Diagnosis of Acute Promyelocytic Leukemia (APL)
When APL is suspected based on morphology or clinical presentation (especially with coagulopathy), immediately initiate ATRA therapy before genetic confirmation and perform rapid FISH for PML-RARA on bone marrow samples. 1
Initial Clinical Recognition
Suspect APL in any patient presenting with:
- Characteristic morphology showing abnormal promyelocytes with heavy granulation (hypergranular variant) or bilobed nuclei with minimal granulation (microgranular variant) 1
- Severe coagulopathy or disseminated intravascular coagulation (DIC), particularly in the setting of acute leukemia 1
- Immunophenotype showing CD34-negative, HLA-DR-negative/dim blasts with strong CD13/CD33 positivity and abnormally low CD15 expression 1
Mandatory Diagnostic Workup
Immediate Sample Collection
Obtain bone marrow aspirate as the preferred specimen for all diagnostic studies; peripheral blood may be used only if bone marrow is unobtainable or contraindicated, or when blast count is sufficiently high 1. A trephine biopsy is required only if the aspirate is dry and no abnormal cells are present in peripheral blood 1.
Essential Morphologic and Cytochemical Studies
- Perform Romanowsky-derived stains (Wright-Giemsa or May-Grunwald-Giemsa) on bone marrow aspirate smears 1
- Conduct myeloperoxidase (MPO) or Sudan Black B staining for diagnosis and subclassification 1
- Evaluate peripheral blood smear and complete blood count with differential 1
Genetic Confirmation (Mandatory)
Rapid FISH for PML-RARA detection on bone marrow is the critical first-line genetic test when APL is suspected, as it provides results within 24-48 hours 1. This test must be performed immediately upon suspicion.
Molecular confirmation by RT-PCR for PML-RARA fusion transcripts is mandatory for definitive diagnosis and must be performed on bone marrow or peripheral blood samples 1. RT-PCR detects PML-RARA in 100% of cases with cytogenetically confirmed t(15;17) 2.
Conventional karyotyping should be performed to identify t(15;17)(q22;q21) translocation, though this is time-consuming and may miss cryptic rearrangements 1. Karyotyping is also useful for identifying rare APL variants with alternative RARA fusions such as PLZF-RARA, NuMA-RARA, or NPM1-RARA 1.
Discontinue ATRA if t(15;17) translocation or PML-RARA fusion is not confirmed 3. Patients without these genetic markers should not receive APL-directed therapy 1, 3.
Immunophenotyping by Flow Cytometry
Perform multiparameter flow cytometry to increase diagnostic accuracy, particularly for microgranular variants 1. The characteristic APL immunophenotype shows:
- CD34-negative or heterogeneous/dim
- HLA-DR-negative or dim
- CD117-negative/dim
- CD13 and CD33 strongly positive
- CD15 abnormally low (CD15-/dim) versus normal promyelocytes (CD15+++)
- CD2 may be coexpressed in microgranular variants 1
Coagulation Studies
Obtain DIC profile immediately when APL is suspected, including PT, PTT, fibrinogen, D-dimer, and platelet count 1. Coagulation screening must be performed before central line insertion 1.
Additional Molecular Testing for Risk Stratification
Test for FLT3-ITD mutation in all confirmed APL cases for prognostic purposes 1, 4. FLT3-ITD is associated with increased relapse risk and helps identify high-risk patients 4.
Consider testing for additional markers including CD2, CD34, and CD56 expression, as these help identify patients at higher risk of relapse, particularly when associated with high WBC count (>10 × 10⁹/L) 4.
Alternative Diagnostic Methods
Immunofluorescence with anti-PML monoclonal antibody (PG-M3) can provide rapid diagnosis with high sensitivity and specificity, showing a characteristic microgranular nuclear pattern in PML-RARA-positive APL versus normal speckled pattern in non-APL cases 5. This technique is technically simple, fast, and requires only small tissue samples 5.
Critical Pitfalls to Avoid
Do not wait for genetic confirmation before starting ATRA if morphology or clinical presentation suggests APL, as early differentiation syndrome and coagulopathy are life-threatening 1, 3. However, be prepared to discontinue ATRA if genetic tests are negative 1, 3.
Do not assume all morphologically diagnosed APL cases have PML-RARA rearrangement—approximately 10-15% of cases with APL morphology may have alternative genetic abnormalities or no RARA rearrangement 6. These patients have significantly worse outcomes and require different treatment approaches 6.
Ensure all testing is performed in laboratories compliant with regulatory and accreditation requirements 1.