GENETIC DISORDER OF GENETIC DISORDER OF HAEMOGLOBINHAEMOGLOBIN

Haemoglobinopathies and Thalassaemias

1

A conjugated protein consisting of iron-containing heme and protein (globin)Globin chains are of different types: -chains and non -chains Each molecule is a tetramer of two - and non chains. Each globin binds a haem in a haem binding site.Haemoglobin binds and transports oxygen from lungs to the tissues, while it transports CO2 from tissues to the lungs.

A conjugated protein consisting of iron-containing heme and protein (globin)Globin chains are of different types: -chains and non -chains Each molecule is a tetramer of two - and non chains. Each globin binds a haem in a haem binding site.Haemoglobin binds and transports oxygen from lungs to the tissues, while it transports CO2 from tissues to the lungs.

Haemoglobin

2

3

Types of Hemoglobin in adults

Globin genes Gene product Tetramers Name of Conc. in

Chromosome (globin) in RBCshaemoglobin adult

16 11

, -chain 2 2 Hb A 96-97

, -chain 2 2 Hb A2 2.3-3.5

,-chain 2 2 Hb F <1.0

----------------------------------------------------------------- 4

Chromosome 11

AG

2 1 2 1

Chromosome 16

Structure of each Globin gene

Exon 1Intron 1Exon 2 Intron 2 Exon 3

5’ 3’

3’

3’5’

5’

5

βδγα α

Chromosome 16 Chromosome 11

25% 25%

α α βδγ

25% 25% 48%

48%

1.5%0.5%

1.5%0.5%

HEMOGLOBIN SYNTHESIS

6

HEMOGLOBINOPATHY; DEFINITION

An inherited mutation of the globin genes leading to a qualitative or quantitative abnormality of globin synthesis.

7

STRUCTURAL STRUCTURAL HEMOGLOBINOPATHYHEMOGLOBINOPATHY

Amino acid substitution in the globin chain e.g. sickle hemoglobin (HbS)

8

THE THE THALASSEMIASTHALASSEMIAS

Syndromes in which the rate of synthesis of a globin chain is reduced:-

beta thalassemia – reduced beta chain synthesis

alpha thalassemia – reduced alpha chain synthesis

9

Disorders of HaemoglobinDisorders of Haemoglobin

Haemoglobinopathies(Structural disorder

of Hb)

Co-existingstructural /

biosyntheticdisorders

Thalassaemias(Biosynthetic

disorderof Hb)

Constitute a major health problem in severalpopulations of the world

(particularly those residing in malariaendemic region)

10

HaemoglobinopathiesHaemoglobinopathies

• Genetic structural disorder.• Due to mutation in the globin gene of

haemoglobin.• Mostly autosomal recessive inheritance.• Result in haemoglobin variants with altered

structure and function.• Altered functions include:

Reduced solubility Reduced stability Altered oxygen affinity- increased or decreased Methaemoglobin formation

• Genetic structural disorder.• Due to mutation in the globin gene of

haemoglobin.• Mostly autosomal recessive inheritance.• Result in haemoglobin variants with altered

structure and function.• Altered functions include:

Reduced solubility Reduced stability Altered oxygen affinity- increased or decreased Methaemoglobin formation

11

HEMOGLOBINOPATHIHEMOGLOBINOPATHIESES

Decrease, lack of, or abnormal globin May be severe hemolytic anemia Abnormal Hb with low functionality Mutation may be deletion, substitution, elongation Hb electrophoresis may be helpful

12

• Point mutation: a change of a single nucleotide base in a DNA giving rise to altered amino acids in the polypeptide chains

(e.g. Hb S , Hb C)

• Deletions and additions: Addition and deletion of one or more bases in the globin genes

• Unequal crossing over: as in Hb-lepore and Hb-antilepore associated with -thalassaemias.

________________________________________________________*Most abnormal Hbs are produced by mutations in the structural genes which determine the amino acid sequence of the globin chains of the Hb molecule.

• Point mutation: a change of a single nucleotide base in a DNA giving rise to altered amino acids in the polypeptide chains

(e.g. Hb S , Hb C)

• Deletions and additions: Addition and deletion of one or more bases in the globin genes

• Unequal crossing over: as in Hb-lepore and Hb-antilepore associated with -thalassaemias.

________________________________________________________*Most abnormal Hbs are produced by mutations in the structural genes which determine the amino acid sequence of the globin chains of the Hb molecule.

Types of Mutations in HaemoglobinTypes of Mutations in Haemoglobin

13

Geographical distribution of common Hb variantsGeographical distribution of common Hb variants

Variant Occurrence predominantly in:

Hb S (6GluVal) Africa, Arabia, Black Americans

Hb C (6Glulys) West Africa, China

Hb E (26Glulys) South East Asia

Hb D (121GluGln) Asia

Hb O (121GluVal) Turkey and Bulgury

Variant Occurrence predominantly in:

Hb S (6GluVal) Africa, Arabia, Black Americans

Hb C (6Glulys) West Africa, China

Hb E (26Glulys) South East Asia

Hb D (121GluGln) Asia

Hb O (121GluVal) Turkey and Bulgury

14

Sickle Cell Haemoglobin

GAG GTG

RBC

Haemolysis

Sickle Cell

6

15

Inheritance of Sickle Cell Anaemia

AS AS

SSAAAS AS

AR

16

Lungs↑pO2

Tissues↓pO2

Red cell sickling

17

Sickling of the red cell during deoxygenation, as the HbShas low solubility at low O2 partial pressure and precipitates. Chronic haemolytic anaemia due to repeated sickling in tissues and unsickling in the lungs. Plugging of microcapillaries by rigid sickled cells leading to sickle cell crises i.e severe pain and edema. This causes significant damage to internal organs, such as heart, kidney, lungs and endocrine glands. Repeated infections. Frequent cerebrovascular accidents. Hand-foot syndrome (in small,i.e.around age of 3y); Swollen hands andfeet may be the first signs of sickle cell anemia in babies. The swelling is caused by sickle-shaped red blood cells blocking blood flow out of their hands and feet Bone deformation – bossing of the forehead. Hepato-spleenomegaly. Growth retardation. Frequent blood transfusion requirements. Psychosocial problems.

Major abnormalities & problems in SCA

18

Site of Sickling

Clinical Features

Management

Bone Painful crises Pain relief and hydration. Hydroxyurea

Lung Acute chest syndrome

Transfusion regimen, pain relief and hydration

Brain Stroke Transfusion regimen.

Heart Myocardial infarction

Transfusion regimen, pain relief and hydration

Spleen Acute splenic sequestration:

Transfusion, pain relief and hydration

Retina Proliferative retinopathy

Retinal surveillance. Laser

19

SICKLE CELL TRAITSICKLE CELL TRAIT

Heterozygous state for HbS (HbAS) No serious clinical consequences Sudden death during intensive training Hematuria, isosthenuria (renal papillary

necrosis)

20

Thalassaemias

Genetic disorders resulting fromdecreased biosynthesis of globin chains

of haemoglobin.

21

• A group ( not single identity) of Genetic defects.• Due to mutations in and around the globin genes.• Decreased production of one or more of the globin

chains.• Result in an imbalance in the relative amounts

of the - and non -chains. Altered /non- ratio.

• As a consequences of thalassaemias there is excess production of the other chains, and a decreased over all haemoglobin synthesis.

• A group ( not single identity) of Genetic defects.• Due to mutations in and around the globin genes.• Decreased production of one or more of the globin

chains.• Result in an imbalance in the relative amounts

of the - and non -chains. Altered /non- ratio.

• As a consequences of thalassaemias there is excess production of the other chains, and a decreased over all haemoglobin synthesis.

Thalassaemias

22

Types of Thalassaemias

- Thalassaemia* -Thalassaemia*

- Thalassaemia - Thalassaemia

- Thalassaemia * Most common23

Deletion of one or more alpha genes from chromosome 16

-a/aa: silent career with little signs --/aa: cis double deletion more common in

SEA -a/-a: trans double deletion --/-a: Hb H disease --/--: Hb Bart’s hydrops fetalis

α Thalassemia

24

ALPHA THALASSEMIAALPHA THALASSEMIA

αα/αα Normal

αα/α- Mild microcytosis

αα/- -

α-/α-

Mild microcytosis

α-/- - Hemoglobin H disease

- -/- - Hemoglobin Barts – Hydrops Fetalis25

- Thalassaemia

Hb In - Thalassaemia Decreased production of - chains

- Decreased / ratio

- Thalassaemia

Normal =

Accumulation of 26

Point Mutation producing - Thalassaemia

Less Frequent

exon1 exon2 exon3Chromosome 16

Introns

5’ 3’

2bp del

5bp del

Base Substitution

Chain TerminationDefect

Poly A signalMutation

27

Mutations Producing - Thalassaemia

DeletionsMost frequent:

/ -/- --/ --/----/--/

-thal 2hetero

-thal 1hetero

HbHDisease

Hydropsfetalis

-thal 2homo

Normal

Chromosome 16

28

• One -gene deletion.

-chain production is only about 75% of normal.

• May be homo- (- /- ) or heterozygous (- / )

• The patient usually shows a normal phenotypic appearance but there might be mild thalassaemia symptoms.

• Hypochromic-microcytic RBC’s due to partial reduction of -chain.

• One -gene deletion.

-chain production is only about 75% of normal.

• May be homo- (- /- ) or heterozygous (- / )

• The patient usually shows a normal phenotypic appearance but there might be mild thalassaemia symptoms.

• Hypochromic-microcytic RBC’s due to partial reduction of -chain.

- Thalassaemia-2

29

• Two -genes deletion- (o ) thal.

• The patient synthesizes -chain but it is decreased to about 50% of normal.

• Anaemic symptoms- hypochromic microcytic anaemia.

• May be homozygous (- -/- -) or heterozygous(--/ ). If the patient is homozygous than there is no -chain synthesis, and if heterozygous then there is decreased synthesis of the -chain to half normal level.

• Two -genes deletion- (o ) thal.

• The patient synthesizes -chain but it is decreased to about 50% of normal.

• Anaemic symptoms- hypochromic microcytic anaemia.

• May be homozygous (- -/- -) or heterozygous(--/ ). If the patient is homozygous than there is no -chain synthesis, and if heterozygous then there is decreased synthesis of the -chain to half normal level.

- Thalassaemia-1

30

• Three -gene (three alleles) deletion.

• The Hb present during foetal life is “Hb Bart’s” (4), while during adulthood the Hb present is “Hb H” (4).

• Some of the symptoms include: hepatosplenomegaly, impairment of erythropoiesis, and hypochromic-microcytic haemolytic anaemia.

• Three -gene (three alleles) deletion.

• The Hb present during foetal life is “Hb Bart’s” (4), while during adulthood the Hb present is “Hb H” (4).

• Some of the symptoms include: hepatosplenomegaly, impairment of erythropoiesis, and hypochromic-microcytic haemolytic anaemia.

Hb H Disease

31

• Homozygous o-thalassaemia.

• There is a complete absence of -chain (all -genes are deleted).

• The Hb produced at birth is Hb Barts (4).

• Hydrops foetalis is lethal and the baby is born dead.• Hydrops fetalis is a severe, life-threatening

problem of severe edema (swelling) in the fetus and newborn. It is also called hydrops.

• Symptoms include: Hepatosplenomegaly, severe hypochromic- microcytic anaemia.

• Homozygous o-thalassaemia.

• There is a complete absence of -chain (all -genes are deleted).

• The Hb produced at birth is Hb Barts (4).

• Hydrops foetalis is lethal and the baby is born dead.• Hydrops fetalis is a severe, life-threatening

problem of severe edema (swelling) in the fetus and newborn. It is also called hydrops.

• Symptoms include: Hepatosplenomegaly, severe hypochromic- microcytic anaemia.

Hydrops foetalis

32

- Thalassaemia

HbIn - Thalassaemia Decreased

production of - chains

Increased / ratio

- Thalassaemia

Normal =

Accumulation of 33

BETA THALASSEMIABETA THALASSEMIA

Impaired production of beta chain beta thalassemia minor – heterozygous (or trait) beta thalassemia major - homozygous

34

TYPES OF TYPES OF - THALASSAEMIA- THALASSAEMIA

Thalassemia Major (Cooley's anemia)

-severe form of beta thalassemia

- presence of two abnormal genes that cause either a severe decrease or complete lack of beta globin production.

Thalassemia Minor - presence of one normal

gene and one with a mutation

- causes mild to moderate mild anemia.

ETIOLOGY OF ETIOLOGY OF - THALASSAEMIA- THALASSAEMIA

Beta thalassemia is caused by a deficiency of Beta globin inherited in an autosomal recessive pattern, which means both copies of the HBB(Hemoglobin beta) gene in each cell have mutations.

The parents of an individual with an autosomal recessive condition each carry one copy of the mutated gene, but they typically do not show signs and symptoms of the condition.

BETA THALASSEMIA TRAITBETA THALASSEMIA TRAIT

No symptoms Mild microcytic anemia

38

BETA THALASSEMIA MAJORBETA THALASSEMIA MAJOR

No beta chain produced (no HbA) Severe microcytic anemia occurs gradually in the

first year of life Marrow expansion Iron overload Growth failure and death

39

Usually point mutation in the control region. The absence of beta-globin is referred to as beta-zero

(B0) thalassemia. Other HBB gene mutations allow some beta-globin to be

produced but in reduced amounts. A reduced amount of beta-globin is called beta-plus (B+) thalassemia β+ has minimal production.

β+/β+ or βo/βo is thal major or Cooley’s anem Often not apparent at birth until β chain takes over γ

chain production. High Hb A2, Hb F. Related: Hb Lepore (δ-β usion), High Persistence of

Foetal Hb; (HPFH).

- Thalassaemia

40

• It is characterized by either no -chain synthesis (i.e. o) or decreased synthesis of -chain (+).

• Excess -chains precipitate in RBC’s causing severe ineffective erythropoiesis and haemolysis.

• The greater the -chains, the more severe the anaemia.

• Production of -chains helps to remove excess -chains and to improve the -thalassaemia. Often HbFlevel is increased.

• Majority of -thalassaemia is due to point mutation.

• It is characterized by either no -chain synthesis (i.e. o) or decreased synthesis of -chain (+).

• Excess -chains precipitate in RBC’s causing severe ineffective erythropoiesis and haemolysis.

• The greater the -chains, the more severe the anaemia.

• Production of -chains helps to remove excess -chains and to improve the -thalassaemia. Often HbFlevel is increased.

• Majority of -thalassaemia is due to point mutation.

- Thalassaemia

41

• The -chain is totally absent.• There is increase in HbF with absence of HbA.• This is combined with ineffective erythropoiesis.• In majority of the cases, -gene is present but

there is complete absence of mRNA.• Characteristics of this disorder are:

• Skeletal deformities (e.g. enlargement of upper jaw, bossing of skull and tendency of bone fractures).

• Severe hypochromic- microcytic anaemia.• Survival depends on regular blood transfusion.• This leads to iron overload (iron accumulates in

the blood and tissues, causing tissue damage). • Death usually occurs in the 2nd decade of life

(i.e. at age of about 20 years) if measures are not taken to avoid iron overload by chelation therapy.

• The -chain is totally absent.• There is increase in HbF with absence of HbA.• This is combined with ineffective erythropoiesis.• In majority of the cases, -gene is present but

there is complete absence of mRNA.• Characteristics of this disorder are:

• Skeletal deformities (e.g. enlargement of upper jaw, bossing of skull and tendency of bone fractures).

• Severe hypochromic- microcytic anaemia.• Survival depends on regular blood transfusion.• This leads to iron overload (iron accumulates in

the blood and tissues, causing tissue damage). • Death usually occurs in the 2nd decade of life

(i.e. at age of about 20 years) if measures are not taken to avoid iron overload by chelation therapy.

o-Thalassaemia

1. Mutations affecting transcription initiation2. Mutations affecting RNA splicing3. Mutations affecting translation initiation4. Non-sense Mutations.5. Mutations of polyadenylation site.

Mutations affecting the -Globin gene.

>200 -Thalmutations reported

to-date Worldwide

Chromosome 11

43

Clinical Classification of ThalassaemiasClinical Classification of Thalassaemias

1. Thalassaemia major:The patient depends on blood transfusions

especially if he is homozygous.2. Thalassaemia intermediate:

• Homozygous mild +-thalassaemia. • Co-inheritance of -thalassaemia.• Heterozygous -thalassaemia.• Co-inheritance of additional -globin genes. -thalassaemia and hereditary persistence of foetal

Hb• Homozygous Hb lepore• Hb H disease.

3. Thalassaemia minor (trait): o-thalassaemia trait. +-thalassaemia trait.• Hereditary persistence of foetal Hb only. -thalassaemia trait. o- and +-thalassaemia trait.

Clinical Classification of ThalassaemiasClinical Classification of Thalassaemias

1. Thalassaemia major:The patient depends on blood transfusions

especially if he is homozygous.2. Thalassaemia intermediate:

• Homozygous mild +-thalassaemia. • Co-inheritance of -thalassaemia.• Heterozygous -thalassaemia.• Co-inheritance of additional -globin genes. -thalassaemia and hereditary persistence of foetal

Hb• Homozygous Hb lepore• Hb H disease.

3. Thalassaemia minor (trait): o-thalassaemia trait. +-thalassaemia trait.• Hereditary persistence of foetal Hb only. -thalassaemia trait. o- and +-thalassaemia trait.

44

BETA THALASSEMIA MAJOR BETA THALASSEMIA MAJOR TREATMENTTREATMENT

Transfusion Iron chelation Stem cell transplant

45

• This is an abnormal Hb due to unequal crossing-over of the - and -genes to produce a polypeptide chain consisting of the - chain at its amino end and - chain at its carboxyl end.

• The -fusion(hybrid) chain is synthesized inefficiently and normal and -chain production is abolished.

• The homozygotes show thalassaemia intermediate and heterozygotes show thalassaemia trait.

• This is an abnormal Hb due to unequal crossing-over of the - and -genes to produce a polypeptide chain consisting of the - chain at its amino end and - chain at its carboxyl end.

• The -fusion(hybrid) chain is synthesized inefficiently and normal and -chain production is abolished.

• The homozygotes show thalassaemia intermediate and heterozygotes show thalassaemia trait.

Hb-Lepore

Chromosome 11

AG

3’5’

HEMOGLOBINOPATHY-ANTENATAL HEMOGLOBINOPATHY-ANTENATAL DIAGNOSISDIAGNOSIS

Test partners of heterozygous or affected individuals Antenatal diagnosis from DNA obtained by chorionic

villus sampling, or by amniocentesis

47