Ron Trent

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Prenatal Diagnosis of the Haemoglobinopathies
Ron Trent Dept of Molecular Clinical
Genetics, RPA Hospital Dept of Medicine,
University of Sydney
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HAEMOGLOBINOPATHIES inherited disorders of
globin
divided into

• Thalassaemia Syndromes
• Variant Haemoglobins

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Some DifferencesBetween Thalassaemia Variants
Thalassaemia Variant Hb Family
history can be spontaneous (not HbE or
HbS) MCV low MCV normal (except HbE)
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(No Transcript)
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Classification of the Thalassaemias

• a thalassaemia
• b thalassaemia
• (db)o thalassaemia
• Hereditary persistence fetal Hb (HPFH)
• d thalassaemia

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Whatinheritancepatterns may be displayed by
thalassaemia ?
A

• Autosomal recessive (both parents are carriers 1
  in 4 risk to offspring) AKA autosomal
  co-dominant (carrier has low MCV)
• autosomal dominant ? thalassaemia (rare)
• compound states e.g. ? ?o thalassaemia HbH
  disease
• various combinations of ? and ? thalassaemias

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Pathogenesis Thalassaemia Syndromes

• a/b globin chain imbalance (N1)
• production, destruction
• iron overload
• hypersplenism
• complications blood transfusions
• gene/ gene interactions

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Diagnosis of b thalassaemia

• microcytic, hypochromic RBC (anaemia)
• blood film
• HbA2
• a/b biosynthesis ratio
• DNA analysis (point mutations)
• NB Family Studies

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b Thalassaemia
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Diagnosis of a thalassaemia

• normal haematology
• microcytic, hypochromic RBC (anaemia)
• blood film
• HbH inclusions
• a/b biosynthesis ratio
• DNA analysis (deletions)
• NB Family Studies

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a Thalassaemia (Normal 4 a Globin Genes)
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z2
yz1
ya1
a2
a1
a deletions
3.7 4.2
ao deletions
SEA
MEDIT
THAI
BRIT
SPAN
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• DNA testing
• Ultrasound
• Chromosome analysis
• Biochemical screen
• Fetal sampling (blood, other tissue)

Options for Prenatal Diagnosis
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• Chorion villus sample
• (CVS) 10/52 the AIM
• of PND is 1st Trimester CVS
• Amniocentesis 15/52. 2nd
• trimester test not ideal
• Fetal Blood 18/52 rarely
• done

Sources of Fetal DNA
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• Termination of pregnancy
• (to 20/52)
• Nothing, but appropriate
• follow up of newborn. NB
• ß globin defect shows up
• 6 months after birth.
• Possibility of treatment (e.g.
• blood transfusion, bone/stem
• cell transplant

Consequences of PND
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• Risk couple identified
• (MCV, MCH key parameters)
• Risk couples ethnic background
• important
• Are they related
• MOST IMPORTANT
• Do they want PND

Workup for PND involving Haemoglobinopathies
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• Hb Barts Hydrops Fetalis
• ( aa/-- in each partner)
• ß Thalassaemia Major
• (ßßT in each partner)
• HbH disease (asymptomatic to
• severe disorder difficult to predict)
• (aa/-- in one and aa/-a in other)
• (i.e. - a/- -)
• Various Hb variant combinations
• with ß Thalassaemia e.g. HbS, HbE

Indications for PND involving Haemoglobinopathies
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CVS only had IVS1,110 paternal and maternal
DNA samples showing typical mutations. No need
to test for maternal contamination. Child born,
and about 6 months of age shown to have ?
thalassaemia major CVS retested (after
re-extracted) and faint band in the CD41-42
position now detected. Problem allele-drop out
with PCR (probably suboptimal DNA and PCR
conditions marginal) Solution 2 different PCRs
now used for each prenatal diagnosis
CASE STUDY PCR ERROR
Pregnant couple (wife Chinese, husband Middle
Eastern) Mutations CD41-42 (-4) and the father
had typical IVS1,110
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• Essential to know what you are dealing
• with i.e. GOOD HAEMATOLOGY
• Couple need to be informed of potential
• consequences
• Problem in 2003 most will NOT have
• exposure to the underlying disorder
• Cultural and religious sensitivities
• Communication issues
• Medico-legal issues

Counselling Issues PND Haemoglobinopathies
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• Maternal blood
• (fetal cells, free fetal DNA)
• Preimplantation genetic
• diagnosis (PGD)

Other potential sources of Fetal DNA
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• Involves IVF
• Biopsy taken of dividing blastomere
• In theory useful for a wide number of
• genetic disorders
• Limited by technology (few cells are
• being studied)
• Requires confirmation by amniocentesis
• or CVS
• Sexing and simple genetic (DNA)
• tests feasible

Preimplantation Genetic Diagnosis (PGD)
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• Prefer CVS because good quality DNA
• 1st trimester diagnosis
• Amniocentesis may be only source
• available, especially in rural regions
• Fetal blood sampling with a / ß globin
• chain synthesis option no longer available
• PGD limited options
• Ultrasound with a thalassaemia
• (Hb Barts Hydrops Fetalis)

Practical Options for PND Haemoglobinopathies
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Prenatal Diagnosis for Thalassaemia
DNA challenge in a multicultural society
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Haemoglobinopathies and ethnic predisposition

• comes down to selection by malaria
• exception perhaps is British type ?o thalassaemia
  (??/--),
• and the Polynesians (lots of ? thalassaemia
  -?/??)
• at risk groups include
• ? thalassaemia SEA / Chinese / Mediterranean
• ? thalassaemia above Europeans, Middle East
  and Indian populations
• HbS Mediterranean, Middle Eastern and
  Black Africans
• HbE SEA

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Mediterranean (Greek, Italian) Chinese / SE
Asian Middle Eastern Indian subcontinent Unusua
l groups (Maldives, Eastern Europe)
RPA Experience with Ethnic Mix
Rare to get a referral from RNSH / Hunter
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Greek couple husbands mother thalassaemia
carrier Pregnant female, with classic ?
thalassaemia trait Partner MCV 66.2 fl (NR
80-100) HbA2. 1.8 (NR 1.5-3.5) HbF 1.3 (NR
lt1) target cells, no HbH inclusions (similar
results in 2 different laboratories) Interpretati
on Normal HbA2 ? thalassaemia or ?? thalassaemia
Case Study The Haematology Laboratory Is Still
The Best Start
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Family study (parents travelling overseas
mother said to have ? thalassaemia) Wifes ?
thalassaemia mutation IVS1,110 (common) CVS did
not have IVS1,110 Husband did not have 11 ?
thalassaemia mutations, before sequencing his
DNA, laboratory asked to review haematology
results
Case Study The Haematology Laboratory Is Still
The Best Start
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On review HbH inclusions bodies found DNA
testing of husband showed ?0 thalassaemia of
Mediterranean type (??/--) Haematology testing
of mother also showed HbH inclusions present with
a similar haematology profile to son (and a
brother)
Case Study The Haematology Laboratory Is Still
The Best Start
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What are the implications of 1 parentbeing a?
thalassaemia carrier,and the partnerhaving?o
thalassaemiatrait ?
Q
A
? / ? ratio is still close to 1 and so there
are no clinical consequences to offspring
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Q
A
? / ? is now going to be gt1. The
offspring could even have b thalassaemia intermedi
a-like picture.
What are the implications of 1 parenthaving
aaa/aa(i.e. there is an extraa globin gene)
the partnerhavingb thalassaemiatrait ?