Includes the new updates by World Health Organisation in brief.

1. RECENT CHANGES IN
MYELOPROLIFERATIVE NEOPLASMS
PRESENTED BY- DR. NILADRI SARKAR
MODERATED BY- DR. T.K. DAS, HOD, DEPT. OF
PATHOLOGY, RGKMCH
1

2. INTRODUCTION
▸ MPN refers to clonal disorders of haemopoiesis that lead
to an increase in the numbers of one or more mature
blood cell progeny.
▸ The proliferation is associated with relatively normal
maturation that is effective, resulting in increased numbers
of granulocytes, red blood cells, and/or platelets in the
peripheral blood, in contrast to ineffective haematopoiesis
observed in myelodysplastic syndrome.
▸ Splenomegaly and hepatomegaly are common occurrence
in these set of diseases.
2

3. UPDATES
▸ The World Health Organisation (WHO) classification of
tumours of haematopoietic and lymphoid tissue was last
updated in 2008.
▸ There has been numerous advances in the identification of
unique biomarkers associated with some myeloid
neoplasms and acute leukemias, largely derived from
gene expression analysis and next generation sequencing.
▸ This can significantly improve diagnostic criteria as well as
the prognostic relevance of entities.
▸ The 2016 edition represents a revision of the prior
classification rather than an entirely new classification.
3

4. ▸ Attempts to incorporate new clinical, prognostic,
morphologic, immunophenotypic, and genetic data that
have emerged since the last edition.
4

5. CLASSIFICATION
▸ Chronic myeloid leukaemia(CML), BCR-ABL1 positive
▸ Chronic neutrophilic leukaemia
▸ Polycythaemia vera(PV)
▸ Primary myelofibrosis(PMF)
PMF, prefibrotic/ early stage
PMF, overt fibrotic stage
▸ Essential thrombocythaemia(ET)
▸ Chronic eosinophilic leukaemia, not otherwise specified
▸ MPN, unclassifiable
Mastocytosis, however, is no longer considered a subgroup of the MPNs.
5

6. CHRONIC
“MYELOID”
LEUKEMIA
6

7. CHRONIC MYELOID LEUKAEMIA
▸ Name change to “MYELOID”.
▸ Originates in an abnormal pluripotent bone marrow stem
cell and is consistently associated with the
Philadelphia(Ph) chromosome.
▸ Initial major finding is neutrophilic leukocytosis, abnormal
fusion gene is found in all myeloid lineage as well as in
some lymphoid cells.
▸ The disease is bi- or triphasic: an initial indolent chronic
phase(CML-CP) is followed by one or both of the
aggressive transformed stages, accelerated phase(CML-
AP) and blast phase(CML-BP).
7

8. CHRONIC MYELOID LEUKAEMIA
▸ Epidemiology
Can occur at any age, but the median age at diagnosis is
in the fifth and sixth decades of life, with slight male
predominance.
▸ Site of involvement
Haematopoietic tissue, primarily the blood, bone
marrow, spleen, and liver. During CML-BP,
extramedullary tissues, including lymph nodes, skin, soft
tissue and CNS show infiltration by blasts.
8

9. The peripheral blood demonstrates neutrophilia with a left shift, eosinophilia and basophilia. Typically
myelocytes predominate. Wright-Giemsa, 500×.
CHRONIC MYELOID LEUKAEMIA
9

10. The BMB is hypercellular with myeloid and megakaryocytic hyperplasia. Clusters of small hypolobated
megarkyocytes seen. Hematoxylin and eosin, 500×.
CHRONIC MYELOID LEUKAEMIA
10

11. CHRONIC MYELOID LEUKAEMIA
▸ Morphology: CHRONIC PHASE
▸ Most cases can be diagnosed from peripheral blood findings
combined with detection of t(9;22)(q34.1;q11.2) or,
specifically, BCR-ABL1 by molecular genetic techniques.
▸ PB shows leucocytosis(median 170x109
/L), blasts usually 2%,
absolute basophilia invariably present. Eosinophilia in some
cases. Platelet count remains normal or increased.
▸ Bone marrow biopsy is hyper cellular, with blasts fewer than
5%. Megakaryocytes are smaller with hypolobated nuclei.
Reticulin fibres are increased in 40% cases.
11

12. CHRONIC MYELOID LEUKAEMIA
▸ BM aspirate is essential to ensure sufficient material for
complete karyotype and morphologic evaluation(to
confirm the phase)
▸ Regular monitoring for BCR-ABL1 burden and for
evidence of genetic evolution and development of
resistance to TKI therapy is essential (to detect disease
progression)
12

13. CHRONIC MYELOID LEUKAEMIA
▸ Morphology: ACCELERATED PHASE
▸ Less common in the era of TKI therapy.
▸ The criteria for AP in revised WHO Classification include
haematologic, morphologic, and cytogenetic parameters
which are supplemented by additional parameters usually
attributed to genetic evolution, and manifested by
evidence of resistance to TKIs.
▸ The ‘response to TKI therapy' criteria for AP are
considered as “provisional” until further supported by
additional data.
13

14. CHRONIC MYELOID LEUKAEMIA
Criteria for CML, accelerated phase
Any 1 or more of the following criteria or response to TKI criteria:
▸ Persistent or incurring WBC(>10x109
/L),
unresponsive to therapy
“Provisional” response to TKI criteria
▸ Persistent or increasing splenomegaly,
unresponsive to therapy
▸ Hematologic resistance to the first TKI or
failure to achieve a complete hematologic
response or
▸ Persistent thrombocytosis(>1000x109
/L),
unresponsive to therapy
▸ Any haematological, cytological, or molecular
indications of resistance to 2 sequential TKIs
or
▸ Persistent thrombocytopenia(<100x109
/L),
unrelated to therapy
▸ Occurence of 2 or more mutations in BCR-
ABL1 during TKI therapy
▸ 20% or more basophils in the PB
▸ 10-19% blasts in the PB and/or BM
▸ Additional clonal chromosomal abnormalities
in Ph positive cells at diagnosis
▸ Any new clonal chromosomal abnormality in
Ph positive cells that occurs during therapy
14

15. CHRONIC MYELOID LEUKAEMIA
▸ COMPLETE HAEMATOLOGICAL RESPONSE:
WBC <10 x 109
/L
Platelet count <450 x 109
/L
No immature granulocytes in the differential, and
Spleen non palpable
15

16. CHRONIC MYELOID LEUKAEMIA
▸ Morphology: BLAST PHASE
▸ Requires either at least 20% blasts in the blood or BM or
the presence of an extramedullary accumulation of blasts.
▸ The finding of any bona fide lymphoblast in the blood
or marrow, even if <10% should prompt concern that
lymphoblastic transformation may be imminent and
warrants further clinical and genetic investigation.
16

17. CHRONIC
NEUTROPHILIC
LEUKEMIA
17

18. CHRONIC NEUTROPHILIC LEUKAEMIA
▸ Rare, characterised by sustained peripheral blood
neutrophilic, bone marrow hypercellularity due to
neutrophilic granulocyte proliferation, and
hepatosplenomegaly.
▸ No Philadelphia chromosome or BCR-ABL fusion gene.
▸ Diagnosis is one of exclusion of all causes of reactive
neutrophilic and of all other myeloproliferative diseases.
▸ The CSF3R mutation is strongly associated with CNL.
18

19. CHRONIC NEUTROPHILIC LEUKAEMIA
▸ Epidemiology
True incidence is unknown, generally affects older adults, but
has been reported in adolescents. The sex distribution is
nearly equal.
▸ Site of involvement
Blood and BM are always involved, spleen and liver shows
leukaemic infiltrates.
▸ Clinical features
Splenomegaly, hepatomegaly with history of bleeding from
much cutaneous surfaces or GIT, gout and pruritus are other
possible symptoms.
19

20. CHRONIC NEUTROPHILIC LEUKAEMIA
Diagnostic criteria for CNL
1. PB WBC ≥ 25 x 109
/L
Segmented neutrophils plus band forms ≥ 80% of WBCs
Neutrophilic precursors (promyelocytes, myelocytes, and metamyelocytes) <10% of WBC
Myeloblasts rarely observed
Monocyte count <1 x 109
/L
No dysgranulopoiesis
2. Hypercellular BM
Neutrophilic granulocytes increased in percentage or number
Neutrophil maturation appears normal
Myeloblasts <5% of nucleated cells
3. Not meeting WHO criteria for BCR-ABL1 Positive CML, PV, ET, or PMF
4. No rearrangement of PDGFRA, PDGFRB, or FGFR1, or PCM1-JAK2
5. Presence of CSF3R T618I or other activating CSF3R mutation
OR
20

21. CHRONIC NEUTROPHILIC LEUKAEMIA
Diagnostic criteria for CNL
In the absence of a CSF3R mutation, persistent neutrophilia (at least 3
months), splenomegaly and no identifiable cause of reactive neutrophilia
including absence of plasma cell neoplasm or, if present, demonstration of
clonality of myeloid cells by cytogenetic or molecular studies.
21

22. POLYCYTHAEMIA
VERA
22

23. POLYCYTHAEMIA VERA
▸ Characterised by increased red blood cell production that is
independent of the mechanisms that normally regulate
erythropoiesis.
▸ Two phases- polycythaemic phase, associated with an increased red
cell mass and a “spent” or post polycythaemic phase, in which
cytopenias, including anaemia are associated with BM fibrosis and
hypersplenism.
▸ All causes of secondary erythrocytosis as well as inheritable
polycythaemia must be excluded prior to making the diagnosis of PV
▸ PV is possibly under diagnosed using the haemoglobin levels of
previous edition and the utility of BM morphology as a
reproducible criterion for the diagnosis of PV is recognised.
23

24. POLYCYTHAEMIA VERA
▸ Epidemiology
Mean age at diagnosis is 60 years with a slight male
predominance.
▸ Site of involvement
Blood and BM being the major site, spleen and liver are
also affected.
▸ Clinical features
Related to hypertension or to vascular abnormalities
caused by increased red cell mass. Venous or arterial
thrombosis, headache, visual disturbances, paraesthesias.
24

25. POLYCYTHAEMIA VERA
▸ Morphology: POLYCYTHAEMIC PHASE
PB- Normocytic, normochromic. Neutrophilia and basophilia are
common.
Normoblastic erythroid marrow proliferation.
Hypercellular for age BMB. Panmyelosis account for increased
cellularity. Megakaryocytes are conspicuous. They tend to
cluster around marrow sinusoids and show pleomorphic aspect.
Silver stains show a normal reticulin finer network(70% cases).
Stainable iron is lacking in more than 95% cases.
Spleen and liver show mainly congestion.
25

26. POLYCYTHAEMIA VERA
▸ Morphology: SPENT PHASE
▸ Later stages of PV, red cell mass normalises and the decreases and
spleen enlarges further.
▸ Most common pattern of progression is post-polycythaemic
myelofibrosis and myeloid metaplasia (PPMM).
▸ It is characterised by a leukoerythroblastic blood smear, RBC
poikilocytosis with tear drop shaped cells, and splenomegaly.
Hallmark of this state is reticulin and even collagen fibrosis of
marrow.
▸ More than 10% blasts in blood/ marrow unusual and signals
transformation to a myelodysplastic syndrome or acute leukemias.
26

27. POLYCYTHAEMIA VERA
WHO criteria for PV
Major criteria
1. Hemoglobin >16.5 g/dl in men, >16.0 g/dl in women or,
Hematocrit >49% in men, >48% in women or,
Increased red cell mass (RCM) {More than 25% above mean normal predicted value}
2. BM biopsy showing hypercellularity for age with trilineage growth(panmyelosis) and
megakaryocytic proliferation with pleomorphic, mature megakaryocytes.*
3. Presence of JAK2V617F or JAK2 exon 12 mutation.
Minor criteria
Subnormal serum erythropoietin level.
DIAGNOSIS OF PV REQUIRES EITHER ALL 3 MAJOR CRITERIA OR FIRST 2 MAJOR AND MINOR
CRITERION
* BM biopsy may not be required in cases with haemoglobin levels >18.5 g/dl in men, >16.5
g/dl in women if major criterion 3 and the minor criterion are present.
27

28. The BMB is hypercellular, with panmyelosis, and increased number of erythroid precursors, and large
atypical megakaryocytes. Hematoxylin and eosin, 200×.
POLYCYTHAEMIA VERA
28

29. BMB showing increased fibrosis and osteosclerosis. Hematoxylin and eosin, 400×.
POLYCYTHAEMIA VERA(POST
POLYCYTHAEMIC PHASE)
29

30. BMB showing increased fibrosis and osteosclerosis. Hematoxylin and eosin, 400×.
POLYCYTHAEMIA VERA(POST
POLYCYTHAEMIC PHASE)
30

31. PBS showing numerous giant platelets, myelocyte, lymphocyte and cluster of blasts. Wright-
Giemsa, 600×.
TRANSFORMATION OF POLYCYTHAEMIA
VERA
31

32. ESSENTIAL
THROMBOCYTHEMIA
32

33. ESSENTIAL THROMBOCYTHAEMIA
▸ Clonal myeloproliferative disease that involves primarily
the megakaryocytic lineage.
▸ characterised by sustained thrombocytosis in the blood
and increased number of large, mature megakaryocytes in
the BM, and clinically, by episodes of thrombosis and/or
haemorrhage.
▸ It is necessary to differentiate “true” essential
thrombocythaemia(ET) from prefibrotic/ early primary
myelofibrosis(prePMF) by, among other features, the
morphologic findings in the BM biopsy, including the lack
of reticulin fibrosis at onset. (Prognostic implications)
33

34. ESSENTIAL THROMBOCYTHAEMIA
▸ Epidemiology
Most cases are diagnosed at 50-60 years of age with no
major predilection. A second peak occurs at 30 years of age,
and them women are more often affected.
▸ Site of involvement
Blood and BM being the principal sites. Spleen shows
minimal extramedullary hematopoiesis.
▸ Clinical features
More than half are asymptomatic but 20-25% cases have
manifestations of vascular occlusion or haemorrhage.
Modest splenomegaly in 50% cases.
34

35. Blood smear reveals anisocytosis and a greatly increased number of platelets.
ESSENTIAL THROMBOCYTHEMIA
35

36. BMB is mildly hypercellular with atypical megakaryocyte clustering. Hematoxylin and eosin,
500×.
ESSENTIAL THROMBOCYTHEMIA
36

37. ESSENTIAL THROMBOCYTHAEMIA
WHO criteria for ET
MAJOR CRITERIA
1. Platelet count ≥ 450 x 109
/L
2. BM biopsy showing proliferation mainly of the megakaryocyte lineage with increased numbers
of enlarged, mature megakaryocytes with hyperlobated nuclei. No significant increase or left shift
in neutrophil granulopoiesis or erythropoiesis and very rarely minor increase in reticulin fibres.
3. Not meeting WHO criteria for BCR-ABL1 Positive CML, PV, PMF, myelodysplastic syndromes, or
other myeloid neoplasms.
4. Presence of JAK2, CALR, or MPL mutation.
MINOR CRITERIA
Presence of a clonal marker or absence of evidence of reactive thrombocytosis.
DIAGNOSIS OF ET REQUIRES MEETING ALL 4 MAJOR CRITERIA OR THE FIRST 3 MAJOR
CRITERIA AND THE MINOR CRITERION.
37

38. PRIMARY
MYELOFIBROSIS
38

39. PRIMARY MYELOFIBROSIS
▸ Characterised by proliferation of mainly megakaryocytic and
granulocytic elements in the BM, associated with active
deposition of BM connective tissue and with extramedullary
haematopoiesis (EMH).
▸ Stepwise evolution of the disease process with an initial
prefibrotic state that is characterised by a hypercellular BM
with minimal reticulin fibrosis that merges into a fibrotic stage,
wherein, the BM demonstrates marked reticulin or collagen
fibrosis and often osteomyelosclerosis.
▸ The minor clinical criteria in prePMF that may have a major
impact on diagnosis and prognosis has been explicitly
defined.
39

40. PRIMARY MYELOFIBROSIS
▸ Epidemiology
Commonly in the seventh decade.
▸ Site of involvement
Blood and BM always involved. Spleen and liver most
common sites of EMH. Other sites like lymph nodes, kidney,
adrenal gland, dura mater, breast and skin.
▸ Clinical features
30% cases asymptomatic and are discovered by detection of
splenomegaly or routine PB shows anaemia or
thrombocytosis. Fatigue, weight loss, night sweats, bleeding.
40

41. PBS showing leukoerythroblastosis with rare blasts, nucleated red blood cells, large platelets,
and teardrop shaped red cells. Wright-Giemsa, 1,000×.
PRIMARY MYELOFIBROSIS
41

42. PRIMARY MYELOFIBROSIS
WHO criteria for prePMF
MAJOR CRITERIA
1. Megakaryocytic proliferation and atypia, without reticulin fibrosis > grade 1, accompanied by
increased age-adjusted BM cellularity, granulocytic proliferation, and often decreased
erythropoiesis.
2. Not meeting WHO criteria for BCR-ABL1 Positive CML, PV, ET, myelodysplastic syndromes, or
other myeloid neoplasms.
3. Presence of JAK2, CALR, or MPL mutation or in absence of these, presence of clonal markers, or
absence of minor reactive BM reticulin fibrosis.
MINOR CRITERIA
Presence of at least 1 of the following, confirmed in 2 consecutive determinations:
a. Anemia not attributed to a comorbid condition
b. Leukocytosis ≥ 11 x 109
/L
c. Palpable splenomegaly
d. LDH increased to above upper normal limit of institutional reference range
DIAGNOSIS OF prePMF REQUIRES MEETING ALL 3 MAJOR CRITERIA, AND AT LEAST 1 MINOR
CRITERION
42

43. PRIMARY MYELOFIBROSIS
WHO criteria for overt PMF
MAJOR CRITERIA
1. Megakaryocytic proliferation and atypia, accomapianed by either reticulin and/or collagen
fibrosis grades 2 or 3
2. Not meeting WHO criteria for BCR-ABL1 Positive CML, PV, ET, myelodysplastic syndromes, or
other myeloid neoplasms.
3. Presence of JAK2, CALR, or MPL mutation or in absence of these, presence of clonal markers, or
absence of reactive myelofibrosis.
MINOR CRITERIA
Presence of at least 1 of the following, confirmed in 2 consecutive determinations:
a. Anemia not attributed to a comorbid condition
b. Leukocytosis ≥ 11 x 109
/L
c. Palpable splenomegaly
d. LDH increased to above upper normal limit of institutional reference range
e. Leukoerythroblastosis
DIAGNOSIS OF prePMF REQUIRES MEETING ALL 3 MAJOR CRITERIA, AND AT LEAST 1 MINOR
CRITERION
43

44. GRADING OF MYELOFIBROSIS
Semiquantitative grading of BM fibrosis(MF) with minor
modifications concerning collagen and osteosclerosis. Fiber
density should be assessed only in haematopoietic areas.
MF- 0
Scattered linear reticulin with no
intersections(crossovers) corresponding to
normal BM
MF- 1
Loose network of reticulin with many
intersections, especially in perivascular areas
MF- 2
Diffuse and dense increase in reticulin with
extensive intersections, occasionally with focal
bundles of thick fibers mostly consistent with
collagen, and/or focal osteosclerosis **
MF- 3
Diffuse and dense increase in reticulin with
extensive intersections and coarse bundles of
thick fibers consistent with collagen, usually
associated with osteosclerosis **
** In grades MF- 2 and MF- 3 an additional trichrome stain is recommended.
44

45. M/L NEOPLASMS
ASSOCIATED WITH
EOSINOPHILIA
45

46. MYELOID/ LYMPHOID NEOPLASMS ASSOCIATED WITH EOSINOPHILIA AND
REARRANGEMENT OF PDGFRA, PDGFRB, OR FGFR1 OR WITH PCM1-JAK2
▸ The criteria for the diagnosis of the eosinophilia-related
proliferations associated with specific molecular genetic
changes are retained in the classification.
▸ Although it is noted that eosinophilia may be absent in a
subset cases.
▸ In the 2016 revision, this disease group will
incorporate the myeloid neoplasm with t(8;9)
(p22;q24.1A); PCM1-JAK2 as a new provisional entity.
46

47. M/LN-E AND ABNORMALITIES 47

48. M/LN-E AND ABNORMALITIES
NEW ENTITY:
Disease Presentation Genetics Treatment
PCM-JAK2
Eosinophilia
Rarely presents
with T-LBL or B-
ALL
Bone marrow
shows left shifted
erythroid
predominance
and lymphoid
aggregates
t(8;9)(p22q24.1)
PCM-JAK2
May respond to
JAK2 inhibitors.
48

49. REFERENCES
▸ The updated WHO classification of haematological
malignancies- bloodjournal.org
▸ The Hematologist- American Society of Hematology
▸ WHO Classification of Tumors- Tumors of Hematopoietic
and Lymphoid tumors
▸ Wintrobe’s Clinical Hematology- 13th edition
▸ Leukemia Classification 2016: What, when, who -
California Society of Pathologists
49

50. SUMMARY OF CHANGES
▸ Presence of even low numbers of bona fide lymphoblast- imminent blast phase.
▸ Revised criteria for diagnosing accelerated phase CML.
▸ CNL now includes specific mention of CSF3R T6181I as a major diagnostic
criterion.
▸ Diagnostic criteria for PV are notably changed to lower the hemoglobin
threshold and thus prevents underdiagnosis. Bone marrow morphology is now
considered a major criteria.
▸ The diagnostic criteria for ET now adds CALR and MPL mutations, a change that
also affects PMF.
▸ The criteria for prefibrotic PMF as opposed to overt PMF further clarified.
▸ Removal of mastocytosis.
▸ Inclusion of PCM1-JAK2 rearrangement in myeloid/lymphoid neoplasms
associated with eosinophilia.
50

51. THANK YOU
Keynote Presentation