Haematology Rocks – Detailed Study Notes

Objectives of the Lecture

Understand the breadth and sub-specialties of Haematology
- Malignant (Leukaemias, Lymphomas, Myeloma)
- Transplantation & Immunotherapy
- Haemostasis & Thrombosis
- Transfusion Medicine / Blood Banking
- Red-Cell Medicine (e.g.
  Sickle-cell anaemia, Thalassaemias)
- Laboratory Haematology (morphology, flow cytometry, cytogenetics, NGS)
Recognise recent paradigm-shifting advances (Immunotherapy, Gene Editing)
Grasp the “bench → bedside” pipeline unique to haematology

Big Picture: Why “Haematology Rocks”

Rapid translation of lab discoveries to clinical care (shortest lag among medical specialities)
Opportunity to • cure cancers • correct monogenic disorders • study fundamental biology
Three canonical monogenic targets: Sickle-cell disease (SCD), β-Thalassaemia, Haemophilia

Classical Chemotherapy vs. Modern Selective Strategies

Chemotherapy = non-selective cytotoxin; kills all rapidly dividing cells → alopecia, mucositis, neutropenia, diarrhoea, immune suppression (“Chernobyl” effect)
Modern aim: “Give eyes to chemotherapy” → kill only tumour clone, spare normal tissue
- Antibodies (naked, conjugated, bispecific)
- Cellular therapies (CAR-T)
- Small-molecule targeted inhibitors (based on NGS signatures)

Antibody Era

1985 CHOP (Cyclophosphamide + Doxorubicin + Vincristine + Pred) = first chemo backbone for B-cell lymphoma
1997 Rituximab (anti-CD20; naked mAb) added → $\text{R–CHOP}$ improved survival curves
- Attacks only CD20⁺ B-cells (malignant & normal) → fewer systemic toxicities

Beyond Naked mAb: BiTEs

Bi-specific T-cell Engagers possess two arms
- One arm → tumour antigen (e.g.
  CD19)
- Second arm → CD3 on any T-cell
- Forces an immunologic synapse → T-cell–mediated cytolysis of bound tumour

CAR-T Cells

“Chimeric Antigen Receptor” = synthetic receptor inserted into patient’s own (autologous) T-cells
- Ectodomain (“lion’s head”) = tumour-binding scFv (CD19, CD22, etc.)
- Endodomain (“snake tail”) = signalling modules that unleash cytokine storm → tumour lysis
Manufacturing: leukapheresis → viral transduction → expansion → conditioning chemo → infusion
Indications (UK): up to 25 yrs for ALL; also Large B-cell, Mantle-cell lymphoma; myeloma licensing imminent
Limitation: Ineffective in solid tumours (hypoxic, acidic micro-environment + heterogeneous antigens)

Landmark Example – Emily Whitehead

2012: First paediatric ALL patient to receive CAR-T (1.2 million cells)
Developed severe Cytokine Release Syndrome (CRS) – IL-6 driven
- Managed with Tocilizumab (IL-6R blocker) – path-finding use, now standard of care
By day 23: $\text{T-cells } 74.5\% \rightarrow \text{blasts } 0.4\%$
Achieved MRD <10^{-6} (no detectable residual disease in $10^{6}$ cells)

Immune Checkpoints & Macrophage Checkpoints

PD-L1 / PD-1 axis
- ICIs: Nivolumab, Pembrolizumab
- Haem-indication: Hodgkin lymphoma (because Reed–Sternberg cells over-express PD-L1)
CD47 “don’t-eat-me” signal
- Anti-CD47 mAb (Magrolimab) restores macrophage phagocytosis
- Shows promise in $\text{TP53}^{\text{mut}}$ AML (p53 loss → poor prognosis)

Gene Editing: CRISPR–Cas9

Nobel-prize technology (Charpentier & Doudna, 2020)
- Programmable endonuclease makes double-strand break
- Repair template allows correction, knock-out, or activation
Ethical pause after Chinese embryo editing for HIV resistance

Haematology Applications

β-Thalassaemia (quantitative)
- Edit erythroid enhancer of $BCL11A$ to re-activate fetal Hb (HbF) → ↑ γ-globin, ↓ transfusion need
Sickle-cell disease (qualitative)
- Either correct $HBB^{E6V}$ mutation or, again, raise HbF ≥10 % to inhibit HbS polymerisation
Early clinical data
- Patient 1: pre-therapy Hb $7.2\,\text{g/dL} \rightarrow 12\,\text{g/dL}$ ; HbF 43 %; transfusion-free >15 mo
- Swimmer plots: drastic fall in vaso-occlusive crises (VOC) post-edit; similar trend in thalassaemia → transfusion-independent ≥12 mo

Bottleneck = conditioning busulfan (myelo-ablation)
- Causes infertility; increases cost (≈ $£1.6\,\text{million}$ + fertility preservation)
- Research ongoing for chemo-free delivery platforms

Socio-Ethical Lens

Disparity in drug approvals: 15 new CF drugs vs. 2 SCD drugs over 12 yrs despite 3× prevalence → highlights structural racism

Viral Vector Gene Transfer (Haemophilia)

Distinct from CRISPR (no genomic integration; episomal)
Use replication-defective AAV capsid to deliver missing gene to hepatocytes

Haemophilia B (Factor IX)

Goal: raise \text{FIX}\,>\,5\% (severe <1 %)
Padua variant (gain-of-function): activity 8–10× normal → durable expression 7–10 yrs
NHS list price ≈ $£2.3\,\text{million}$ one-off

Haemophilia A (Factor VIII)

FVIII cDNA = $\approx 7\,\text{kb}$ (won’t fit into AAV 4.7 kb)
- Solve via B-domain deletion (BDD) + codon optimisation
Early UK trial: 7 patients reached 70–90 % FVIII, but waned to 15–20 % by year 4–5 (unknown mechanism)

Blood-Film Morphology: “Cell Narratives”

Concept of Poikilocytosis

Any abnormally shaped red cell; pattern recognition + story behind shape

1 Sickle & C-crystals

HbS (β6 Glu→Val) polymerises on de-oxygenation → irreversibly sickled crescents
HbC (β6 Glu→Lys) crystallises on oxygenation → parallelogram crystals
SC poikilocyte = cell carrying one S & one C allele – half-sickle, half-crystal; typical in Afro-Caribbean adults (e.g.
59-yr-old woman with bilateral hip osteonecrosis)

2 Spherocytes (loss of membrane)

Appearance: small, dense, no central pallor
Mechanisms
1. Hereditary spherocytosis (germ-line defects in spectrin, ankyrin, band 3)
- Neonatal jaundice, anaemia (\text{Hb}_{\text{neonate}}<180\,\text{g/L})
- Negative Coombs test
1. Auto-immune haemolytic anaemia (warm IgG) – Coombs-positive
2. Severe burns, Clostridial sepsis, certain malarias

3 Target Cells (excess membrane OR shrunken cytoplasm)

“Bull’s-eye”/dartboard appearance
Causes: cholestatic/obstructive jaundice, alcoholic liver disease, HbC, α/β-thalassaemia
Diagnostic utility limited (non-specific)

4 Spiky Cells

Acanthocyte (thorny, irregular projections)
- Alcoholic cirrhosis; Neuro-acanthocytosis (e.g.
  McLeod syndrome – seizures, cardiomyopathy in 20–30 yr male)
Echinocyte (sea-urchin; many symmetric blunted spikes)
- Artefact (old EDTA sample) → “cremated cells”
- Uraemia / dialysis → “Burr cells”
- Pyruvate-kinase deficiency → “prickle cells”; now treatable with PK activator (oral)

5 Bite / Blister / Keratocyte Cells

G6PD deficiency (X-linked; boys)
- Oxidant stress (infection, fava beans, primaquine) → Hb denatured into Heinz bodies
- Splenic macrophage “bites” out Heinz body → keratocyte with 1–2 horns
- Blood-film triad: keratocytes + hemi-ghosts/blister cells + dark urine (intravascular haemolysis)
- Dark urine = haemoglobinuria (free Hb), not haematuria (intact RBCs)

Intravascular vs. Extravascular Haemolysis (Quick Review)

Extravascular: spleen/liver macrophages; jaundice but no haemoglobinuria
Intravascular: free Hb in plasma → binds haptoglobin; overflow → haemoglobinuria; nitric-oxide scavenging → vasoconstriction, renal injury
G6PD crisis = commonest intravascular haemolysis in exams

Numerical & Statistical Pearls

Minimal Residual Disease (MRD) sensitivity in haematology: $1\,\text{leukaemic cell}/10^{6}$ normal cells
AAV cargo limit ≈ $4.7\,\text{kb}$ ; FVIII gene size ≈ $7\,\text{kb}$ ; FIX gene size ≈ $1.4\,\text{kb}$
Busulfan-based CRISPR protocol cost ≈ $£1.6\,\text{million}$ (+ fertility preservation)
Padua FIX variant patient’s activity: $940\%$ normal → inspiration for gene therapy

Practical / Philosophical Points

Benchmark for success in SCD gene editing: ≥10 % rise in HbF markedly reduces VOC
Allogeneic “off-the-shelf” CAR-T aims to cut 3-week autologous manufacturing lag; risk = GvHD & rejection
Persistent need to balance cure (chemotherapy, busulfan) vs. quality-of-life (fertility, long-term toxicity)
Hidden bias: slower therapeutic innovation for diseases affecting predominantly Black patients (SCD) compared with those affecting White patients (CF)
Bench discoveries (e.g.
PK activator, Tocilizumab for CRS) rapidly re-shape bedside protocols → stay current!

Exam-Day Quick Reference

Rituximab → CD20; BiTE → CD19/CD3; CAR-T → custom (CD19 most common)
ICIs (PD-1/PD-L1) licensed only for Hodgkin in haematology context
Magrolimab → anti-CD47, useful in $TP53^{mut}$ AML/MDS
CRISPR editing target in Thalassaemia/SCD = $BCL11A$ enhancer; HbF protective
Spherocytes + Coombs(–) infant jaundice → hereditary spherocytosis; Coombs(+) adult → AIHA
Burr cells + high creatinine → uraemia; Prickle cells + anaemia → PK deficiency
Bite cells + dark urine male → G6PD crisis (trigger? fava, infection, drug)
Acanthocytes + neuro-muscular + cardiomyopathy → McLeod (neuro-acanthocytosis)
Cost remembrance: $\text{CAR-T}\approx£0.3–£0.4\,\text{M}$ ; CRISPR $£1.6\,\text{M}$ ; Haemophilia B gene therapy $£2.3\,\text{M}$