1. Myeloproliferative disorders
Clonal hematopoiesis (stem cell disorder)
Marrow hypercellularity
Overproduction of one or more cell lines (effective hematopoiesis)
Exception: myelofibrosis
Differentiation nearly normal

2. The “common” bcr-abl negative myeloproliferative disorders
Polycythemia vera
Essential thrombocythemia
Myelofibrosis/myeloid metaplasia

3. Less common conditions
Undifferentiated myeloproliferative disorder
Mast cell disease
Hypereosinophilic syndrome
Chronic neutrophilic leukemia
Myelodysplastic/myeloproliferative disorders
CMML
Atypical CML (BCR-ABL negative)
Juvenile CML
“Unclassifiable” MDS/MPD

4. JAK-2
Tyrosine kinase involved in signaling pathways initiated by EPO, TPO, G-CSF and other growth factors
V617F mutations in most cases of PV, ET, MF
Other JAK-2 mutations (exon 12, etc) in a minority of PV cases (almost all cases of PV have a JAK-2 mutation of some type)
Some cases of ET, MF lack JAK-2 mutation
Mutation releases cells from dependence on growth factors
Homozygosity for mutation seen mainly in P vera, associated with disease progression
Specific inhibitors of the kinase now in clinical trials

5. Structure-function relationships in the JAK2 receptor
Science 2014; 344:703

6. In Panel A, in the absence of ligand, the erythropoietin receptor (EPOR) binds JAK2 as an inactive dimer. In cells with wild-type JAK2 protein, the binding of erythropoietin (Epo) to its receptor induces conformational changes in the receptor, resulting in phosphorylation (P) of JAK2 and the cytoplasmic tail of the receptor. This leads to signaling through pathways made up of Janus kinases and signal transducers and activators of transcription (JAK–STAT), phosphatidylinositol 3 kinase (PI3K), and RAS and mitogen-activated protein kinase (RAS–MAPK). In cells with the V617F mutation, the signaling is constitutively increased, even in the absence of erythropoietin. In Panel B, the JAK2 protein binds to multiple cytokine receptors — EPOR, thrombopoietin receptor (MPL), granulocyte colony-stimulating factor receptor (G-CSFR), and probably others — that are important for hematopoietic stem-cell biology and differentiation. Therefore, the JAK2 protein with the V617F mutation exerts its effects at various stages of differentiation and in various lineages. In Panel C, the development of homozygosity for the V617F mutation is a two-step process, with the initial point mutation followed by mitotic recombination of chromosome 9p between the JAK2 locus and the centromere. This results in the loss of heterozygosity but a diploid DNA copy number.
NEJM 2006; 355:2452

8. NEJM 2006; 355:2452

9. Polycythemia vera Elevated RBC mass, typically high platelets and WBC
Some patients present with thrombocytosis and develop erythrocytosis subsequently
Hypercellular marrow with variable degree of reticulin fibrosis
Morphology fairly normal, some clustering and dysmorphism of megas
Splenomegaly in 70%, constitutional sx, increased risk of arterial & venous thrombosis (esp portal vein), microvascular disease (erythromelalgia, pruritus, headache, etc), hypermetabolic sx & gout

10. Differential diagnosis of erythrocytosis
H&P
COPD or other possible causes of hypoxemia?
Smoker?
Splenomegaly, pruritus, erythromelalgia?
Concomitant thrombocytosis and/or leukocytosis?
Serum EPO
JAK-2 mutation testing

11. Polycythemia Vera Natural History
Life expectancy decreased (3 deaths/100 pts/yr)
Cardiovascular mortality increased 1.4x
Major thrombosis in 3/100pts/yr
Risk of death from leukemia increased 36-fold
Progressive myelofibrosis

12. NEJM 2004; 350:99

13. NEJM 2004; 350:99

14. Oxygen delivery vs Hematocrit
J Clin Invest 1963;42:1150

15. Vaso-occlusion in P vera

16. Thrombosis in myeloproliferative disorders
41% of deaths in PV from cardiovascular causes
15% coronary dz
8% CHF
8% non hemorrhagic stroke
8% PE
Complex pathophysiology
Abnormal RBC/WBC/plt function
Activated PMNs/cytokines affect vascular endothelium
Prothrombotic microparticle release
Increased whole blood viscosity

17. Risk factors for thrombosis in myeloproliferative disorders
Hx of thrombosis
Disease category (PV > ET, MF)
Cardiovascular risk factors (lipids, blood pressure, smoking, diabetes)
Age
OCP therapy (splanchnic vein clots)
JAK2 mutation status/allele burden
High hematocrit (PV) & WBC
Extreme thrombocytosis increases bleeding risk
Thrombophilic genetic traits?

18. Prothrombotic effects of erythrocytosis
Barbui et al. Blood 2013;122:

19. Hematology 2005:201

20. Recommendations for treatment of patients with polycythemia vera
Keep Hct < 45 (plebotomy)
Low dose ASA unless contraindicated
Manage reversible risk factors (BP, lipids, smoking)
Consider cytoreduction if:
Intolerant to phlebotomy
Thrombocytosis or substantial leukocytosis
Symptomatic splenomegaly
Choice of cytoreductive therapy:
Age < 40: Interferon-alpha
Age > 40: hydroxyurea
Anagrelide or busulfan for patients intolerant of, or not responsive to, above choices
Hematology 2005:201

22. Aspirin vs placebo in P vera

23. Essential thrombocytosis
Thrombocytosis (typically > 600K) with variable leukocytosis, normal RBC count
About 50% have JAK2 mutation
Normal to mildly hypercellular marrow with marked increase in megakaryocytes, often with clustering
Minimal reticulin fibrosis
Splenomegaly in about 50%
Many patients asymptomatic at diagnosis
Increased risk of arterial and venous thrombosis as well as hemorrhage

24. Thrombocytosis Differential diagnosis
Essential thrombocythemia
Other MPD
P vera
Myelofibrosis
CML
Myelodysplasia (5q- et al)
Reactive
Inflammation
Surgery/trauma
Non-myeloid malignancy
Iron deficiency
Hemolysis
Acute blood loss
Absence of spleen
Rebound following thrombocytopenia

25. Hematology 2005:201

26. Essential Thrombocytosis Natural History
1-2 cases/100,000/yr
Most patients >50, but occurs in young adults
Woman more often affected than men
Life expectancy normal to slightly decreased
Patients > 60, or with prior thrombosis, at increased risk for thrombotic events
Very high platelet counts (>1.5 million) increase risk of bleeding, not thrombosis
Risk of myelofibrosis about 8% at 10 years
Risk of AML about 1% overall

27. Essential Thrombocytosis Treatment
Low-risk patients (young, no vascular risk factors) may not require treatment
Aspirin decreases thrombotic events
Hydroxyurea
Anagrelide
Interferon
Other: busulfan (leukemogenic)

28. Recommendations for treatment of patients with essential thrombocytosis
Manage reversible risk factors (BP, lipids, smoking)
If hx of thrombosis, or age >60, or plts > 1.5 million:
Hydroxyurea
Low dose ASA unless for plts > 1.5 m
2nd line: Anagrelide or interferon-alpha
Age < 60, no other risk factors:
Low dose ASA
Hematology 2005:201

30. Myelofibrosis 0.5-1.5 cases/100,000/yr Most patients >60
Marrow fibrosis with extramedullary hematopoiesis in spleen, liver, many other tissues
“Prefibrotic” stage with hypercellular marrow & abnormal megakaryocytes
Peripheral leukoerythroblastosis, teardrop cells, often with anemia. WBC and platelet counts may be high, normal or low
Splenomegaly, constitutional symptoms
Poor prognosis: marrow failure, AML in 5-30%

31. Improving survival in MF
Median survival 6.5 yrs
Median survival 4.5 yrs
J Clin Oncol 2012;30:2981

32. DDx of Marrow Fibrosis Myeloproliferative disorders
MF > > P vera, ET, CML
Other heme neoplasm
Megakaryocytic leukemia
Hodgkins
Hairy cell leukemia
Non-heme cancer
Non-malignant conditions
Renal osteodystrophy
Autoimmune disease
Vitamin D deficiency

33. SPLENOMEGALY IN MYELOFIBROSIS
Mayo Clin Proc 2004;79:503
SPLENOMEGALY IN MYELOFIBROSIS

34. Myelofibrosis Treatment
Supportive care
Transfusions
Splenectomy
Thalidomide/lenalidomide (low dose)
Reduce transfusion requirements, reduce spleen size
Chemotherapy (limited data; myelosuppression often dose-limiting)
Cladribine
Azacytidine/decitabine
Low dose cytarabine
Marrow transplant in selected patients
JAK inhibitor therapy: Ruxolitinib (inhibits JAK-1 and JAK-2)

35. Ruxolitinib treatment of myelofibrosis
Spleens shrink
Symptoms improve
Better survival
NEJM 2012;366:799

36. Eosinophilia Non-clonal/secondary (common) Clonal
Idiopathic hypereosinophilic syndrome
Familial (very rare)
Mild: AEC
Moderate: AEC
Severe: AEC > 5000

37. Hypereosinophilia (AEC > 5000)
Parasitic infection
Drug reaction
Allergic gastroenteritis
Eosinophilic fasciitis
Churg-Strauss syndrome
Pulmonary eosinophilia
Eosinophilia-myalgia syndrome
IL-2 therapy
Hyper-IgE syndrome
AML
Eosinophilic leukemia
Hypereosinophilic syndrome

38. Clinical manifestations of hypereosinophilia
Skin: pruritus, angioedema, ulcers, papular or nodular lesions
Heart: endocardial fibrosis, valvular disease, mural thrombi, cardiomyopathy, incr troponin
Lungs: infiltrates, nodules, pleural effusion
Neurologic: poly- or mononeuropathy, transverse myelitis, optic neuritis, CNS vasculitis
GI: hepatosplenomegaly, gastroenteritis, sclerosing cholangitis
Heme: marrow fibrosis, cytopenias
Kidney: thrombotic microangiopathy

39. Secondary Eosinophilia
Infection
Usually parasitic
Allergy
Asthma, allergic rhinitis, dermatitis, urticaria/angioedema, drug reaction
Autoimmune/inflammatory disorders (many)
Paraneoplastic
Solid tumors, lymphomas (Hodgkin>NHL)
Endocrinopathy
Adrenal insufficiency, growth hormone deficiency
Clonal T-cell disorder (→ IL-5)
Most cases with IL-5 overproduction

40. Clonal Eosinophilia Acute leukemia (AML>ALL)
Chronic myeloid disorders
CML (bcr-abl)
Systemic mastocytosis (c-Kit)
8p11 myeloproliferative disorder (FGFR-1)
PDGFRA and PDGFRB mutations
“Classic” MPD (JAK-2)
MDS
Chronic eosinophilic leukemia

41. Clonal eosinophilic disorders caused by mutations in tyrosine kinase genes
PDGFRA, PDGFRB
May be cytogenetically silent→ FISH testing for dx
Marrow fibrosis, incr mast cells, elevated tryptase → variant of mast cell dz?
Male predominance
Imatinib-responsive ( mg/d)
FGFR1
Associated with 8p11 translocations
Stem cell disorder → aggressive MPD associated with T-lymphoblastic lymphoma
High dose chemotherapy + allo-HSCT

42. Chronic eosinophilic leukemia WHO diagnostic criteria
Eosinophil count > 1500
No Ph chromosome or BCR-ABL fusion
No PDGFRB, PDGFRA or FGFR1 rearrangement
Blast count in blood and marrow < 20%
No inv (16) or other variant characteristic of AML
Presence of clonal cytogenetic or molecular abnormality OR blasts >2% in blood or >5% in marrow

43. Idiopathic Hypereosinophilic Syndrome
Persistent moderate or severe eosinophilia (AEC > 1500) with end-organ damage
Other causes of eosinophilia ruled out
Male predominance
Can evolve into overt myeloid or lymphoid neoplasm
Potentially fatal – 10 yr survival < 50%
Treatment
Asymptomatic: corticosteroids vs watchful waiting
Symptomatic patients: corticosteroids, IFNα, hydroxyurea, imatinib, various myelosuppressive agents

44. Laboratory evaluation of hypereosinophilia
Marrow biopsy with cytogenetics
FISH for PDGFR mutations
Serum tryptase
T-cell immunophenotyping and gene rearrangement study
Serum IL-5
Serum IgE
Screen for organ damage
Echo, troponin level
CXR, PFTs

45. Classification of Mast Cell Disease WHO classification and frequency in Mayo Clinic series
Indolent – often limited to skin (urticaria pigmentosa (46%)
Associated with myeloproliferative disorder (40%)
“Aggressive” mastocytosis – with lymphadenopathy & eosinophilia (12%)
Mast cell leukemia (1%)
Blood 2009;113:5727

46. WHO Diagnostic Criteria for Mast Cell Disease
Major: multifocal dense infiltrates of mast cells (15+) in marrow or other extracutaneous organs
Minor
>25% spindle shaped or otherwise atypical MC
c-KIT codon 816 mutation
MC in marrow or other extracutaneous organ express CD2 and/or CD25
Serum tryptase persistently >20 ng/ml in the absence of another clonal myeloid disorder
Diagnosis requires major + 1 minor criterion or at least 3 minor criteria

47. Stuff in mast cell granules that could make you sick
Metcalfe, Blood 2008;112:946

48. Clinical & laboratory features of mast cell disease
Skin: Urticaria pigmentosa, Darier sign (dermatographism), telangiectasias
Lymphadenopathy
Hepatosplenomegaly
Constitutional symptoms
Recurrent anaphylaxis
GI symptoms (N/V, diarrhea, abd pain, peptic ulcer)
Elevated serum/urinary histamine levels
Elevated serum tryptase (96%)
Cytopenias
Eosinophilia
Coagulopathy (heparin in mast cells) – rare
Lytic or blastic bone lesions

49. Blood 2009; 113: 5727
ISM: Indolent MCD
SM-ANMD: associated with clonal non-mast cell lineage disease
ASM: Aggressive MCD
MCL: Mast cell leukemia

50. Mastocytosis in bone marrow
H&E
Tryptase

51. Mastocytosis in bone marrow

52. Skin findings in mast cell disease
Dermatographism
Urticaria pigmentosa
Telangiectasia macularis eruptiva perstans

53. Molecular biology of mast cell disease
A majority of patients have D816V mutation in c-KIT tyrosine kinase gene (imatinib-insensitive)
A minority have other mutations, may be imatinib-sensitive
Occasionally found together with JAK2 V617F
Some patients have eosinophilia & PDGFRA mutation (imatinib-responsive)

54. Treatment of mast cell disease
Antihistamines, PPIs
Vit D/Calcium, bisphosphonates
Glucocorticoids
Epinephrine (for anaphyactic reactions)
For aggressive disease:
Interferon
Imatinib (if there is an imatinib-sensitive mutation)
Other TKIs (dasatinib, nilotinib)?
Cladribine
Combination chemotheapy
Allo-HSCT

55. Survival of patients with MCD
Blood 2009; 113: 5727