What are the basic investigations required for the diagnosis of pediatric Myeloproliferative Neoplasms (MPN)?

Diagnosis of Pediatric Myeloproliferative Neoplasms: Basic Investigations

The diagnostic workup for pediatric MPN requires a comprehensive laboratory and molecular evaluation that mirrors adult criteria, with the critical caveat that driver mutations (JAK2, CALR, MPL) are identified in only 22-26% of pediatric cases compared to the majority of adults, making diagnosis heavily reliant on clinical, laboratory, and histopathologic assessment with rigorous exclusion of secondary causes. 1, 2

Essential Initial Laboratory Studies

Complete Blood Count and Peripheral Blood Assessment

• CBC with differential to assess for erythrocytosis, thrombocytosis, leukocytosis, and presence of immature cells in circulation 3
• Peripheral blood smear examination to evaluate cell morphology, identify dysplastic features, circulating blasts, and leukoerythroblastosis 3, 4
• Document monocyte count, white blood cell count, and presence of circulating immature myeloid cells 4

Comprehensive Metabolic and Biochemical Panel

• Comprehensive metabolic panel including uric acid, lactate dehydrogenase (LDH), and liver function tests 3
• Serum erythropoietin (EPO) level - particularly important in suspected polycythemia vera, where levels are typically low or normal despite erythrocytosis 3
• Serum iron studies to exclude iron deficiency as a cause of reactive thrombocytosis 3

Critical Molecular and Cytogenetic Studies

Exclusion of BCR-ABL1 (Mandatory First Step)

• FISH or RT-PCR for BCR-ABL1 to definitively exclude chronic myeloid leukemia, which is the essential first step in diagnosing any MPN 3
• If BCR-ABL1 is positive, the patient has CML and requires different management entirely 3

Driver Mutation Testing (Sequential Approach)

• JAK2 V617F mutation testing as the initial molecular test - found in most adult PV/ET/PMF but only 22-26% of pediatric cases 3, 4, 1
• If JAK2 V617F is negative: test for CALR and MPL mutations (for ET and MF patients) 3, 4
• If JAK2 V617F is negative in suspected PV: test for JAK2 Exon 12 mutations 3
• Comprehensive myeloid mutation panel including SF3B1, TET2, ASXL1, SRSF2, and TP53 for diagnostic clarification and prognostic information 4

Bone Marrow Evaluation (Essential for Definitive Diagnosis)

• Bone marrow aspirate and biopsy with trichrome and reticulin staining to assess cellularity, fibrosis, and megakaryocyte morphology 3
• Bone marrow cytogenetics (karyotype ± FISH) to identify clonal abnormalities and provide prognostic information 3
• Flow cytometry immunophenotyping to detect abnormalities in erythroid, immature myeloid, granulocyte, and monocyte compartments 4

Additional Specialized Testing

Coagulation Assessment (Selected Patients)

• Coagulation tests to evaluate for acquired von Willebrand disease and other coagulopathies in patients with elevated platelet counts, splenomegaly, or unexplained bleeding 3
• Expanded panel including von Willebrand factor antigen, Factor VIII activity, and VWF multimers may be useful in high-risk surgical candidates 3

Clinical Assessment Components

• History and physical examination documenting spleen size by palpation, history of thrombotic/hemorrhagic events, and cardiovascular risk factors 3
• Family screening is recommended in JAK2 V617F-negative cases to differentiate from rare familial disorders caused by mutations of TPO or MPL (particularly MPLS505N) 3
• In JAK2 V617F-negative erythrocytosis with normal/reduced EPO, search for rare erythropoietin receptor mutations if family history is positive 3

Critical Pediatric-Specific Considerations

The absence of driver mutations in 74-78% of pediatric MPN cases means diagnosis must rely heavily on:

• Rigorous exclusion of secondary causes including infectious diseases, solid tumors, autoimmune conditions, prior chemotherapy/radiation, and environmental exposures 4, 1
• Serial clinical and laboratory assessments over time, as diagnosis may require longitudinal observation 5
• Application of WHO 2016 diagnostic criteria with recognition that pediatric cases may not meet all molecular criteria 3, 1, 2

Common diagnostic pitfall: Do not dismiss the diagnosis of MPN in children simply because driver mutations are absent - the majority of pediatric cases are mutation-negative, making clinical and histopathologic correlation paramount 1, 2

Diagnostic Algorithm Priority

1. First tier (immediate): CBC with differential, peripheral smear, BCR-ABL1 testing, comprehensive metabolic panel with LDH and uric acid 3, 4
2. Second tier (concurrent): JAK2 V617F testing, serum EPO level, iron studies 3, 4
3. Third tier (if JAK2 negative): CALR, MPL, JAK2 Exon 12 mutations, comprehensive myeloid panel 3, 4
4. Fourth tier (definitive): Bone marrow aspirate/biopsy with cytogenetics and flow cytometry 3, 4

Referral to specialized centers with expertise in pediatric MPN management is strongly recommended given the rarity and diagnostic complexity of these disorders in children. 3, 1, 2