European vs 2015-World Health Organization clinical molecular and pathological classification of myeloproliferative neoplasms

Table 1 The 1980 Rotterdam clinical and pathological criteria for essential thrombocythemia and polycythemia vera

1A The 1980 RCP major (A) and confirmative (B) criteria for prefibrotic ET
A1 Persistent platelet count in excess of 400 × 109/L
A2 Increase and clustering of enlarged megakaryocytes in bone marrow biopsy
A3 No or slight increase of reticulin fibers (RF 0 or RF 1)
B1 Presence of large platelets in a peripheral blood smear
B2 Absence of any underlying disease for reactive thrombocytosis and normal ESR
B3 No splenomegaly (< 12 cm) or slight splenomegaly on palpation or scan (< 15 cm)
B4 Increase of LAP-score and no signs of fever or inflammation
Exclusion criterion
Ph+ chromosome and any other cytogenetic abnormality in blood or bone marrow cells
1B The 1980 RCP major (A) and minor (B) criteria for prefibrotic PV
A1 Raised red cell mass. Male > 36 mL/kg, female > 32 mL/kg consistent with erythrocyte count of > 6 × 1012/L (Dameshek[1,2])
A2 Absence of primary or secondary erythrocytosis by clinical and laboratory tests
A3 Slight, moderate or marked increase in bone marrow biopsy of clustered, enlarged pleomorphic megakaryocytes with hyperlobulated nuclei and moderate to marked increase cellularity of megakaryopoiesis/erythropoiesis or typically trilinear mega-erythro-granulopoiesis. A typical PV bone marrow excludes erythrocytosis. No or presence of reticuline fibers and no collagen fibers (no dry tap)
B1 Thrombocythemia, persistant increase of platelet > 400 × 109/L
B2 Leukocytosis, leucocyte count > 109/L and low erythrocyte sedimentation rate
B3 Raised leukocyte alkaline phosphatase score > 100, absence of fever or infection
B4 Splenomegaly on palpation or on isotope/ultrasound scanning
A1 + A3 plus one of B establishes PV and excludes any variant of erythrocytosis
1C Grading of bone marrow biopsy content of RF according to Ellis et al[41], Georgii et al[35,36] and Wilkins et al[171] and WHO grading of MF[98-101]
Grading RF[41]	Grading MF 2008 WHO91	Description of RF and reticulin/collagen fibers in MF as a secondary event in MPN
Normal RF 0	N MF 0	No reticulin fibers, occasional individual fibers or focal areas with tiny amount of reticulin fiber network
Slight increase RF 1	+ MF 0	Fine reticulin fiber network throughout much of section and no course reticulin fibers
Moderate increase RF 2	++ MF 1	Diffuse fine reticuline network with focal collections of thick course reticulin fibers and no collagenisation
Marked increase RF 3	+++ RCF = MF 2	Diffuse and dense increase in reticulin with extensive intersections, and presence of collagen fibers and no or minor O
OS Dry tap RF 4	Sclerotic RCF and O = MF 3	Diffuse and dense reticulin with with coarse bundles of collagen associated with significant O

MF: Myelofibrosis; MPN: Myeloproliferative neoplasms; O: Osteosclerosis; RCP: Rotterdam clinical and pathological; ET: Essential thrombocythemia; RF: Reticulin fibrosis; PV: Polycythemia vera.

Table 2 Polycythemia vera study group criteria for polycythemia vera[10] and diagnostic differentiation of polycythemia vera from all variants of primary and secondary erythrocytoses by bone marrow histology[17]

Major criteria PV	Minor criteria PV
A1 RCM, males > 36 mL/kg females > 32 mL/kg. Hemoglobin	B1 Thrombocytosis Platelet count > 400 × 109/L
18.5 g/dL male and > 16.5 g/dL females (PVSG, WHO)	B2 Leukocytosis > 12 × 109/L
A2 Normal arterial oxygen saturation > 92%	B3 Raised neutrophil alkaline phosphatase score > 100 or raised B12
A3 Splenomegaly on palpation	(> 900 ng/L) or raised unsaturated B12 binding capacity (> 2200 ng/L)
	B3 is replaced by spontaneous EEC as a specific clue to PV
Benign erythrocytosis: 1980 RCP criteria	Myeloproliferative PV: 1980 RCP criteria
RCM, males > 36 mL/kg females > 32 mL/kg or increased erythrocytes above 6 × 1012/L	Increased red cell counts > 6 × 1012/L or increased RCM and increase of clustered large megakaryocytes with hyperlobulated nuclei is a pathognomonic diagnostic clue to PV
Normal platelet and leukocyte counts	Normal RCM = inapparent erythrocytosis is not associated with splenomegaly
Normal bone marrow histopathology: normal cellularity and erythropoiesis, and normal size, morphology and	and shows normal bone histology, whereas IPV is associated with splenomegaly and show typical features of PV bone marrow histology
distribution of megakaryocytes
Classification of erythrocytoses[74]	Notes anno 1980-1999[74]
Congenital or primary erythrocytosis including mutation	Increased RCM does not distinguish between PV and primary erythrocytosis
truncated EPO recepotor, disrupted oxygen homeostasis in	Increased RCM does not distinguish between PV and IPV. IPV is featured by
Chuvash erythrocytosis, high oxygen affinity	advanced PV with normal hb, Ht and erythrocyte count due to splenomegaly and
hemoglobinopathy, and congenital autonomous EPO	hypersplenism and with increase of reticulin fibrosis with typical PV bone
production	marrow features
Secondary erythrocytosis due to autonomous EPO production in renal diseases or by tumour cels or due to hypoxia	In IPV the values of hemoglobin hematocrit and erythrocytes are normal but RCM is increased due to splenomegaly with absence of hypervolemic symptoms
Idiopathic erythrocytoses

Diagnostic criteria for polycythemia vera (PV) proposed in 1975 by the polycythemia vera study group (PVSG)[10] and used in the 2001 and 2008 World Health Organization Classifications of MPD and MPN[75,76]. Diagnosis of PV is acceptable if the following combinations are present: A1 + A2 + A3 or A1 + A2 + any two from category B. Diagnostic differentiation of benign erthrocytosis from myeloproliferative polycythemia vera by histopathology from bone marrow sections according to the Rotterdam clinical and pathological (1980 RCP criteria for PV) (Table 1)[73,74]. RCM does not distinguish between PV and primary or secondary erythrocytosis. RCM does not distinguish between PV and inapparent PV with splenomegaly in splanchnic vein thrombosis (IPV, Table 3). In IPV RCM is increased due to splenomegaly with absence of hypervolumemic symptoms. Bone marrow histology distinguishes PV from all variants of erythrocytosis with a sensitivity and specificity of 100%. RCM: Raised red cell mass; EEC: Erythroid colony formation; IPV: Inapparent PV.

Table 3 Comparion of clinical and laboratory features between polycythemia vera study group defined polycythemia vera (group A) and inapparent polyctemia vara (group B)

Clinical feature	Group A PV	Group B IPV	P-value
No. of cases	85	18
Age (range)	61 (27-83)	52 (28-82)	NS
Sex male/female	56/42%	39/61%	NS
Splenomegaly	44 (52%)	15 (83%)	< 0.005
Leukocytes > 12 × 10/L	31 (36%)	5 (28%)	NS
Platelets > 500 × 10/L	40 (47%)	10 (56%)	NS
Red cell counts × 10/L: males	6.2 (4.9-7.4)	5.2 (4.7-5.9)	< 0.0002
Red cell mass males	48.2 (36-60)	43.3 (41-61)	NS
Red cell counts females	6 (4.2-7.3)	4.7 (3.7-5.5)	< 0.003
Red cell mass females	40.1 (32-59)	37.3 (34-46)	NS
Plasma volume PV vs IPV
Increase/theoretical norm (%)	10 (-11, 61)	36 (20, 98)	< 0.00001

Lamy et al[44]. Conclusion: inapparent polycythemia vara (IPV) is featured by normal values of haemoglobin, haematocrit and erythrocyte counts, in the absence of hypervolumic symptoms and in IPV red cell mass is increased related to the degree of increased spleen size. NS: Not significant.

Table 4 Clinical and hematological findings in thrombocythemia of various myeloproliferative diseases polycythemia vera, primary myelofibrosis and essential thrombocythemia in 395 myeloproliferaive disease patients from the Cologne Institute of Pathology 1980-1989

Diagnosis cologne criteria	PV	PMF	True ET	NV
No. of patients	55	250	40
Thrombocythemia > 500 × 109/L (%)	48	48	100	< 350
Thrombocythemia > 1000 × 109/L (%)	6	17	65
Age (median years)	63	66	58
Male/female	20/35	58/62	14/26
Platelets, × 109/L mean ± SD	808 ± 288	960 ± 361	1386 ± 541	< 350
Erythrocytes, × 10/L mean ± SD	6.7 ± 0.2	4.5 ± 0.1	4.6 ± 0.7	< 6.0
Hemoglobin, g/dL	17.7 ± 0.4	12.8 ± 0.2	13.7 ± 2
Leukocytes	17 ± 1	15 ± 9	13 ± 5
Leukocyte alkaline phosphatase score	164 ± 91	98 ± 83	57 ± 43	< 10
Spleen size increase on palpation (cm)	2.0 ± 3.3	2.6 ± 3.1	0.4 ± 0.8	NP
Observed 10 yr survival (mo)	106	85	170
Specific loss of life expectancy (%)	19	22	3
Bone marrow histopathology	PV	PMF	True ET	RT
Megakaryocytes	Pleomorph	Immature giant	Staghorn	N
Frequency/mm hematopoietic area	123/27	112/37	157/45	98/39
Size (μm2)	385 ± 102	386 ± 197	425 ± 117	328 ± 84
Erythropoiesis × 10	44 ± 8	9 ± 4	22 ± 5	27 ± 4
Granulopoiesis × 10	65 ± 12	58 ± 27	47 ± 15	57 ± 18
Reticulin fibers × 10 (mm )	21 ± 11	97 ± 41	15 ± 7	15 ± 6
No. of patients	120	40
LAP score	110 ± 60	57 ± 43	< 100
Spleen size	1.7 ± 1.4	0.4 ± 0.8	NP
Observed survival (mo)	77	170	-
Specific loss in life expectancy (%)	53	3	0

Data from Thiele et al[90]. MPD: Myeloproliferaive disease; PV: Polycythemia vera; PMF: Primary myelofibrosis; ET: Essential thrombocythemia; LAP: Leukocyte alkaline phosphatase; RT: Reactive thrombocytosis; NV: Normal value; NP: Not palpable; N: Normal.

Table 5 The 2002 European Clinical and Pathological criteria for the diagnosis of "true" essential thrombocythemia and chronic idiopathic myelofibrosis or primary megakaryocytic granulocytic myeloproliferation according to Michiels et al[91]

Clinical ECP criteria of "true" ET	Pathological ECP criteria of "true" ET
A1 Persistent increase of platelet count grade 1 400-1500 × 109/L, grade 2 > 1500 × 109/L	B1 Predominant proliferation of enlarged to giant megakaryocytes wit hyperlobulated staghorn-like nuclei and mature cytoplasm, lacking conspicious cytological abnormalities
A2 Normal spleen or only minor splenomegaly on echogram	B2 No proliferation or immaturity of granulopoisis or erythropoiesis
A3 Normal LAP score, normal ESR and increased MPV	B3 No or only borderline increase in reticulin fibers
A4 Spontaneous megakaryocyte colony formation (CFU-Meg)	The combination of A1 and B1 + B2 establish "true" ET. Any other criterion confirms ET
A5 No signs or cause of reactive thrombocytosis
A6 No preceding or allied other subtype of MPN, PV, MDS or CML
A7 Absence of Philadelphia chromosome
Clinical ECP criteria of CIMF or PMGM	Pathological ECP criteria of CIMF or PMGM
A1 No preceding or allied other subtype of MPN, PV, CML or MDS	B1 PMGM and relative or absolute reduction of erythropoiesis (erythroid precursors). Abnormal clustering and increase of atypical immature medium-sized large to giant megakaryocyte containing (Cloud-like) hypolobulated nucle and definitive maturation defects
Early clinical stage	Staging of myelofibrosis: MF in ET, PV and PMGM
Normal hemoglobin, or anemia grade 1: Hemoglobin	MF 0 No reticulin fibrosis RF 0/1
> 12 g/dL, slight or moderate splenomegaly on palpation	MF 1 Slight reticulin fibrosis RF 2
or > 11 cm on ultrasound or CT. Thrombocythemia	MF 2 Marked increase RF grade 3 and slight to moderate collagen fibrosis
around 1000 × 109/L	MF 3 Advanced collagen fibrosis-osteosclerosis (endophytic bone formation)
Intermediate clinical stage
Anemia grade 2, hemoglobin > 10 g/dL, definitive leuko-erythroblastic blood picture and/or tear-drop erythrocytes. Splenomegaly on palpation, no adverse signs
Advance clinical stage
Anemia grade 3, hemoglobin < 10 g/dL, significant splenomegaly and one or more adverse signs

ECP: European Clinical and Pathological; CIMF: Chronic idiopathic myelofibrosis; PMGM: Primary megakaryocytic granulocytic myeloproliferation; ET: Essential thrombocythemia; LAP: Leukocyte alkaline phosphatase; ESR: Erythrocyte sedimentation rate; MPV: Mean platelet volume; MPN: Myeloproliferative neoplasm; PV: Polycythemia vera; MDS: Myelodysplastic syndrome; RF: Reticulin fibrosis; CML: Chronic myeloid leukemia.

Table 6 The 2000 European Clinical and Pathological criteria for the diagnosis of polycythemia vera defined by Michiels[73] in 1997[9,74], and in 2000[93]

Clinical ECP criteria of PV	Pathological ECP criteria of PV
A1 Increased erythrocytes > 6 × 1012/L. Raised RCM: RCM (optional) male > 36 mL/kg, female > 3.2 mL/kg or increased red cell counts above 6 × 1012/L	B1 Thrombocytheia: platelet count > 400 × 109/L
A2 Absence of any cause of primary or secondary erythrocytosis by clinical and laboratory investigations	B2 Granulocytes > 10 × 109/L and raised LAP score in the absence of fever or infection
A3 Histopathology of bone marrow biopsy	B3 Splenomgaly on palpation or on echogram > 11 cm
(1) Increase and clusters of pleomorph large megakaryocytes with hyperploid nucei	B4 Spontaneous erythroid colony formation in the absence of EPO and low plasma or serum EPO level
(2) Increased cellulartity due to increased erythropoiesis or erythropoiesis and granulopoiesis (panmyelosis)	Staging of MF related to reticulin fibrosis
(3) No or slight increase of reticulin fibers	MF 0 No reticulin fibrosis RF 0/1
	MF 1 Slight reticulin fibrosis RF 2
	MF 2 Marked increase RF grade 3 and slight to moderate collagen fibrosis
	MF 3 Advanced collagen fibrosis-osteosclerosis

Diagnosis of idiopathic erythrocythemia (IE), early prodromal polycythemia vera (PV), classical PV or latent primary myeloproliferative disease (subclinical PV) are acceptable when[73,74,93]: A1 + A2 + A3 and none of B (except B4) is consistent with early erythrocythemic PV, labelled as idiopathic erythrocythemia: IE; A3 + B1 + B4 is consistent with prodromal PV with PV features in the bone marrow; A1 + A2 + A3 + plus one of B1 to 3 is consistent with classical PV; B4 confirms all variants of IE, prodromal PV and classical PV; A3 + B3 and none of the others is consistent with latent subclinical primary myeloproliferative diease (PMD) usually preceding masked cases of PV or MF during long term follow-up. ECP: European Clinical and Pathological; MF: Myelofibrosis; RF: Reticulin fibrosis; EPO: Erythropoetin; RCM: Red cell mass; LAP: Leukocyte alkaline phosphatase.

Table 7 2015 World Health Organization Clinical Molecular and Pathological criteria for the diagnosis of prodromal, masked and classical JAK2 mutated polycythemia vera vs primary or secondary erythrocytoses[77,78]

CM criteria	Bone marrow pathology (P) criteria (WHO)
Major criteria for PV	P1 Bone marrow pathology: increased cellularity (60%-100%) due to trilinear increase of erythropoiesis, megakaryopoiesis and granulopoiesis and clustering of small to giant (pleomorph) megakaryocytes with hyperlobulated nuclei
A1 Hematocrit > 0.51/> 0.48 in male/female Erythrocytes > 5.8 × 1012/L males > 5.6 × 1012/L females	Absence of stainable iron. No pronounced inflammatory reaction
A2 Presence of heterozygous and/or homozygous JAK2V617F or JAK2 exon 12 mutation	P2 Erythrocytosis. Normal erythropoiesis, normal granulopoiesis and megakaryocytes of normal size, morphology and no clustering
A3 Low serum Epo level	Grading of RF and MF
Minor	Prefibrotic: RF-0/1 = MF-0
B1 Persistent increase of platelet count × 109/L:	Early fibrotic: RF-2 = MF-1
grade 1: 400-1500, grade 2: > 1500	Fibrotic: RCF 3 = MF-2
B2 Granulocytes > 10 × 109/L or Leukocytes > 12 × 109/L and raised	Post-PV MF: RF 4 = MF-3
LAP-score or increased CD11b expression in the absence of fever or infection
B3 Splenomegaly on ultrasound echogram (> 12 cm length in diameter) or on palpation
B4 Spontaneous EEC formation (optional)

2015 WHO-CMP criteria for staging of prodromal, erythrocythemic, and advanced polycythemia vera (PV). A2 + B1 + P1 establish early PV (mimicking ET) prodromal PV CMP stage 0; A1 + A2 + A3 + P1 and none of B establish idiopathic erythrocythemia (IE) or stage 1 PV; A1 + A2 + A3 + P1 and one or more of B establish classic stages of PV stage 2 and 3; A2 + B3 + P1 detect masked cases of PV with splenomegaly and hypersplenism to be labelled as inapparent PV (IPV) frequently seen Budd-Chiari syndrome or splanchnic vein thrombosis. CM: Clinical and molecular; RF: Reticulin fibrosis; MF: Myelofibrosis; ET: Essential thrombocythemia; WHO-CMP: World Health Organization clinical molecular and pathological; EEC: Endogenous erythroid colony.

Table 8 2015 World Health Organization clinical molecular and pathobiological criteria for diagnosis of JAK2^V617F mutated essential throbocythemia[77,78]

CM criteria	Bone marrow pathology (P) criteria (WHO)
ET	Normocellular ET
(1) Platelet count of > 350 × 109/L and the presence of large platelets in a blood smear	Proliferation and clustering of enlarged mature pleomorphic megakaryocytes with hyperlobulated nuclei and mature cytoplasm, lacking conspicuous morphological abnormalities
(2) Heterozygous JAK2V617F mutation, and low JAK2 allele mutation load	Normocellular bone marrow (< 60%) and no proliferation or immaturity of granulopoiesis or erythropoiesis
(3) Normal erythrocytes < 5.8 × 1012/L males, < 5.6 × 1012/L females	RF 0 or 1
(4) Hb and ht normal or in the upper range of normal
Prodromal PV	ET with bone marrow features of PV
(1) Platelet count of > 350 × 109/L Hb and Ht normak or in the upper range of normal, normal erythrocyte < 5.8 × 1012/L males, < 5.6 × 1012/L females	Increased cellularity (60%-80%) due to increased erytropoiesis or trilineage myeloproliferation (i.e., panmyelosis). Proliferation and clustering of medium sized to large (pleomorphic) mature megakaryocytes
(2) Presence of JAK2V617F mutation and variable JAK mutation load	Absence bone marrow features consistent with congenital polycythemia and secondary erythrocytosis
(3) Low serum EPO level and increased LAP score	RF 0 or 1
(4) Spontaneous EEC
Prefibrotic hypercellular ET	EMGM
(1) Platelet count of > 350 × 109/L	Hypercellular ET due to chronic megakaryocytic and EMGM and normal or reduced erythroid precursors
(2) Presence of JAK2V617F mutation and high JAK2 mutation load	Loose to dense clustering of more pleiomorphic megakaryocytes with hyperploid or clumpsy nuclei (not or some cloud-like)
(3) Slight or moderate splenomegaly on ultrasound or on palpation
(4) No preceding or allied CML, PV, PMGM, RARS-T or MDS
Clinical stage 1: No anemia with Hb and Ht in the normal or low normal range: hb > 12 g/dL, normal LDH and CD34+	Grading of reticulin fibrosis and MF in EMGM
Clinical stage 2: Slight anemia Hb < 12 to > 10 g/dL, LDH↑, and splenomegaly	Prefibrotic: RF 0/1 = MF 0, no/minor splenomegaly
Clinical stage 3: Anemia, Hb < 10 g/dL, LDH↑↑, CD34+, leukoerythroblastose and, tear drop	Early fibrotic EMGM: RF 2 = MF 1 and minor or moderate splenomegaly
	Fibrotic EMGM: RF 3, RCF = MF 2 and overt splenomegaly
	Post-ET MF: RF 3/4 = MF 2/3 (WHO criteria)

ET: Essential thrombocythemia; MF: Myelofibrosis; EPO: Erythropoetin; CM: Clinical and molecular; Hb: Hemoglobin; ht: Hematocrit; WHO: World Health Organization; PV: Polycythemia vera; EPO: Erythropoetin; EEC: Endogenous erythroid colony; CML: Chronic myeloid leukemia; PMGM: Primary megakaryocytic granulocytic myeloproliferation; MDS: Myelodysplastic syndrome; EMGM: ET with hypercellular megakaryocytic-granulocytic myeloproliferation; RF: Reticuline fibrosis; LDH: Lactodehydrogenase; RCF: Reticulin collagen fibrosis.

Table 9 2015 WHO Clinical Molecular and Pathological criteria for the diagnosis of normocelular essential thrombocythemia carrying one of the MPL⁵¹⁵ mutations[78]

CM JAK2 wild type ET	Bone marrow pathology (P) criteria (WHO)
(1) Platelet count > 350 × 109/L and presence of large platelets in blood smear	P1 Proliferation of large to giant mature megakaryocyte with hyperlobulated, staghorn-like nuclei in a normocellular bone marrow (< 65%)
(2) Hemoglobin, haematocrit and erythrocyte count in the normal range	No increase of erythropoiesis, and no increase or immaturity of granulopoiesis or erythropoiesis, no or slight increase in reticulin RF 0/1
(3) Presence of MPL515 mutation and JAK2 wild type	ET → MF
(4) Normal serum EPO	Increased reticulin fibrosis around dense clustered megakaryocytes in a normocellular bone marrow and reduced erythropoiesis. Follow-up data of RF and MF related to splenomegaly in MPL515 ET transltional states to MF are lacking. Grading of RF and MF similar as described for PV
(5) Normal LAP score and CD11b expression
(6) No or slight splenomegaly
(7) No leukoerythroblastosis
(8) No preceding or allied CML, PV, RAS-T or MDS

This entity is identical to “true” ET as defined in 2002 by Michiels and Thiele in Table 5[91]. ET: Essential thrombocythemia; CM: Clinical and molecular; RF: Reticulin fibrosis; MF: Myelofibrosis; EPO: Erythropoetin; LAP: Leukocyte alkaline phosphatase; CML: Chronic myeloid leukemia; PV: Polycythemia vera; MDS: Myelodysplatic syndrome; WHO: World Health Organization.

Table 10 World Health Organization-clinical molecular and pathological criteria for hypercellular essential thrombocythemia associated with primary megakaryocytic, granulocytic myeloproliferation caused by calreticulin mutations[78]

Clinical CM criteria JAK2 wild type PMGM	Pathological ECP criteria of CALR MGM
A1 No preceding or allied other subtype of myeloproliferative neoplasm PV, CML, MDS. The main presenting features is pronounced isolated thrombocythemia with platelet count around or above 1000 × 109/L	P1 PMGM and relative or absolute reduction of erythropoiesis and erythroid precursors. Abnormal dense clustering and increase in atypical medium sized, large to giant immature megakaryocytes containing bulbous (cloud-like) hypolobulated nuclei and definitive maturation defects
A2 Presence of CALR mutation and JAK2 wild type
C Clinical stages of CALR MGM	MF Grading RF, MF
C1 Early clinical stage: Hb > 12 g/dL, slight to moderate splenomegaly, thrombocytosis around or above 1000 × 109/L, normal LAP score	MF 0 Prefibrotic CALR MGM, no reticulin fibrosis RF 0/1
C2 Intermediate clinical stage: slight anemia Hb < 12 to > 10 g/dL, decreasing platelet count, splenomegaly, increased LDH and definitive tear drop erythrocytes	MF 1 Early fibrotic CALR MGM slight reticulin fibrosis RF 2
C3 Advanced stage: anemia Hb < 10 g/dL, tear drop erythrocytes, increased LDH, increased CD34+ cells, pronounced splenomegaly, normal or decreased platelet counts, leucocytosis or leukopenia	MF 2 Fibrotic CALR MGM increase RF grade 3 and slight to moderate collagen fibrosis
	MF 3 Advanced fibrotic CALR MGM with collagen fibrosis-osteosclerosis

The combination of A1 + A2 and P1 establishes calreticulin (CALR) ET and various clinical stages(C1, C2, C3) related to the degree of myelofibosis (MF). This entity has been defined as CMGM in the 1990 Bone Marrow Classification by Georgii et al[35] as the third MPD entity and as chronic idiopathic myelofibrosis false ET or prefibrotic or primary megakaryocytic granulocytic myeloproliferation in 2002 by Michiels and Thiele (Table 5)[91]. PMGM: Primary megakaryocytic, granulocytic myeloproliferation; RF: Reticulin fibrosis; CM: Clinical and molecular; PV: Polycythemia vera; MDS: Myelodysplatic syndrome; LAP: Leukocyte alkaline phosphatase; CML: Chronic myeloid leukemia; Hb: Hemoglobin; LDH: Lactodehydrogenase; MGM: Megakaryocytic granulocytic myeloproliferation.

Table 11 Staging of JAK2^V617F positive prodromal polycythemia vera, erythrocythemic polycythemia vera, classical polycythemia vera, early myelofibrosis, inapparent polycythemia vera, spent phase polycythemia vera and post-polycythemia vera myelofibrosis according to 2015 World Health Organization-Clinical Molecular and Pathological criteria related to therapy

PV: WHO-ECMP stage	0	1	2	3	4	5	6
WHO-ECMP	Prodromal	Erythrocy-themic PV	Early PV	Manifest PV	PV early MF	Inapparent	Spent PV
Clinical diagnosis	PV			Classical PV	Masked PV	PV	Post-PV MF
LAP-score	↑	↑	↑	↑	↑/↑↑	↑	Variable
EEC	+	+	+	+	+	+	+
Serum EPO	N/↓	N/↓	↓	↓	↓	↓	Variable
Erythrocytes × 1012/L	> 5.8	< 5.8	> 5.8	> 5.8	> 5.8	Normal < 5.5	Decreased
Leukocytes × 109/L	< 12	< 12	< or > 12	< or > 15	> 15	N or ↑	> 20
Platelets × 109/L	> 400	400	< or > 400	> 400	< or >1000	N low or ↑	Variable
WHO-ECMP bone marrow	Early PV	Early PV	Early PV	Trilinear PV	Trilinear PV	Prilinear PV	Myelofibrosis
Bone marrow cellularity (%)	50-80	50-80	60-100	80-100	80-100	60-100	Decreased
Grading reticulin fibrosis: RF	RF 0-1	RF 0-1	RF 0-1	RF 0/1	RCF1/2/3	RCF 1/2/3	RCF 3/4
Grading myelofibrosis: MF57	MF 0	MF 0	MF 0	MF 0	MF 0/1	MF 0/2	MF 2/3
Splenomegaly on palpation	No/+	No	No/+	+	++/+++	++/+++	/large
Spleen size, echogram (cm)	< 12-15	< 13	12-15	12-16	18 > 20	16 > 20	> 20
Spleen size on palpation (cm)	0-3	NP	0-3	4-6	> 6	> 6	> 8
JAK2V617F in granulocytes %	Low	Low	Moderate < 50	High > 50	High > 50	Mod/High	High > 50
JAK2V617F in BFU-e (exon 12)	+(++)	+(++)	+(++)	++	++	+	++
Risk stratification → therapeutic implications anno 2014	Low risk	Low risk	Low risk	Intermediate risk PV	High risk	Wait/see	Post-PV MF
					PV early MF	IFN	Spent phase PV
						JAK2
First line Aspirin/Phlebotomy	Aspirin	Aspirin	Phlebotomy	Phlebotomy	If IFN resistant →	If IFN	JAK2
Second line IFN vs HU	Phlebotomy	Phlebotomy	Aspirin	Aspirin	HU or JAK2	Resistent	Inhibitor →
Third line JAK2 inhibitor			Low dose IFN → responsive	IFN → resistant → HU	inhibitor	JAK2 inhibitor	Bone marrow transplant

↑: Increased; ↓: Decreased; N: Normal; +: Present or heterozygous; ++: Homozygous; MF: Myelofibrosis; EPO: Erythropoetin; PV: Polycythemia vera; WHO-ECMP: World Health Organization European Clinical Molecular and Pathological; RF: Reticulin fibrosis; LAP: Leukocyte alkaline phosphatase; EEC: Endogenous erythroid colony; BFU-e: Burst forming units erythropoiesis; RCF: Reticulin collagen fibrosis; IFN: Interferon; HU: Hydroxy urea.

Table 12 2015 update on the molecular landscape findings in the chronic phase of essential thrombocythemia, polycythemia vera and myelofibrosis and during blast phase of myeloproliferative neoplasms transformation[121]

Gene	Chronic phase ET, PV and MF	Blast phase/AML
JAK2V617F	PV: 95%-98%; ET and MF: 50%-60%
MPL	ET: 1.5%; MF: 5%-10%
TET2	PV: 7%-16%; ET: 4%-11%; MF: 8%-17%
ASXL	PV: 2%; ET: 5%-8%; MF: 7%-17%	19%
DNMT3A	PV: 7%; ET: 3%; MF: 7%-15%	17%
CBL	MF: 6%
LNK	PV, ET, MF: < 5%	10%
IDH 1/2	MF: 4%	21%
IKZF		19%
EZH2	MPNs: 5%-13%
P53		27%
SRSF2		19%

MF: Myelofibrosis; PV: Polycythemia vera; ET: Essential thrombocythemia; AML: Acute myeloid leukemia.