Diagnostics

BCR::ABL1-negative myeloproliferative neoplasms (MPN) - overview

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Classification
Diagnostics
Prognosis

Myeloproliferative neoplasms (MPN) are rare, clonal diseases of the hematopoietic stem cell, which have many similarities. Especially in the early stages, they are often difficult to distinguish from each other and can also merge in individual cases. According to the WHO classification 2022, polycythemia vera (PV) shows an overall incidence of 1.57/100,000 person-years, essential thrombocythemia (ET) an overall incidence of 1.55/100,000 person-years, and primary myelofibrosis (PMF) in the fibrotic stage an annual incidence of 0.44-1.5 cases per 100,000 person-years (WHO 2022). MPNs usually affect people of older age with a median of 60-65 years (Barbui 2012). Characteristic of MPN is hypercellularity of the bone marrow, specifically the myeloid cell series, and increased numbers of erythrocytes, granulocytes, and/or platelets in the peripheral blood, depending on the entity (Haferlach et al. 2008, WHO 2022).

Myeloproliferative neoplasms: Classification

Classification into BCR::ABL1-positive CML and BCR::ABL1-negative myeloproliferative neoplasms is based on the WHO classification (Table 1):

Table 1: MPN WHO classification 2022 (WHO 2022)

Myeloproliferative neoplasms (MPN)
Chronic myeloid leukemia (CML)	BCR::ABL1 positive
Polycythemia vera (PV)	BCR::ABL1 negative
Essential thrombocythemia (ET)
Primary myelofibrosis (PMF)
Chronic neutrophilic leukemia (CNL)
Chronic eosinophilic leukemia (CEL)
Juvenile myelomonocytic leukemia (JMML)
Myeloproliferative neoplasms, not otherwise specified (NOS)

Myeloproliferative neoplasms: Diagnostic methods and their relevance

Cytomorphologic assessment in MPNs involves cellularity in the whole as well as in individual hematopoietic series. It is also important to determine the proportion of blasts in the blood and bone marrow. If there is fibrosis of the bone marrow, e.g. in primary myelofibrosis (PMF), the cytomorphological assessability of the preparations is often limited (punctio sicca).

In all cases, the histology is decisive for the assessment of the degree of fibers and the bone marrow architecture, which should always be performed in the case of a suspected or confirmed MPN.

The chromosomal alterations occurring in MPN present very heterogeneously. Chromosomal aberrations are most frequently observed in PMF (40%), followed by PV (35%), whereas alterations in ET are rarely found (~ 3%). Over 80% of chromosomal aberrations in MPN are unbalanced alterations (Bacher et al. 2005). An overview of typical chromosomal aberrations is given in Table 2.

Table 2: Overview of chromosomal aberrations in classical BCR::ABL1-negative MPN (Reilly 2008, Tang et al. 2017, Gangat et al. 2022, WHO 2022, Tefferi 2023)

Aberration	PMF	PV	ET
+1q	+	+
+8	+	+	+
+9	+	+	+
-5	+
del(5q)	+	+	+
-7	+	+
del(7q)	+	+
del(11q)	+
del(12p)	+	+
del(13q)	+	+	+
del(20q)	+	+	+
i(17q)	+

In the context of MPN diagnostics, FISH analysis is predominantly to be seen as a complementary method to classical chromosome analysis. It is used specifically to answer certain questions. To exclude a BCR::ABL1 rearrangement as a reliable differentiation from CML, a FISH analysis and/or PCR should always be performed in cases of suspected MPN, since a cryptic BCR::ABL1 rearrangement may be present even in the absence of a Philadelphia translocation in the chromosome analysis (so-called Philadelphia-negative BCR::ABL1-positive CML).

By means of a FISH "screening" on interphase nuclei only a fraction of the aberrations possible in MPN would be detected. Therefore, FISH analysis cannot replace classical chromosome analysis.

Molecular genetics is the most important tool in MPN diagnostics today. It is used to exclude a BCR::ABL1 rearrangement and is also used to differentiate a MPN from a reactive alteration and to diagnose progression. For this purpose, a large number of mutations (especially JAK2 V617F, MPL, CALR) has been identified in the last ten years (see Table 3).

Table 3: Frequency of the different mutations in PV, ET and PMF (Luque Paz et al. 2023)

Gene mutation	PMF	PV	ET
JAK2	60	98	55
MPL	7-10	0	5-7
CALR	20-30	0	25-30
TET2	10-20	10-20	3-10
DNMT3A	8-12	5-10	1-5
IDH1/2	5-6	1-2	1-2
ASXL1	15-35	2-7	5-10
EZH2	7-10	1-2	1-2
NRAS	2-4	<2	<2
KRAS	2	<2	<2
SH2B3	2-4	2-9	1-3
CBL	4	<2	<2
SRSF2	6-14	<2	<2
SF3B1	5-7	2-3	2-5
U2AF1	7-10	<2	<2
NFE2	3-5	3-6	1-7
RUNX1	2-3	<2	<2
TP53	4-5	<2	<2

In familial MPN, mutations in the VHL, EPOR, EPAS1 and EGLN1 genes can occasionally be found. These mutations are very rare and should be investigated when all other markers are negative, the patient is very young, and there is a positive family history (Jones & Cross 2013, Rumi & Cazzola 2017, McMullin 2019).

Myeloproliferative neoplasms: Prognosis

In addition to increased age, leukocytosis, and thrombosis (Gangat et al. 2011, Barbui et al. 2018), in general, the detection of chromosomal alterations at diagnosis of an MPN seems to be associated with a less favorable prognosis. The presence of complex karyotypes during the course of an MPN increases the likelihood of transition to blast phase (Haferlach et al. 2008). SNP array analyses showed clustered deletions of the genes ETV6 (Chr. 12), TP53 (Chr. 17), or RUNX1 (Chr. 21) in blast phase (Thoennissen et al. 2010).

Gene mutations influence the risk profile of "classic" BCR::ABL1-negative MPNs

The JAK2 V617F mutation is the most common mutation in MPN. PV is almost always associated with this mutation (98%). In addition to the JAK2 V617F mutation, mutations in the CALR (20-30%) and MPL (5-10%) genes often occur in ET and PMF (Luque Paz et al. 2023). These mutations are among the driver mutations in MPN, but do not allow specific diagnosis due to their cross-entity occurrence.

81% of PMF patients, 53% of PV patients, and 53% of ET patients show mutations in addition to driver mutations (non-driver mutations, Fig. 1). IDH2 mutations are classified as a risk factor in all three classic BCR::ABL1-negative MPN (Tefferi et al. 2016, Tefferi & Vannucchi 2017). Furthermore, each of the three entities has an entity-specific risk profile (Fig. 1).

Figure 1: Recurrent non-driver gene mutations in PMF, PV and ET with prognostic impact (adapted from Tefferi et al. 2016, Tefferi & Vannucchi 2017).

For a detailed insight into the individual MPNs and their genetic risk profiles, please refer to the infotexts of the respective entity:

Bacher U et al. Conventional cytogenetics of myeloproliferative diseases other than CML contribute valid information. Ann Hematol 2005;84(4):250-257.

Barbui T. How to manage children and young adults with myeloproliferative neoplasms. Leukemia 2012;26(7):1452-1457.

Barbui T et al. Philadelphia chromosome-negative classical myeloproliferative neoplasms: revised management recommendations from European LeukemiaNet. Leukemia 2018;32(5):1057-1069.

Gangat N et al. DIPSS plus: a refined Dynamic International Prognostic Scoring System for primary myelofibrosis that incorporates prognostic information from karyotype, platelet count, and transfusion status. J Clin Oncol 2011;29(4):392-397.

Gangat N et al. Cytogenetic abnormalities in essential thrombocythemia: Clinical and molecular correlates and prognostic relevance in 809 informative cases. Blood Cancer J 2022;12(3):44.

Haferlach T et al. The diagnosis of BCR/ABL-negative chronic myeloproliferative diseases (CMPD): a comprehensive approach based on morphology, cytogenetics, and molecular markers. Ann Hematol 2008;87(1):1-10.

Jones AV, Cross NC. Inherited predisposition to myeloproliferative neoplasms. Ther Adv Hematol 2013;4(4):237-253.

Luque Paz D et al. Genetic basis and molecular profiling in myeloproliferative neoplasms. Blood 2023;141(16):1909-1921.

McMullin MF. Diagnostic workflow for hereditary erythrocytosis and thrombocytosis. Hematology Am Soc Hematol Educ Program 2019;2019(1):391-396.

Reilly JT. Pathogenetic insight and prognostic information from standard and molecular cytogenetic studies in the BCR-ABL-negative myeloproliferative neoplasms (MPNs). Leukemia 2008;22(10):1818-1827.

Rumi E, Cazzola M. Advances in understanding the pathogenesis of familial myeloproliferative neoplasms. Br J Haematol 2017;178(5):689-698.

Tang G et al. Characteristics and clinical significance of cytogenetic abnormalities in polycythemia vera. Haematologica 2017;102(9):1511-1518.

Tefferi A. Primary myelofibrosis: 2023 update on diagnosis, risk-stratification, and management. Am J Hematol 2023;98(5):801-821.

Tefferi A et al. Targeted deep sequencing in polycythemia vera and essential thrombocythemia. Blood Adv 2016;1(1):21-30.

Tefferi A, Vannucchi AM. Genetic Risk Assessment in Myeloproliferative Neoplasms. Mayo Clin Proc 2017;92(8):1283-1290.

Thoennissen NH et al. Prevalence and prognostic impact of allelic imbalances associated with leukemic transformation of Philadelphia chromosome-negative myeloproliferative neoplasms. Blood 2010;115(14):2882-2890.

WHO Classification of Tumours Editorial Board. Haematolymphoid tumours [Internet; beta version ahead of print]. Lyon (France): International Agency for Research on Cancer; 2022. (WHO classification of tumours series, 5th ed.; vol. 11). Available from: https://tumourclassification.iarc.who.int/chapters/63.

Status: Dezember 2023

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BCR::ABL1-negative myeloproliferative neoplasms (MPN) - overview

Myeloproliferative neoplasms: Classification

Table 1: MPN WHO classification 2022 (WHO 2022)

Myeloproliferative neoplasms: Diagnostic methods and their relevance

Cytomorphology

Chromosome analysis

Table 2: Overview of chromosomal aberrations in classical BCR::ABL1-negative MPN (Reilly 2008, Tang et al. 2017, Gangat et al. 2022, WHO 2022, Tefferi 2023)

Fluorescence in situ hybridisation (FISH)

Molecular genetics

Table 3: Frequency of the different mutations in PV, ET and PMF (Luque Paz et al. 2023)

Myeloproliferative neoplasms: Prognosis

Gene mutations influence the risk profile of "classic" BCR::ABL1-negative MPNs

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