Clinical epigenetics and its relation to ART

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Clinical epigenetics and its relation to ART

• M. De Rycke
• Centre for Medical Genetics
• UZ Brussel - VUB

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Epigenetics and ART

• What is epigenetics?
• Genomic imprinting
• Review data on imprinting disorders in ART
  children

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What is epigenetics?
Chromatin structure DNA and histones
Chromatin structure histones and DNA
-

-

-

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What is epigenetics?
gene expression
gene silencing
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What is epigenetics?
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What is epigenetics?
euchromatin
heterochromatin
gene expression
gene silencing
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What is epigenetics?

• epigenetics
• study of changes in gene expression that occur
  without changes in the DNA code
• gene expression
• patterns

inheritable through mitosis and meiosis
changes during cell differentiation
influence by environmental factors
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Epigenetic galaxy
genotype space
G
G
Z
Z
genotype space
early embryo
gametogenesis
epigenesis
A
A
phenotype space
J
J
phenotype space
Epigenetic variation
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Genomic imprinting
Paternal copy is expressed
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Genomic imprinting

• Imprinted genes monoallelic expression
• gt 80 imprinted genes (0.1-1 of all genes)
• Key role in embryonic growth and placental
  function, cognition and maternal behaviour
• Defective imprinting involved in carcinogenesis
  and in human genetic diseases (Angelman and
  Beckwith-Wiedemann syndrome)
• Maternal and paternal alleles carry different
  epigenetic modifications (imprints)

paternal
open chromatin
maternal
compact chromatin
no DNA methylation
DNA methylation
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Imprinting
somatic cell line
maintenance and monoallelic expression
p
m
gametogenesis
imprint resetting
embryo
maintenance and monoallelic
expression
early embryonic development imprint maintenance
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Angelman syndrome
- Incidence 1/10000 - 1/30000
- Neurogenetic disorder - severe mental
retardation - ataxia - happy puppet
syndrome - absence of speech
- Caused by genetic or epigenetic defects in an
imprinted region on chr15q11-13, gt loss of UBE3A
expression
- Inheritance most cases are sporadic - 85-90
genetic defects - lt 5 epigenetic defects
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ART and AS
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ART and AS

• Ludwig et al., 2005 correlation between
  infertility treatment and epigenetic defects
  (cohort 79 AS patients)

RR relative risk HT hormone treatment LOM
loss of maternal methylation TTP time to
pregnancy
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Beckwith-Wiedemann syndrome
- Incidence 1/15000
- Overgrowth syndrome with predisposition for
embryonal tumours
- Caused by genetic or epigenetic defects in
an imprinted region on chr11p15
- Inheritance - 15 familial cases, autosomal
dominant
- 85 sporadic cases - 20 genetic (pUPD,
mutations) - 60 epigenetic (aberrant
methylation) - 20 unknown
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ART and BWS
RR relative risk LOM loss of maternal
methylation
overlapping patients
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ART and BWS

• epidemiological data
• molecular analysis an epigenetic defect (loss of
  maternal methylation at BWSIC2) in 24/25 BWS
  patients after ART
• general population epigenetic defect in 50 of
  cases

Association between ART and BWS to be considered
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ART and imprinting disorders

• Other reports on imprinting disorders after ART
• no higher incidence of ART in other imprinting
  registers
• gt only higher incidence of AS and BWS after ART

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Iceberg theory
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Iceberg theory

• Moll et al., 2003 link between ART and
  Retinoblastoma
• Risk Ratio 4.9 (1.6-11.3) RR 7.2 (2.4-17.0)
• No higher risk of childhood cancer in 4 studies
• Doyle et al. 1998, (UK n 2057), Bruinsma et al.
  2000, (Australia, n 5249), Klip et al. 2001,
  (the Netherlands, n 9484), Lerner-Geva et al.
  2000, (Israel n 332)

• Lidegaard et al., 2005
• systematic follow up of 6052 ART singletons
  (1995-2001) till 2002 vs a control group of
  442.349 non-ART
• the incidence rate of childhood cancer, mental
  diseases, congenital syndromes and developmental
  disturbances was equal in the two groupsgt no
  increased risk for imprinting diseases after ART

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Iceberg theory

• Rossignol et al., 2006abnormal methylation
  patterns at other imprinted regions than BWS
  region for some BWS patients with an epigenetic
  defect (11 ART, 29 naturally conc) gt mosaic
  patterns suggest failure in methylation
  maintenance in the early embryos

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Cause of imprinting disorders after ART

• causes? both IVF and ICSI gt common element
  may interfere with
• imprint maintenance during embryo culture
• imprint resetting during gametogenesis
• in-vitro culture systems
• hormonal stimulation
• parental infertility

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Cause of imprinting disorders after ART

• Khosla et al. 2001 culture (serum) of
  preimplantation mouse embryos reduced fetal
  development (lower birthweight) and expression of
  imprinted genes
• Young et al. 1998 after exposure in vitro
  unusually large offspring syndrome (LOS) in
  relation with imprinted genes
• Young et al. 2000 epigenetic changes in IGF2R
  are associated with fetal overgrowth (LOS) after
  sheep embryo culture

The mammalian preimplantation embryo is very
sensitive to culture conditions epigenetic
changes induced at the early stages may lead to
altered phenotypes at later stages
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Cause of imprinting disorders after ART
use of hormonal ovarian stimulation
hormonal stimulation may interfere with
epigenetic reprogramming and imprint resetting
during oocyte development
gt epigenetic deregulation in embryos from
superovulated mice compared to embryos from
naturally ovulated mice Shi and Haaf, 2002
gt retrospective case analysis of mothers (12) of
BWS children after ART review of the
reproductive endocrine records gt hormonal
stimulation was the only common factor Chang et
al., 2005
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Cause of imprinting disorders after ART
parental infertility
subfertile couples are predisposed to epigenetic
defects imprinting defects and subfertility have
a common cause
gt Incomplete establishment of imprints
(incomplete methylation of H19) in spermatozoa
from men with fertility problems Marques et
al., 2004
gt AS cohort study subfertile couples have
an increased risk of conceiving a child with an
imprinting defect TTPgt 2 years, no therapy RR
6.25 TTPgt 2 years, treatment RR 12.5 Ludwig et
al., 2005
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Conclusion and perspectives (1)

• (limited) evidence for a higher risk of
  imprinting disorders after ART
• Absolute risk still low
• Time for studies of larger series of children
  since rare events have to be studied
• Prospective multi-centre studies with control
  groups

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Conclusion and perspectives (2)

• Need for long-term follow up
• birth defects and neonatal outcome
• imprinting disorders, cancer, neurobehavioural
  development
• Record data on infertility history, hormonal
  stimulation, in vitro culture conditions
• Need for basic epigenetic and imprinting research
  
• in animal models
• human gametes and embryos

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ART and BWS

• Chang et al., 2005 review of reproductive
  endocrine records
• 12 BWS children after ART (10 IVF, 2 ovarian
  stimulation)
• Analysis of treatment type all had some type of
  ovarian stimulation
• No increased risk due to
• specific type of culture medium
• type of stimulation
• type of ART
• type of infertility

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ART and BWS

• Remarks on studies
• Methodological errors?
• Lack of appropriate controls
• Underestimation of ART history in the syndrome
  registers
• Underestimation of the ART frequency in the
  general population
• Strict diagnosis ?
• Enhanced awareness since case reports
• Statistical variation?
• Clustering , large confidence intervals in small
  studies

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Cause of imprinting disorders after ART
use of in-vitro culture systems
culture and manipulation of gametes and embryos
during ART would induce epigenetic changes
imprinted genes vulnerable
may interfere with - imprint maintenance during
embryo culture - imprint resetting during IVM
(GV to MII)
gt Evidence from animal studies
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Human studies

• Analysis of human epidemiological data
• data on Angelman syndrome
• data on Beckwith-Wiedemann syndrome