Myeloproliferative Neoplasms - MPNs | Choose the Right Test

Medical Experts

Contributor

Li

Peng Li, MD, PhD

Associate Professor of Pathology (Clinical), University of Utah

Medical Director, Hematopathology, ARUP Laboratories

Contributor

Abujasen

Kaileigh Bingham Abujasen, MD

Resident, Pathology, University of Utah

Myeloproliferative neoplasms (MPNs) are a group of blood cancers that cause excess production of erythrocytes, platelets, and/or white blood cells in the bone marrow. MPNs include chronic myeloid leukemia (CML, discussed separately), polycythemia vera (PV), essential thrombocythemia (ET), primary myelofibrosis (PMF), chronic neutrophilic leukemia (CNL), chronic eosinophilic leukemia (CEL), and myeloproliferative neoplasms not otherwise specified or unclassified (MPN-NOS/U). The World Health Organization (WHO) also classifies juvenile myelomonocytic leukemia (JMML) as an MPN. MPNs are characterized by mutations that lead to clonal blood cell expansion. For example, CML is defined by the presence of the BCR::ABL1 (BCR-ABL1) fusion gene, also referred to as the Philadelphia chromosome. The BCR::ABL1-negative MPNs are driven by mutations involving JAK2, CALR, MPL, TET2, ASXL1, DNMT3A, and other genes.

Quick Answers for Clinicians

How do myeloproliferative neoplasms differ from myelodysplastic syndromes/myeloproliferative neoplasms?

Myeloproliferative neoplasms (MPNs) are distinct from myelodysplastic syndromes (MDSs) as they have no significant unilineage or multilineage dysplasia at presentation. If a disorder has both clonal myeloid proliferation and significant dysplasia (>10%), it is more appropriately characterized as a myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN). MDS/MPN disorders include chronic myelomonocytic leukemia (CMML), including its precursor conditions, clonal cytopenia with monocytosis of undetermined significance (CCMUS) and clonal monocytosis of undetermined significance (CMUS). Additional subtypes include MDS/MPN with neutrophilia (MDS/MPN-N, previously known as atypical chronic myeloid leukemia [aCML]), MDS/MPN with SF3B1 mutation and thrombocytosis, and MDS/MPN not otherwise specified (MDS/MPN-NOS).^,

What is the role of next generation sequencing in the diagnosis of myeloproliferative neoplasms?

Next generation sequencing (NGS) panels that include JAK2, MPL, and CALR genes may be used in place of individual molecular tests in the initial diagnostic workup of myeloproliferative neoplasms (MPNs). NGS may be used to establish clonality in triple-negative MPNs (those without JAK2, MPL, and CALR gene mutations) by demonstrating the presence of mutations in genes like TET2, SF3B1, ASXL1, and others that are associated with MPNs. After a diagnosis of MPN is established, NGS testing is recommended to evaluate for additional mutations associated with disease progression to determine mutational prognosis and better estimate outcomes such as leukemic transformation, overall survival, and disease-free survival.

Is further testing recommended after a diagnosis of myeloproliferative neoplasms is made?

Next generation sequencing (NGS) is recommended in those with myeloproliferative neoplasms (MPNs) to determine mutational prognosis. Given the increased risk of thrombosis and major bleeding events in those with MPNs compared to the general population, testing for coagulopathies is recommended in patients with an elevated platelet count, splenomegaly, unexplained bleeding, and/or those undergoing high-risk surgical procedures. A coagulopathy workup includes testing prothrombin time, partial thromboplastin time, fibrinogen, and/or testing specifically for acquired von Willebrand disease (VWD). If an allogeneic hematopoietic cell transplant is being considered, human leukocyte antigen (HLA) testing is necessary for donor matching.

Indications for Testing

The symptoms of MPNs vary widely depending on the specific subtype (ie, PV, ET, PMF, etc.). Often, patients will present with pruritus, fatigue, weight loss, and/or splenomegaly with associated symptoms. In early cases, patients may be asymptomatic at the time of diagnosis and are detected incidentally through a routine CBC test with an abnormal result such as leukocytosis, thrombocytosis, or erythrocytosis.^, A complete diagnostic workup for an MPN should be considered in the presence of unexplained thrombotic or hemorrhagic events.

If an MPN is suspected, further evaluation via additional laboratory testing, molecular testing, cytogenetic analysis, and bone marrow evaluation is indicated.

Diagnostic Criteria

The diagnostic and classification criteria of MPNs come from the WHO and the International Consensus Classification (ICC).^, The National Comprehensive Cancer Network (NCCN) does not advocate for one classification system over the other. WHO and ICC criteria include specific findings from a CBC, peripheral blood smear, and bone marrow analysis correlated with clinical history as well as the presence of certain molecular markers and the exclusion of other disorders.^,

Laboratory Testing

Diagnosis

Along with medical history, symptom assessment, and physical examination, the following laboratory tests are recommended as part of the initial evaluation in the diagnostic workup of MPNs: CBC, peripheral blood smear, comprehensive metabolic panel (CMP) (including uric acid, lactate dehydrogenase [LDH], and liver function tests [LFTs]), serum erythropoietin (EPO), and serum iron studies. Human leukocyte antigen (HLA) testing is indicated in patients who might undergo an allogeneic hematopoietic cell transplant.

If the results of these tests suggest the possibility of an MPN, bone marrow evaluation, cytogenetics, and molecular testing are indicated. A diagnosis is made when the results of these studies meet the specific diagnostic criteria for a particular MPN subtype outlined by the WHO or ICC.^, Distinguishing between MPN subtypes and grades is essential as treatments vary and long-term clinical outcomes differ significantly.

Testing for BCR::ABL1 Mutation

To exclude a diagnosis of CML, test peripheral blood by karyotyping, fluorescence in situ hybridization (FISH), or multiplex reverse transcriptase polymerase chain reaction (RT-PCR) for the BCR::ABL1 fusion gene. Karyotyping, FISH, and multiplex RT-PCR are recommended initial screening tests; the specific method used varies based on institutional preference. FISH may be better able to detect unusual breakpoints and cryptic translocations. The absence of BCR::ABL1 excludes CML, and testing for other MPN-associated mutations through next generation sequencing (NGS) should be performed. For testing specific to CML, refer to the ARUP Consult Chronic Myelogenous Leukemia - CML topic.

Bone Marrow Evaluation

Bone marrow evaluation is essential to assess the morphologic features of the various MPN subtypes, the blast percentage, and the degree of fibrosis. Evaluation includes a biopsy with reticulin and trichrome stains to grade the degree of fibrosis in PMF and evaluate for progression of ET or PV to myelofibrosis. Progression to myelofibrosis must be ruled out before initiating cytoreductive therapy. Another component of a bone marrow evaluation is an aspirate iron stain to assess for the presence of ringed sideroblasts and overall iron stores.

Cytogenetics

Cytogenetic analysis is important in diagnosing subtypes of MPNs. Cytogenetics may provide evidence of clonality at the time of diagnosis (supplementary to NGS). Repeat cytogenetic testing can demonstrate clonal evolution over time, help determine if the patient needs an allogeneic hematopoietic cell transplant, and/or be used to assess treatment. Chromosomal evaluation (ie, karyotyping) is performed on bone marrow aspirate samples when possible, with or without FISH; if a bone marrow aspirate is not available, peripheral blood may be used for karyotyping and/or FISH.

Although specific chromosomal abnormalities have not been identified for MPN subtypes other than CML, abnormal karyotypes may be present in varying degrees in non-CML MPNs. These abnormalities have prognostic significance and are used to determine risk stratification. They may also predict the transformation of an MPN to acute myeloid leukemia (AML).

Additional Molecular Testing

Molecular testing is used to assess clonality and detect MPN-specific mutations. Molecular testing of blood or bone marrow should be performed first for the JAK2 V617F mutation in all patients as it is the most common mutation, occurring in >90% of PV cases and approximately 60% of ET and PMF cases. If the JAK2 V617F mutation is not detected, testing for CALR and MPL mutations should follow in patients with ET or PMF, and testing for JAK2 exon 12 mutations should be performed in patients with PV. In those without JAK2, MPL, or CALR mutations (triple-negative MPNs), an NGS myeloid panel may be used to establish clonality. Initial testing using a multigene NGS panel that includes JAK2, CALR, and MPL may be used instead of single-gene tests. After the diagnosis of an MPN is established, if it was not already performed, NGS should be used to evaluate for additional mutations (eg, ASXL1, CBL, CSF3R, DNM3TA, EZH2, IDH1, IDH2, SF3B1, TET2, TP53, and U2AF1), which are used to determine prognosis.

Patients with molecular evidence of PDGFRA, PDGFRB, FGRF1, JAK2, FLT3, ETV6, and other tyrosine kinase gene rearrangements/fusions are considered under the category of myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions.^, Refer to the ARUP Consult Eosinophil-Related Disorders - Eosinophilia topic for more information.

Prognosis

The overall survival of patients with MPNs is variable depending on the patient’s age, symptoms, laboratory test results, MPN subtype, degree of fibrosis, blast percentage, molecular/cytogenetic profile, and other factors. Several scoring systems and prognostic models have been developed for the risk stratification of patients with MPNs. Refer to the NCCN for recommendations on risk stratification.

Monitoring

Regular monitoring for signs and symptoms of disease progression and therapeutic response is recommended, which may include repeat labs such as a CBC, molecular testing, and bone marrow evaluation. Medications commonly used in the treatment of MPNs, such as janus kinase (JAK) inhibitors (eg, ruxolitinib, fedratinib, and pacritinib), may have side effects. Refer to the NCCN for specific recommendations regarding monitoring for MPNs.