Diagnosis
Common tests and WHO criteria for the diagnosis of myelofibrosis
Myelofibrosis (MF) is a heterogeneous clonal myeloid stem cell cancer and belongs to a group of conditions called myeloproliferative neoplasms (MPN)1. MF primarily affects older adults, over the age of 601. The annual incidence of MF in the UK is approximately 0.6 per 100,000 persons2, with a 10-year prevalance of 3.2 per 100,000 persons2.
Overview of Myelofibrosis
According to the REALISM study, of 200 patients with MF surveyed in the UK, 58.9% present with symptoms (n = 117) and 47% present with splenomegaly (n = 94) at diagnosis10. However, up to 30% of patients can be asymptomatic and are diagnosed through routine examination or discovered through abnormal laboratory tests3.
MF is diagnosed using a combination of physical examination, blood tests, imaging, bone marrow biopsy and molecular testing4,5,7.
Blood Tests4
Complete Blood Counts (CBC) with differential
Peripheral blood smear
Blood chemistry profile
CBC with differential to assess White Blood Cell (WBC), Platelets (PLT) counts and haemoglobin levels.
Peripheral blood smear to assess morphology of circulating cells. Myelofibrosis (MF) can be characterised by leucoerythroblastic with left-shifted granulocytes and an abnormal presence of blast cells in circulation.
Blood chemistry to assess Lactate Dehydrogenase (LDH) levels.
Renal and liver function, iron status and serum uric acid are usually also analysed to assess hepatic and renal conditions.
Bone Marrow Examination4,7
Aspiration and biopsy
To assess morphological characteristics such as:
Proliferation of atypical megakaryocytes
Clustering and abnormal localisation of megakaryocytes with hyperchromatic or bulbous nuclei
Increased reticulin network with focal or diffuse collagen
Extensive osteosclerosis in advanced stages
Reticulin grading is essential to establish MF diagnosis, with a minimum of grade 2 (0 – 3 grading system)
Imaging5,7
Ultrasound
X-rays
MRI
Imaging tests can be used to assess spleen size.
Clinical palpation or ultrasound imaging of the left costal margin are the easiest ways to assess spleen size, which is recoded in centimetres.
In MF trials the preferred method to assess splenomegaly according to the International Working Group-MPN Research and Treatment (IWG-MRT) is spleen volume in cm3, which is usually measured by tomography or magnetic resonance.
Spleen volume determination is not a requirement in routine clinical practice.
Molecular Testing for Genetic Mutations and Chromosome Abnormalities4,7
DNA Sequencing
Cytogenetic analysis (Karyotyping)
Patients with suspected MF usually undergo molecular testing for the most prevalent mutations:
JAK2
CALR
MPL
If a patient is triple negative for the classic MF mutations, a myeloid gene panel containing other known High Molecular Risk (HMR) mutations can be used as a molecular test.
A Cytogenetic analysis and/or Single Nucleotide Polymorphism (SNP) arrays can also be performed to aid the diagnosis of MF.
Diagnosis of Myelofibrosis is based on the 2022 WHO criteria6,7.
All 3 major criteria and at least 1 minor criterion are in two consecutive determinations required for diagnosis:
Post-PV, post-ET MF
- Documentation of a previous established diagnosis of PV or ET
- Bone marrow fibrosis of Grade 2 - 3 on a scale of 0 - 3
Primary MF
- Bone marrow biopsy showing megakaryocytic proliferation and atypia, accompanied by reticulin and/or collagen fibrosis grades 2 or 3a
- JAK2, CALR or MPL mutation assessed by sensitive technique or presence of another clonal markerb or absence of minor reactive bone marrow reticulin fibrosisc
- Not meeting the WHO criteria for ET, PV, BCR-ABL1: fusion gene between BCR and ABL1 genes found in certain forms of Chronic Myeloid Leukaemia (CML), myelodysplastic syndromes or other myeloid neoplasms
Post-PV, post-ET MF
- Anaemia and a > 20 g/L decrease from baseline haemoglobin concentration. A sustained loss of requirement of either phlebotomy (in the absence of cytoreductive therapy) or cytoreductive treatment for erythrocytosis
- Development of any two (or all three) of the following constitutional symptoms: >10% weight loss in 6 months, night sweats and unexplained fever (> 37.5oC)
- Increased palpable splenomegaly > 5 cm from the baseline or newly palpable
- Elevated Lactate Dehydrogenase (LDH, for post-ET MF only)
- Leucoerythroblastosis
Primary MF
- Anaemia not attributed to a comorbid condition
- Leucocytosis ≥ 11 x 109/L
- Splenomegaly detected clinically and/or by imaging
- Elevated LDH
- Leucoerythroblastosis
a. in grades MF-2 or MF-3 an additional trichrome stain is recommeded;
b. in the absence of any of the 3 major clonal mutations, the search for the most frequent accompanying mutations (e.g. ASXL1, EXH2, TET2, IDH1/IDH2, SRSF2, SF3B1) are of help in determining the clonal nature of the disease;
c. Bone Marrow (BM) fibrosis secondary to infection, autoimmune disorder or other chronic inflammatory conditions, hair cell leukaemia, or other lymphoid neoplasm
Prognostic scoring systems7
There are multiple prognostic systems to assess risk of MF patients: International Prognostic Scoring System (IPSS), used to assess risk at diagnosis, Dynamic International Prognostic Scoring System (DIPSS or DIPSS plus), used to assess risk at any stage of disease progression, Mutation and Karyotype-enhanced International Prognostic Scoring System for Primary Myelofibrosis (MIPSS70 v 2.0) and a prognostic system used to predict survival in patients with post-polycythaemia vera and post-essential thrombocythaemia myelofibrosis (MYSEC-PM).
| IPSS | DIPSS | DIPSS plus | MYSEC-PM | MIPSS 70 v 2.0 | |||||
|---|---|---|---|---|---|---|---|---|---|
| Newly diagnosed PMF patients | PMF patients at any stage of disease | PMF patients at any stage of disease | Post-PV and Post-ET patients | PMF patients up to 70 years old | |||||
| Variables | Points | Variables | Points | Variables | Points | Variables | Points | Variables | Points |
| Age > 65 y | 1 | Age > 65 y | 1 | Age > 65 y | 1 | Age | 0.15 per year of age | VHR Karyotype | 4 |
| Hb < 100 g/L | 1 | Hb < 100 g/L | 2 | Hb < 100 g/L | 2 | Hb < 110 g/L | 2 | Unfavourable karyotype | 3 |
| WCC > 25 x 109/L | 1 | WCC > 25 x 109/L | 1 | WCC > 25 x 109/L | 1 | Plts < 150 x 109/L | 1 | ≥ 2 HMR mutations | 3 |
| Circulating blasts ≥ 1% | 1 | Circulating blasts ≥ 1% | 1 | Circulating blasts ≥ 1% | 1 | Circulating blasts ≥ 3% | 2 | 1 HMR mutation | 2 |
| Constitutional Symptoms | 1 | Constitutional Symptoms |
1 | Constitutional Symptoms |
1 | CALR mutation absent | 2 | Type 1/like CALR absent | 2 |
| Unfavourable Karyotype |
1 | Constitutional Symptoms | 1 | Hb < 80 g/L (f) Hb < 90 g/L (m) |
2 | ||||
| Plts < 100 x 109/L | 1 | Hb 80-99 g/L (f) Hb 90-109 g/L (m) |
1 | ||||||
| Red blood cell transfusion | 1 | Circulating blasts ≥ 2% | 1 | ||||||
| Constitutional Symptoms | 2 | ||||||||
The sum of all the points of each scoring system is associated to overall survival and can be used to calculate relative risk as per table below.
| Risk Groups | IPSS | DIPSS | DIPSS plus | MYSEC-PM | Risk Groups | MIPSS 70 v 2.0 | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Score | Survival (years) |
Score | Survival (years) |
Score | Survival (years) |
Score | Survival (years) |
Score | Survival (years) |
||
| Low | 0 | 11.3 | 0 | Not reached | 0 | 15.4 | < 11 | Not reached | Very Low | 0 | Not reached |
| Intermediate-1 | 1 | 7.9 | 1-2 | 14.2 | 1-2 | 6.5 | 11-13 | 9.3 | Low | 1-2 | 16.4 |
| Intermediate-2 | 2 | 4 | 3-4 | 4 | 3-4 | 2.9 | 14-16 | 4.4 | Intermediate | 3-4 | 7.7 |
| High | ≥ 3 | 2.3 | > 4 | 1.5 | ≥ 4 | 1.3 | > 16 | 2.0 | High | 5-8 | 4.1 |
| Very High | ≥ 9 | 1.8 | |||||||||
PMF: primary myelofibrosis, PV: polycythaemia vera, ET: essential thrombocythaemia, Hb: haemoglobin, WCC: white cell count, Plts: platelets, CALR: calreticutlin, VHR: very high risk, HMR: high molecular risk
Splenomegaly
- Patients with MF may experience splenomegaly, or an enlarged spleen, due to splenic extramedullary haemtopoiesis (EMH)8
- Progressive splenomegaly is significantly associated with symptoms including early satiety, portal hypertension, decreased physical activity, deteriorative abdominal pain8
- Splenomegaly is associated with the progression of cytopenias due to splenic sequestration of circulatory cells and platelets
47% of MF patients present with an enlarged spleen at diagnosis10
Based on a cohort of 200 adult patients with MF enrolled in the REALISM UK study between 2018 and 2019, which was a multicentre, retrospective, non-interventional study from 15 NHS hospitals across the United Kingdom10
Constitutional Symptoms
- The abnormal production of cytokines is thought to result in constitutional symptoms e.g. night sweats, itchy skin, weight loss in MF9
- Constitutional symptoms are reported in more than 80% of MF patients throughout disease progression and can compromise the quality of life in patients9
- 23% of patients with MF present with at least 1 constitutional symptom at diagnosis (based on a cohort of 200 MF patients from the UK REALISM study)10
- The presence of constitutional symptoms is an adverse prognostic factor when estimating the survival of patients with MF9,11
Constitutional symptoms associated with MF may include night sweats, pruritus, undesired weight loss and fever.9
Most frequently reported symptoms by MF patients*
*as reported by MF patients (n = 174) in the international myeloproliferative neoplasms (MPN) Landmark survey (Harrison, et al, Ann Hematol, 2017)
Progressive bone marrow fibrosis can lead to worsening cytopenias, including anaemia, thrombocytopenia and neutropenia9,13
Anaemia
- Anaemia in MF may result from disruption in normal erythropoiesis due to a proinflammatory environment, fibrotic bone marrow, increased red blood cell (RBC) turnover and splenic sequestration of RBC14
- As MF progresses, anaemia can become more prominent, either through a direct consequence of disease progression or due to therapies used to treat the disease. RBC transfusions are an option in managing the anaemia aspect of MF14
- Anaemia and transfusion dependence are adverse prognostic factors and are associated with increased complications and poor overall survival14
- No anaemia (Hb ≥ 13.5 g/dL for men and ≥ 12.0 g/dL for women), median survival: 7.9 years15
- Mild anaemia (Hb ≥ 10 g/dL but below sex adjusted lower limit of normal), median survival: 4.9 years15
- Moderate anaemia (Hb 8 - <10 g/dL), median survival: 3.4 years15
- Severe anaemia (Hb < 8 g/dL or transfusion dependent), median survival: 2.1 years15
Data from 1109 consecutive PMF patients from the Mayo Clinic cohort with a median follow up (for living patients at the time of writing this publication) of 6.1 years15
Thrombocytopenia
- Thrombocytopenia in MF can result from multiple causes, including ineffective haematopoiesis, splenic sequestration of platelets, and treatment-related effects18
- Thrombocytopenia occurs when there is a deficiency of platelets in the blood, in adults defined as <150 x 109 platelets/L19
- Between 16% and 26% of MF patients present with moderate thrombocytopenia at diagnosis, defined as platelet count of 50 - 100x109/L18
- Between 11% and 16% of MF patients present with severe thrombocytopenia at diagnosis, defined as platelet count less than 50x109/L18
- Thrombocytopenia is an adverse prognostic factor and closely linked to disease progression20
Neutropenia
- Neutropenia, or a low white blood cell count, weakens the immune system, making it harder for the body to fight infection21
- Numerous factors may cause neutropenia through destruction, decreased production, or abnormal storage of neutrophils22
OVERACTIVE JAK-STAT SIGNALLING PATHWAY IS A KEY DRIVER OF MF23,24
- Mutations leading to constitutional activation of the JAK/STAT signalling pathway are found in 90% of PMF patients. Mutually exclusive JAK2 V617F, CALR and MPL mutations can be found in 60%, 20-30% and 5-10% of PMF patients respectively.25
- Constitutional symptoms driven by increased levels of circulating inflammatory cytokines are increasingly recognised as being attributed to overactive JAK1 signalling23
- Overactive JAK2 signalling has been implicated in splenomegaly in patients with myelofibrosis (MF)24
Pathways beyond JAK-STAT are being explored to further understand the drivers that may contribute to key manifestations of MF
Other pathways with evidence of contribution to pathogenesis of MF include:
ACVR1/ALK2
The activin A receptor type 1 (ACVR1) gene, which encodes for the activin receptor-like kinase-2 (ALK2), is involved in bone morphogenetic protein (BMP) signalling. ALK2 activation leads to increased hepcidin production, which in turn promotes anaemia.26
BCL-2/BCL-XL
Myeloproliferative neoplasm (MPN) cells with Janus kinase 2 (JAK2) mutations are believed to be dependent on B-cell lymphoma 2 (Bcl-2) and B-cell lymphoma-extra large (Bcl-xL) proteins for survival. Over expression of these proteins can help mutated cells avoid apoptosis and continue proliferating.27,28
BET
Bromodomain and extra-terminal domain (BET) proteins are epigenetic readers that are believed to contribute to the progression of MF due to their involvement in several signalling pathways. Activation of BET has been shown to increase nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) signalling, thereby increasing inflammation.29
PI3K/AKT
Aberrant activation of the phosphatidylinositol 3-kinase (PI3K) and protein kinase B (Akt) pathway is present in cancer and other diseases involving immune deficiencies and tissue overgrowth. Akt activation leads to proliferation of normal or malignant cells while the activation of the PI3K pathway can promote the production of proinflammatory cytokines.30,31
TELOMERASE
Telomerase is an enzyme that maintains telomere length in rapidly dividing cells. Upregulated telomerase activity has been observed in myeloproliferative (MPN) cells, is thought to enable telomere length maintenance and cell survival in rapidly dividing cells.32
TGF-β
Transforming growth factor beta (TGF-β) expression is elevated in primary myelofibrosis (PMF) patients compared to people without PMF. In addition to inducing fibrosis, TGF-β has been shown to inhibit haematopoiesis and negatively affect erythroid differentiation.33
References
- Harrison C, McLornan D. Myelofibrosis. Hematology. 2014;19(2):120-1.
- Haematological Malignancy Research Network Factsheets. Last accessed July 2025.
- Cervantes F;Blood;2014;124;2635-2642
- Leukemia & Lymphoma Society. Myelofibrosis: diagnosis. Last accessed July 2025.
- Mayo Clinic. Myelofibrosis Diagnosis: Last accessed July 2025.
- Arber DA, et al. Blood. 2016;127(20):2391-2405
- McLornan D.P. et al, BJHaematol, 2023; 00; 1-9
- Song MK et al. Int J Mol Sci. 2018;19(3):898.
- Mughal TI et al. Int J Gen Med. 2014;7:89-101
- Mead AJ et al. Ther Adv Hematol. 2022;13:1-15.
- Mesa RA et al. Leuk Res. 2009;33(9):1199-1203.
- Harrison CN;Ann Hematol;2017;96;1653-1665
- Bose P, Verstovsek S. Curr Hematol Malig Rep. 2018;13(3):164-172.
- Naymagon L;Hemasphere;2017;1;1-9
- Nicolosi M, et al. Leukemia. 2018;32;1254-1258
- Kiladjian JJ;Hemasphere;2023;7;1-15
- Passamonti F, Clin Rev Onc/Hem; 2022; 180:1040-8428
- Sastow D et al. Clin Lymphoma Myeloma Leuk. 2022;22(7):e507-e520.
- Mayo Clinic;2023;1-3;Thrombocytopenia
- Scotch AH et al. Leuk Res. 2017;63:34-40.
- Bloodcancer;2023;1-4;Infection risk and neutropenia
- Mayo;2022;1-5;Neutropenia
- Mascarenhas J. Leuk Lymphoma. 2015;56(9):2493-2497.
- Tremblay D;Ann Hematol;2020;99;1441-1451
- Mascarenhas J;Leukemia;2023;37;255–264
- Aykul S et al. J Clin Invest. 2022;132(12):e153792.
- Tognon R et al. J Hematol Oncol. 2012;5:2.
- Petiti J et al. J Cell Mol Med. 2020;24(18):10978-10986.
- Kleppe M et al. Cancer Cell. 2018;33(1):29-43.e7.
- Bartalucci N et al. J Cell Mol Med. 2013;17(11):1385-1396.
- Fruman DA et al. Cell. 2017;170(4):605-635.
- Tefferi A et al. N Engl J Med. 2015;373(10):908-919.
- Ceglia I et al. Exp Hematol. 2016;44(12):1138-1155.e4.
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