described.57 SVAS is related to the absence of one copy of the Elastin (ELN)
gene which is located in the deleted locus. Mutations in ELN have been identified in patients with
non-syndromic SVAS and associated arteriopathies, although some individuals were described
to have a “Williams-like” mouth or connective tissue abnormalities such as inguinal hernias.58,59
Holt-Oram syndrome
Holt-Oram syndrome is an autosomal dominant ‘heart-hand syndrome’, occurring in about 1 in
100,000 individuals.60-62 Affected individuals show radial ray malformations, including an abnormal
carpal bone and absent, hypoplastic, or triphalangeal thumbs with or without radial dysplasia. The
limb abnormalities may be symmetrical or asymmetrical. Three quarter of patients have CHD,
most commonly ostium secundum ASD and VSD, as well as progressive conduction
disease.61,62 Although rare, other CHD have also been reported.63,64 Lower limb defects or other
extracardiac malformations or disease (including intellectual disability) do not belong to Holt-Oram
syndrome, thus if these are present Holt-Oram syndrome is unlikely.
In 1997, TBX5 was identified as the causal gene in Holt-Oram syndrome.65,66 TBX5 functions as
a transcription factor that has an important role in cardiac growth and development (especially in
cardiac septation), in the development of a cardiac conduction system.67,68 and in forelimb specification and outgrowth.69 TBX5 can interact with other transcription factors including NKX2-5 and
GATA4.70 A pathogenic mutation in TBX5 can be identified in over 70% of patients, mostly but not
always through the mechanism of TBX5 haplo-insufficiency. About 85% of affected individuals
20
have been reported to have Holt-Oram syndrome as the result of a de novo mutation. In a small
number of patients with Holt-Oram syndrome, mutations in SALL4 have been demonstrated.71
CHD recurrence risks
In only a minority of CHD patients, it is possible to provide a CHD recurrence risk based on known
Mendelian inheritance in a family or on figures related to chromosomal abnormalities. When a (genetic) cause is not identified in an individual non-syndromic patient, empirical estimates have to be
used to provide recurrence risk numbers. Several studies have shown that there is an increased
risk of CHD for relatives of CHD patients, mostly in the order of magnitude of 2 to 15% for firstdegree relatives of a sporadic patient.72-74 A recent population-based study in Denmark demonstrated that the relative risk for all types of CHD taken together is 3.2 among first degree relatives
and 1.8 among second degree relatives.75 Risk numbers vary significantly between different types of
CHD, however. For example, the heritability of heterotaxy as well as BAV and other left ventricular
outflow tract lesions is estimated to be much higher.75-77 Within families, the relative risk is highest
for same-type CHD, but the risk is also increased for dissimilar types.75 If more than one family member is affected, the recurrence risk for other relatives increases. Moreover, the risk for offspring of
women with CHD is higher than for offspring of men with CHD72,73; generally the risk for offspring
of female CHD patients is estimated to be about 5-6%, whereas the risk for offspring of male CHD
patients is about 2-3%.78 The reasons for this gender difference are unknown; imprinting mechanisms or maternal and environmental factors may play a role. Sibling recurrence risks are generally
stated to be 2-3%. If two siblings are affected, the recurrence risk increases to 10%.78
Genetic counseling in adult CHD
The population of adult CHD patients is growing, and an increasing number of patients will have
children. These patients may have questions regarding the origin and inheritance of their disease
and the recurrence risk in (future) offspring. For the individual patient, knowledge about the
underlying (genetic) origin of CHD is important because 1) the patient and his or her offspring may
be at risk for extracardiac disease (in syndromic cases); 2) an individualized recurrence risk for
offspring based on the underlying cause can be established; 3) there may be other relatives for
whom genetic or cardiologic examination may be appropriate.79,80
The majority of adults with CHD has not had genetic testing or counseling, and those who did
in childhood were probably too young to participate in the counseling process. In general, the
knowledge of adult CHD patients about inheritance issues, including pregnancy-related topics and
recurrence risk in offspring, has been shown to be low.81-84 Clinical genetic consultation can aid in
increasing awareness and knowledge about such issues. It may help patients to understand the
genetic basis of their CHD (and extracardiac disease), and the implications for family members
including their offspring.
By means of a detailed medical history of cardiac and extracardiac disease, extensive family
history and physical examination aimed at dysmorphic features the clinical geneticist can gather
21
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information to establish a strategy for genetic testing and come to an etiological diagnosis in an
individual patient. Subsequently, recurrence risks targeted to the patient’s specific situation can be
provided as accurate as possible. In addition to the establishment of an etiological diagnosis and
the estimation of a recurrence risk, genetic counseling involves education about management (of
cardiac and extracardiac manifestations), inheritance issues and familial implications. Reproductive options, including prenatal ultrasound investigations, prenatal diagnosis and pre-implantation
diagnosis, may also be discussed. Moreover, the psychological implications of the diagnosis and
recurrence risk are addressed.
As CHD is not uncommon and mostly multifactorial in origin with general recurrence risks applying,
at current times it seems not eligible to counsel all adult CHD patients. It is however important to
identify the subgroup of patients that is especially likely to benefit from genetic counseling. These
patients are summed in Table 3. Physicians caring for adult CHD patients should actively identify
those patients and refer them at a low threshold.
Table 3. Adult CHD patients who may benefit from clinical genetic testing and counseling
• Patients desiring having children (preconceptional counseling)
• Patients with extracardiac abnormalities/disease
- Congenital malformations
- Intellectual disability
- Dysmorphic features
- Multisystem involvement (e.g. endocrinologic, hematologic, immunologic or sensorineural disorders)
- Psychiatric disorders
• Patients with CHD with high risk of 22q11.2 deletion (IAA, TA, TOF, PA with VSD, AAA)
• Patients with a family history of CHD
• Any other patient who is interested in learning about the origin and inheritance of their CHD
CHD, congenital heart disease; IAA, interrupted aortic arch; TA, truncus arteriosus; TOF, tetralogy of Fallot; PA, pulmonary atresia; VSD, ventricular septal defect; AAA, aortic arch anomaly.
22
Outline of this thesis
This thesis focuses on the genetics of non-syndromic and syndromic CHD, the implications it has
for (adult) CHD patients and the adult CHD patients’ perspective on inheritance issues. It presents
different types of studies, including clinical and genetic studies in families with multiple affected
individuals with CHD and in patient cohorts with specific CHD subtypes, a morphologic study of
post-mortem hearts, as well as questionnaire studies.
In Part I of this thesis, clinical and genetic studies in non-syndromic CHD are presented. These
studies were aimed to the identification of genes that are implied in human CHD. Chapter 2
describes the linkage analysis and subsequent identification of a disease locus in a large family
with a novel phenotype, which resembles a mild form of left atrial isomerism. Chapter 3 presents
the analysis of the sarcomere protein gene MYH7 in a cohort of patients with Ebstein anomaly. In
Chapter 4, the role of mutations in the cardiac sodium channel gene SCN5A in CHD is explored;
on the one hand, a cohort of patients with a septal defect and conduction disease was analyzed
for SCN5A mutations. On the other hand, a cohort of SCN5A mutation carriers was evaluated for
the frequency of CHD.
Part II focuses on CHD in association with other (extracardiac) abnormalities, including syndromic
CHD. Chapter 5 is dedicated to the 22q11.2 deletion syndrome. Although it is currently common
practice to test children with specific CHD types for the presence of a 22q11.2 deletion, this has not
routinely been performed by patients who have already reached adult age. We therefore evaluated the prevalence of 22q11.2 microdeletion syndrome in adults with TOF and pulmonary atresia
with VSD, which is described in chapter 5. In Chapter 6, we focus on CHD in Turner syndrome; in
this syndrome left-sided heart defects including aortic valve abnormalities are especially common.
We specifically looked at the morphology of the (bicuspid) aortic valve early in development in a
selected group of TS patients with adverse outcome, namely in post-mortem heart specimens of
Turner syndrome fetuses.
Also in the absence of a recognized syndromic etiology, CHD is associated with several extracardiac
malformations or disease. As common developmental pathways may underlie these associations,
genes implied associated abnormalities may also be implied in cardiac development and CHD,
and vice versa. One of the genes that are already well-established in CHD is NKX2-5. More
recently, NKX2-5 was also implicated in human thyroid dysgenesis. In Chapter 7 we studied a
specific variant in NKX2-5, which we encountered in two unrelated probands with CHD and which
had been identified before in a patient with thyroid dysgenesis. Chapter 8 focuses on childhood
cancer: we evaluated whether CHD is more frequent in children with neuroblastoma, as was
previously reported.
Whereas Part I and II mainly focus on genetic causes and pathogenetic mechanisms leading to
CHD, Part III appraises the clinical implications and patients’ perspectives on genetics and inheritance
23
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of CHD. Chapter 9 reports on the information adult CHD recalled to have received about the
inheritance of their CHD, patients’ knowledge about inheritance and their concerns in this regard.
Chapter 10 focuses on adults with CHD who consulted a clinical geneticist. We report on the etiologic diagnoses that are made by the geneticist in these patients. Additionally, patient satisfaction
with genetic counseling and reproductive choices are presented.
Finally, in Chapter 11, the major findings presented in this thesis are summarized and future
perspectives for research and clinical care are discussed.
24
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