FISH and Chips in Developmental Pediatrics

1
FISH and Chips in Developmental Pediatrics

• Dr. Victoria Mok Siu
• M.D., FRCPC, FCCMG
• Paediatrician and Medical Geneticist

2
Objectives

• Understand the use of genetic diagnostic tests,
  specifically FISH and CGH
• Recognize dysmorphic and behavioral patterns
  associated with genetic diagnoses
• Be able to access genetic databases for diagnosis
  and management

3
Where to go for help
4
www.genetests.org
5
(No Transcript)
6
(No Transcript)
7
(No Transcript)
8
(No Transcript)
9
(No Transcript)
10
OMIM
11
(No Transcript)
12
Investigation of a non-dysmorphic child with
autism/developmental delay

• First tier investigations
• Hearing assessment
• Verify newborn biochemical screening results or
  assess for the possibility of a metabolic
  disorder
• TSH
• Lactate, ammonia, amino acids
• Urine screen MPS, organic acids, amino acids
• Chromosomes (likely to become microarray testing
  when more widely available)
• DNA testing for Fragile X syndrome
•  
• Second tier investigations
• PTEN molecular testing, if macrocephalic
• MECP2 testing, if female
• Microarray analysis
• MRI of brain

13
When to go FISHing

• routine G banded karyotype
• FISH
• telomere probes

14
Giemsa-banded karyotype
15
FISH - region-specific probe
Can detect specific microdeletion syndromes Must
specify which probe is to be used
16
22q microdeletion syndrome

• Congenital heart defect
• Cleft palate nasopharyngeal regurgitation
• Immunodeficiency
• Short palpebral fissures

17
Williams syndrome 7q

• Stellate iris
• Periorbital puffiness
• Thick lips
• Long philtrum
• Cocktail party chatter
• Hyperacusis
• SVAS

18
Williams Syndrome Are people different from
shapes?
Am J Ment Retard. 2001 10694-107
Exp Brain Res. 2004 157137-46
19
Smith-Magenis syndrome 17p11.2

• Polyembolokoilomania (foreign objects in all
  orifices)
• Self-hug
• Limited speech

20
Smith-Magenis syndrome 17q
www.prisms.org
21
Disturbance of circadian rhythm in Smith-Magenis
syndrome
Treatment beta-blocker and melatonin
22
Prader-Willi syndrome 15q

• Initial FTT and hypotonia
• Thick saliva
• Pain insensitivity
• Good with puzzles
• Almond-shaped eyes
• Onset of obesity around 2 to 3 years

www.pwsausa.org
23
Angelman syndrome 15q

• Acquired microcephaly
• Absent speech
• High-stepping gait
• Seizures by 2-3 yr
• prognathism

www.angelman.org
24
Both Prader-Willi syndrome and Angelman syndrome
may be caused by a 15q11-13 microdeletion. These
conditions involve imprinting.
http//www.mgm.ufl.edu/images/driscoll/Fish20pic
20Gradient.gif
25
Genomic Imprinting an epigenetic phenomenon

• Epigenetic modifications reversible changes of
  DNA methylation pattern and chromatin structure
  that can affect gene expression
• Genomic imprinting differential expression of a
  gene, depending on which parent it was inherited
  from
• (Reproductive Health 200416)

26
Genetics of PWS and AS

• The genes associated with Prader-Willi syndrome
  (SNRPN) and Angelman syndrome (UBE3A) are located
  in the same region of chromosome 15

SNRPN UBE3A
SNRPN UBE3A
27
Differential expression of the PWS and AS genes
In a normal individual
paternal
maternal
SNRPN UBE3A
SNRPN UBE3A
methylated
methylated
28
Chromosomal deletion of 15q11-13

• Angelman syndromedeletion on 15 inherited from
  mother

• Prader-Willi syndromedeletion on 15 inherited
  from father

SNRPN UBE3A
SNRPN UBE3A
29
Chromosomal deletion in PWS and AS

• Usually de novo, with parental chromosomes being
  normal
• Different phenotypes depend on parental origin of
  the deleted chromosome

30
Sometimes with PWS or Angelman syndrome, FISH for
15q microdeletion is normal
31
Uniparental disomy a pair of chromosomes
inherited from the same parent

• Prader-Willi syndromeboth 15 chromosomes from
  mother

• Angelman syndromeboth 15 chromosomes
  inherited from father

SNRPN UBE3A
SNRPN UBE3A
SNRPN UBE3A
SNRPN UBE3A
32
Strategy for testing for PWS and AS

• Methylation studies to determine if alleles from
  both parents are present
• If abnormal methylation, do FISH to differentiate
  between microdeletion and UPD
• (if you do only FISH, you will miss 30 of cases
  of PWS or AS)

33
Rett syndrome

• Acquired microcephaly
• Developmental plateau/regression
• Seizures
• Handwringing
• Hyperventilation
• Abdominal bloating
• Bruxism

34
Rett syndrome is another epigenetic disorder

• The normal MeCP2 protein is involved in
  maintaining DNA methylation (ie inhibiting gene
  expression)
• With a mutation in MeCP2, too many genes remain
  activated (like an orchestra where all the
  instruments are playing at once)

35
Overlap between symptoms of Angelman syndrome and
Rett syndrome

• Acquired microcephaly
• Seizures
• Lack of speech

36
Multiple subtelomeric FISH probes

• Used to detect subtelomeric deletions
• Not useful for duplications or translocations
  unless there is an accompanying deletion

37
Microarray CGH

• To detect variation in copy numberie. small
  deletions or duplications
• Need to compare with parental DNA

www.molgen.mpg.de
38
duplication in Xp11-p21.1
http//www.genetik-osnabrueck.de/english_aCGH.html
39
Other behaviors to watch for
40
Diet and eating habits

• Low protein
• urea cycle defect, aminoacidopathy
• Avoidance of fruits
• fructose intolerance
• Nasopharyngeal reflux
• 22q deletion

41
Sleeping habits

• Sleep apnea
• Prader-Willi syndrome
• Altered circadian rhythm
• Smith-Magenis syndrome

42
Speech patterns

• Echolalia
• Fragile-X syndrome
• cocktail party chatter
• Williams syndrome

43
Behavior with illnesses

• Undue lethargy and vomiting, or unusual odour of
  urine or sweat mousey PKU
• sweaty feet isovaleric acidemia
• sweet MSUD

44
Whats new in gene therapy?
45
Normal Duchenne muscular dystrophy
Dystrophin stain
46
Dystrophin protein is involved in DMD and BMD
47
Potential treatments for DMD

• Utrophin upregulationturn on another gene to
  make a protein similar to dystrophin
• Myostatin inactivationturn off a gene to
  enhance muscle growth

48
Myostatin

• Normally acts to inhibit muscle cell growth
• Inactivation of myostatin may result in muscle
  growth
• Use of myostatin inhibitors is being investigated
  for a wide range of disorders, including muscular
  dystrophies and muscle wasting associated with
  HIV