Haemoglobinopathies

Haemoglobinopathies

Blood and Genetics Vertical Module — 2026. Lecturer: Dr. Bokor, Department of Biochemistry / Department of Pathology, University of Otago. Reference: Genetics in Medicine, Chapter 11 (Thompson).

Overview

Haemoglobinopathies are common genetic blood disorders caused by mutations that affect either haemoglobin's structure (haemoglobin variants) or its production (thalassaemias). They are the most prevalent monogenic disorders worldwide, affecting ~500,000 births per year. Most common in individuals of African, Mediterranean, Middle Eastern, and South Asian descent — geographic overlap with historical malaria burden explains carrier frequency selection.

Normal Haemoglobin Structure

Adult haemoglobin HbA = α₂β₂ (two alpha chains + two beta chains), each with a haem group (Fe²⁺ + porphyrin ring) capable of reversible oxygen binding.

• Hb A (α₂β₂): >95% of adult haemoglobin
• Hb A₂ (α₂δ₂): ~2.5%; useful diagnostic marker
• Hb F (α₂γ₂): <1% in adults; predominant fetal haemoglobin (high O₂ affinity — benefits transplacental O₂ transfer)

Gene dosage: 4 alpha-globin genes (2 on each chromosome 16); 2 beta-globin genes (1 on each chromosome 11). This asymmetry means: losing one alpha gene is often silent (3 copies remain); losing one beta gene always causes beta-thalassaemia trait.

Role of 2,3-bisphosphoglycerate (2,3-BPG): stabilises the deoxygenated T-state of Hb by binding to non-alpha subunits. Beta chains (HbA) have strong BPG binding → lower O₂ affinity; gamma chains (HbF) have weak BPG binding → higher O₂ affinity (favours O₂ uptake from maternal blood).

Haemoglobin Variants — Structural Mutations

#### Sickle Cell Disease (HbS — Haemoglobin S)

Molecular basis: Point mutation in codon 6 of the beta-globin gene: GAG → GTG → Glutamate (polar, negatively charged) → Valine (non-polar, uncharged). This is a single nucleotide substitution.

Mechanism of sickling: In the deoxygenated state, the normal hydrophobic pocket on the beta-globin surface is exposed at the Val-6 position. Deoxygenated HbS molecules polymerise end-to-end into rigid polymer fibres (tactoids), distorting red cells into the characteristic sickle shape. In the oxygenated state, Val-6 is buried — so oxygenated HbS does not polymerise.

Gene dosage (dose effect):

| Genotype | Disease | Features |

|----------|---------|---------|

| HbAA | Normal | — |

| HbAS | Sickle cell trait (carrier) | Mild/asymptomatic; protection against P. falciparum malaria |

| HbSS | Sickle cell disease | Severe; chronic haemolytic anaemia + vaso-occlusive crises |

| HbSC | HbSC disease | Intermediate severity; SC compound heterozygote |

| HbS/β-thal | Sickle/β-thalassaemia | Variable; depends on β-thal type |

Clinical features of HbSS (sickle cell disease):

1. Chronic haemolytic anaemia: Hb typically 60–90 g/L; elevated LDH, reticulocytosis, unconjugated bilirubin, low haptoglobin; jaundice; splenomegaly (in children; autosplenectomy develops over time due to repeated infarction)
2. Vaso-occlusive crises: sickled RBCs are rigid, adhesive, and occlude small vessels → ischaemic pain crises (bones most common — long bones, vertebrae), acute chest syndrome (pulmonary infiltrates + chest pain/hypoxia — leading cause of death), stroke (~10% without prophylaxis), priapism, avascular necrosis (femoral head)
3. Functional asplenia: by ~5 years; increased susceptibility to encapsulated bacteria (Streptococcus pneumoniae, Haemophilus influenzae, Neisseria meningitidis) → overwhelming sepsis; requires penicillin prophylaxis + vaccines
4. Acute splenic sequestration: sudden massive trapping of RBCs in spleen → acute anaemia, hypovolemia (life-threatening in young children)
5. Aplastic crises: parvovirus B19 infects erythroid precursors → temporary cessation of erythropoiesis → profound anaemia (no reticulocytes)
6. Chronic organ damage: renal papillary necrosis, sickle nephropathy, pulmonary hypertension, retinopathy, leg ulcers

Management:

• Hydroxyurea (hydroxycarbamide): induces HbF production (gamma chains do not polymerise with HbS) → reduces sickling frequency and severity; reduces vaso-occlusive crises and acute chest syndrome
• Regular blood transfusions: dilutes HbS; used for stroke prevention and acute chest
• Folic acid: supplements increased haemopoietic requirement
• Penicillin prophylaxis + pneumococcal/meningococcal/Hib vaccines: prevent sepsis
• Curative: allogeneic haemopoietic stem cell transplantation (HSCT); gene therapy (beta-globin gene addition or BCL11A silencing to re-induce HbF) — emerging treatments
• Analgesia: paracetamol, NSAIDs, opioids for painful crises; IV fluids; oxygen

Malaria protection: HbS/HbC carrier frequencies are highest in malaria-endemic regions (West Africa: HbS ~30–40%). In malaria-infected RBCs (Plasmodium falciparum consumes O₂ → deoxygenation) → premature sickling/crystallisation → immune system destroys infected RBCs before parasite completes sexual cycle → carrier protection.

#### Haemoglobin C (HbC)

Point mutation at the same beta-6 position: GAG → AAG → Glutamate → Lysine (positively charged). Unlike HbS (which polymerises), the charge change in HbC causes crystal formation (more ordered than HbS fibres) in deoxy state → rigid, less deformable RBCs → mild chronic haemolytic anaemia, target cells on blood film, splenomegaly. HbCC: mild haemolytic anaemia. HbSC: intermediate sickle cell disease severity.

#### Haemoglobin E (HbE)

Point mutation at beta-26: Glutamate → Lysine. Creates an abnormal mRNA splice site → reduced production of normal beta chains (behaves partly like a thalassaemia). Common in Southeast Asia. HbEE: mild microcytic anaemia. HbE/β-thal: clinically significant, can resemble β-thal major.

Thalassaemias

Thalassaemias result from mutations that reduce or abolish production of globin chains (gene expression defect, not structural variant). This leads to imbalanced globin chain production, ineffective erythropoiesis, haemolysis, and anaemia.

#### Alpha-Thalassaemia

Caused predominantly by deletions of alpha-globin genes. There are 4 alpha genes (2 per chromosome 16); severity depends on how many are deleted:

| Alpha genes remaining | Condition | Clinical features |

|-----------------------|-----------|------------------|

| 4 (normal) | Normal | — |

| 3 (−α/αα) | Silent carrier | Normal or very mild microcytosis; usually undiagnosed |

| 2 (−−/αα or −α/−α) | Alpha-thalassaemia trait | Mild anaemia, mild microcytosis; MCV ↓, MCH ↓ |

| 1 (−−/−α) | HbH disease | Moderate-severe haemolytic anaemia; HbH = β₄ tetramers (unstable); jaundice, splenomegaly, transfusion-dependent |

| 0 (−−/−−) | Hb Bart's hydrops fetalis | Hb Bart's = γ₄; virtually no functional Hb; incompatible with extrauterine life; usually fatal in utero or at birth; severe gestational complications for mother |

Pathophysiology: ↓ alpha chains → excess beta chains (in adults) or gamma chains (in fetus). These form unstable tetramers (HbH = β₄; Hb Bart's = γ₄) that bind oxygen too tightly (shifted left on O₂ dissociation curve) → poor tissue O₂ delivery, haemolysis.

Genotyping is essential for carrier couples (especially Southeast Asian populations where −−/αα deletions common on single chromosome) to assess risk of Hb Bart's hydrops in offspring.

#### Beta-Thalassaemia

Caused by point mutations (rarely deletions) of beta-globin genes. Because there is only 1 beta gene per chromosome (2 total), even one abnormal beta gene causes beta-thalassaemia trait. Two mutation classes:

• β⁰: no beta chain production from that allele
• β⁺: reduced (but not absent) beta chain production

| Genotype | Condition | Features |

|----------|-----------|---------|

| β/β | Normal | — |

| β/β⁺ or β/β⁰ | Beta-thalassaemia minor (trait) | Mild microcytic anaemia; usually asymptomatic; ↑ HbA₂ (>3.5%) on Hb electrophoresis — diagnostic |

| β⁺/β⁺ or β⁺/β⁰ | Beta-thalassaemia intermedia | Moderate anaemia (Hb ~70–100 g/L); jaundice, splenomegaly, extramedullary haematopoiesis; may not be transfusion-dependent |

| β⁰/β⁰ | Beta-thalassaemia major (Cooley anaemia) | Severe transfusion-dependent anaemia from 3–6 months of age (when HbF → HbA switch occurs); massive splenomegaly, bone deformities (marrow expansion → frontal bossing, "hair-on-end" skull XR), growth retardation, iron overload (from transfusions + increased gut absorption) |

Pathophysiology of beta-thalassaemia: ↓ beta chains → excess unpaired alpha chains. Free alpha chains are highly toxic — they precipitate within RBC precursors → destruction in bone marrow (ineffective erythropoiesis, hallmark of beta-thal major). Surviving RBCs have short lifespan (haemolysis). Marrow attempts compensatory expansion → bone deformities, hepatosplenomegaly (extramedullary haematopoiesis).

Diagnosis: Hb electrophoresis (HPLC): beta-thal trait shows ↑ HbA₂ (>3.5%). Beta-thal major: near-absent HbA, elevated HbF; confirmed by molecular genotyping.

Treatment of beta-thalassaemia major:

• Regular blood transfusions (every 3–4 weeks) to suppress ineffective erythropoiesis and maintain Hb >90–100 g/L
• Iron chelation therapy (desferrioxamine SC infusion, or oral deferasirox/deferiprone) to prevent iron overload (haemosiderosis) → cardiac, hepatic, endocrine damage
• Folic acid supplementation
• Curative: allogeneic HSCT (HLA-matched sibling donor); gene therapy (betibeglogene/autologous HSC gene therapy — approved for transfusion-dependent thalassaemia)
• Luspatercept (activin receptor ligand trap): reduces ineffective erythropoiesis → reduces transfusion burden

Genetic Counselling and Prenatal Diagnosis

• Thalassaemias and sickle cell disease are autosomal recessive — both parents must be carriers for 25% risk of affected offspring
• Carrier testing offered to at-risk ethnic groups (Mediterranean, African, Middle Eastern, South Asian, Southeast Asian)
• Prenatal diagnosis: CVS (chorionic villus sampling, 10–13 weeks) or amniocentesis (15–18 weeks) → molecular genetic testing of fetal DNA
• Pre-implantation genetic diagnosis (PGD) available for couples undergoing IVF

Haemolytic Anaemias (Overview)

Anaemia caused by premature destruction (haemolysis) of RBCs. Classified as:

• Intravascular (within blood vessels): mechanical trauma (microangiopathic haemolytic anaemia — schistocytes; TTP, DIC, HUS), complement-mediated (PNH)
• Extravascular (reticuloendothelial system, spleen): hereditary spherocytosis, haemoglobinopathies, autoimmune haemolytic anaemia (AIHA)

Laboratory markers of haemolysis: ↑ LDH (released from RBCs), ↑ unconjugated (indirect) bilirubin (haem breakdown), ↓ haptoglobin (binds free Hb → cleared), reticulocytosis (bone marrow compensation), haemoglobinuria (intravascular haemolysis only). Blood film: spherocytes (hereditary spherocytosis, AIHA), sickle cells, target cells, schistocytes.

Direct Coombs test (DAT): detects antibody/complement on RBC surface → positive in immune-mediated haemolytic anaemias (AIHA, haemolytic transfusion reactions). Negative in non-immune haemolysis (sickle cell, hereditary spherocytosis, mechanical).