Updates in the Diagnosis & Classification of Myeloproliferative Disorders
From Disorder, Disease to Neoplasm Jameela Sathar Ampang Hospital 23 April 2010
The story…
Clinical insights • 1892 Louis Henri Vaquez Polycythaemia Vera (PV) • 1890s Gustav Hueck Primary Myelofibrosis (PMF) • 1934 Emil Epstein & Alfred Goedel Essential Thrombocythaemia (ET)
Myeloproliferative ‘Disorders’ William Dameshek 1951 • PV • ET • PMF – Overlap in clinical and laboratory features
PV, ET, PMF • ? Separate diseases • ? Same disease, different manifestations • ? Combination of both
Clonality • 1967 Fialkow Polymorphisms at G6PD locus • 1974 Axelrad, Prchal Endogenous Erythroid Colony formation • 1976 Adamson Single G6PD isoform in red cells, granulocytes and platelets from PV patients
JAK2V617F • 2005 Vainchenker, Kravolics, Levine JAK2V617F mutation • Valine to phenylalanine substitution at codon 617 of JAK2 on chromosome 9p • Constitutive kinase activation of hematopoeitic growth factor receptors
Goldman JM. N Engl J Med 2005;352:1744-1746
JAK2 Domains Cytokine Pseudokinase receptor domain Kinase binding domain JH7 JH6 JH5 JH4 JH3 JH2 JH1 Exon 12 mutation V617F
JAK2V617F • 95% in PV ( JAK2 exon 12 mutation in 5%) • 50% in ET and PMF • <3% in MDS • Not seen in reactive myeloproliferation or lymphoid disorders Levine RL, Blood 2006
JAK2 mutation analysis • Allele-specific PCR assay • Real-time PCR • Pyrosequencing • Restriction enzyme digestion Levine RL 2006; James C 2006; Baxter EJ 2005; Jones AV 2005
? 1Mutation, 3Diseases • Gene dosage – high neutrophil allele burden – higher Hct and WBC, lower platelet, splenomegaly • Homozygosity – homozygous JAK2V617F favours PV over ET • Inherited MPD alleles • Cooperating mutations eg. 20q- Tiedt R 2007; Xing S, Blood 2008
Other mutations that activate JAK2 signaling • MPLW515L/K • 5-10% of JAK2 negative ET and PMF • Higher platelet counts • Marked fibrosis Beer P, Blood 2008 • JAK2 exon 12 • Only identified in JAK2V617F negative PV Percy MJ 2007
WHO Classification
Before JAK2V617F
Chronic Myeloproliferative ‘Diseases’ WHO 2001 • Chronic Myeloid Leukemia (CML) • Chronic Neutrophilic Leukemia (CNL) • Chronic Eosinophilic Leukemia/ hypereosinophilic syndrome (CEL/ HES) • PV • Chronic idiopathic myelofibrosis (CIMF) • ET • MPD, unclassifiable
Discovery of JAK2V617F
Myeloproliferative ‘Neoplasms’ WHO 2008 • CML, Bcr-Abl positive • CNL • PV • Primary Myelofibrosis • ET • CEL-NOS • Mastocytosis • Myeloproliferative Neoplasm, unclassifiable
Polycythaemia Vera
WHO 2008- PV Major criteria Minor criteria 1. Hb >18.5(M); 1. BM trilineage >16.5(F) or other myeloproliferation RCV 2. Subnormal serum 2. Presence of epo level JAK2V617F or exon 3. EEC growth 12 mutation Diagnosis of PV: • both major + 1 minor or • 1 st major + 2 minor
WHO 2001- PV A1: RCM >25% B1: thrombocytosis >400 A2: no 2 0 erythrocytosis B2: WBC >12 A3: splenomegaly B3. PV BM changes A4: clonal abn; non-bcr- B4: low serum epo level abl A5: EEC formation in vitro Diagnosis: First 2A + one other A or 2B
WHO 2008 PV criteria • Omits: – splenomegaly – platelet and leucocyte count – no secondary cause for erythrocytosis • New: – JAK2V617F mutation – Hb>18.5 (M); Hb>16.5 (F) = absolute erythrocytosis – Importance of BM changes
Erythrocytosis • Sets PV apart from other MPDs • No evidence to support substitution of Hb values for red cell mass • Normal Hb or Hct ≠ normal red cell mass • Haemodilution from hypersplenism; iron deficiency anaemia
WHO Hb guidelines (M) True Apparent erythrocytosis Erythrocytosi Hb >18.5 g/dL s (n= 31) Hb <18.5 g/dL RCM and PV (n= 49) True 11 (35%) 20 (65%) erythrocytosis Apparent 7 (14%) 42 (76%) erythrocytosis Johansson PL, BJH 2005
WHO Hb guidelines (F) True Apparent erythrocytosis Erythrocytosi Hb >16.5 g/dL s (n=46) Hb <16.5 g/dL RCM and PV (n=17) True 29 (63%) 17 (37%) erythrocytosis Apparent 6 (35%) 11 (65%) erythrocytosis Johansson PL, BJH 2005
Hematocrit • Hct ≥60 = absolute erythrocytosis 2 studies: Johansson (2005) and Pearson (1984)
Red Cell Mass • Red Cell Mass 51 Cr- and Plasma Volume 125 I • Cumbersome, costly and time-consuming • Mainly to distinguish PV from ET in absence of high Hct • Becoming obsolete with JAK2 mutation
Phlebotomy trial • Absolute erythrocytosis: – Requiring ≥ 2 phlebotomies – To reduce Hct to < 45% (M) or < 42% (F) – 10% Hct rise within 3 months Spivak JL 2008
JAK2V617F in PV • >95% of patients with PV • High negative predictive value in PV • V617F homozygosity is specific for PV Scott LM; Blood 2006
JAK2V617F in Budd Chiari • MPD represent the commonest cause of Budd Chiari syndrome • >50% with unexplained Budd Chiari syndrome are JAK2V617F positive Janssen HL, J Hepatol 2003; Patel RK, Gastroenterology 2006; Kiladjian JJ, Blood 2008
BM morphology • Not specific • Overlap, evolving disease • PVSG study: 281 PV patients at diagnosis – 13% did not have increased marrow cellularity – 11% had moderate to marked increase in reticulin • Inadequate tool to distinguish PV from ET or PMF Ellis JT 1986
?
Serum erythropoeitin • A normal epo level does not exclude PV • A low epo level is not specific for PV • Epo levels do not distinguish PV from V617F- positive ET • Erythrocytosis + presence of JAK2V617F makes epo level redundant Casadevall N 1994
Endogenous Erythroid Colony (EEC) formation • Only in research laboratories • Never standardised • Not specific; can also be observed in ET
‘Realistic’ PV criteria Major criteria Minor criteria • Hct >60 • BM trilineage hyperplasia • Absent bcr-abl • Palpable • JAK2V617F splenomegaly • Thrombocytosis >400 • Leukocytosis >12 PV diagnosis= All 3 major + 1 minor or first 2 major + 2 minor
Essential Thrombocythaemia
ET • The only MPD without a specific phenotype • A diagnosis of exclusion • JAK2V617F identifies 50% of patients with isolated thrombocytosis as possibly having ET
WHO 2008- ET All 4 criteria: 1. Sustained platelet ≥ 450 x 10 9 /L 2. Megakaryocyte proliferation with large and mature morphology. No or little granulocyte or erythroid proliferation 3. Not meeting WHO criteria for CML, PV, MF, MDS 4. JAK2V617F mutation or other clonal marker or no evidence of reactive thrombocytosis
JAK2V617F in ET • High positive predictive value for a MPD vs. reactive states • V617-positive thrombocythemia resembles PV • Higher levels of Hb & white cells and a more cellular BM Campbell PJ 2005
Clinical Consequences of the JAK2 Mutation in ET JAK2V617F JAK2 wild-type p n=414 n=362 Age* (years) 60 (39 − 77) 52 (32 − 75) <0.0001 Hb (g/L) 145 (14) 135 (14) <0.0001 WBC † 10.6 (3.4) 9.3 (2.6) <0.0001 Neutrophils † 7.4 (3.0) 6.2 (2.2) <0.0001 Platelets † 902 (276) 1030 (343) <0.0001 Campbell PJ, Lancet 2005;366:1945-1953
Primary Myelofibrosis
WHO 2001- PMF • Prefibrotic vs. fibrotic stage • Mild vs. marked – PMF BM changes – Leukoerythroblastosis/ dacrocytosis – Anaemia and/or organomegaly – Leukothrombocytosis
WHO 2008- PMF Major criteria Minor criteria 1. Megakaryocyte prolif 1. Leukoerythroblastosis and atypia ± 2. Increased serum LDH reticulin/collagen fibrosis 3. Anaemia 2. Not meeting WHO 4. Palpable criteria for CML, PV, MDS splenomegaly 3. JAK2V617F mutation or other clonal marker or no reactive fibrosis Diagnosis: all 3 major + 2 minor
Emphasis on megakaryocyte atypia in BM histology Polypoid nuclei Bulbous nuclei Loose clusters Dense clusters PMF ET Thiele J 2006
PMF • Biopsy is essential for diagnosis • Primary vs. idiopathic • Presence of MF does not exclude PV or ET • Prefibrotic phase- not possible to identify morphologically • Splenomegaly- minor or major criteria?
JAK2V617F in PMF • Not specific for PMF • Helpful in differentiating PMF from reactive conditions
Impact of V617F on Prognosis
Conflicting evidence • Risk of thrombosis • Risk of leukemic transformation • Survival studies
Risk Factors for thrombosis in ET Patients (n=439) Hazard ratio p (95%CI) value Standard risk factors 1.8 (1.1-3.0) 0.04 WBC of at least 8.7 x 10 9 /L 1.6 (0.9-2.8) 0.06 Platelet at least 784 x 10 9 /L 0.9 (0.5-1.6) NS JAK2V617F 1.4 (0.7-3.0) NS Carobbio A, Blood 2007
Complication Rate in JAK2-positive ET Patients V617F Wt p value n=414 n=362 Arterial thrombosis 38 24 NS In year before diagnosis After trial entry* 25 21 NS Venous thromboembolism 11 2 0.04 In year before diagnosis After trial entry* 12 4 0.06 Major haemorrhage 18 14 NS Death 34 23 NS *After entry into one of three prospective multicentre studies of high-, medium- and low-risk ET patients. Campbell PJ, Lancet 2005
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