New highly sensitive fluorescence in situ hybridization method to detect PML/RARA fusion in acute promyelocytic leukemia
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Stephanie R Brockman1, Sarah F Paternoster, Rhett P Ketterling
1Division of Laboratory Genetics, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA.
Abstract:
We investigated a new fluorescence in situ hybridization (FISH) method to detect PML/RARA fusion and/or anomalies of the RARA gene (alias RARalpha) in interphase nuclei from patients with acute promyelocytic leukemia (APL). This method uses a commercially available product with two different colored fluorescent probes to detect both PML/RARA gene fusion products (double fusion signal or dual-color fluorescence in situ hybridization [D-FISH]). A total of 82 bone marrow specimens were studied, including 30 from normal bone marrow transplant donors, 33 from patients with untreated APL, 14 from patients with treated APL, and 5 from APL patients with known translocation variants or alternate translocations. The signal patterns and percentage of abnormal nuclei were determined in a blinded study on 500 interphase nuclei for each specimen. Based on 25 normal specimens, the normal cutoff was >0.6% and >1.6% for t(15;17) and t(17;var), respectively. The clinical sensitivity for this series of patients was 98% and the clinical specificity was 100%. The results suggest that the new D-FISH probe set can detect all t(15;17)(q22;q21) and all variant forms of this translocation associated with PML and RARA. In addition, this FISH method can detect all alternate translocations involving RARA and not PML. This FISH method can be used both for the accurate diagnosis of APL and to monitor low levels of disease in treated patients.
Insights
A new dual-color fluorescence in situ hybridization (D-FISH) method accurately detects PML/RARA gene fusions in acute promyelocytic leukemia (APL). This sensitive and specific FISH technique aids in APL diagnosis and monitoring treatment response.
Area of Science:
- Hematology
- Molecular Biology
- Cytogenetics
Background:
- Acute promyelocytic leukemia (APL) is characterized by the PML/RARA fusion gene.
- Accurate detection of PML/RARA is crucial for APL diagnosis and management.
- Traditional methods may have limitations in detecting all variant translocations.
Purpose of the Study:
- To evaluate a new dual-color fluorescence in situ hybridization (D-FISH) method for detecting PML/RARA fusion and RARA gene anomalies.
- To assess the diagnostic accuracy and clinical utility of this D-FISH probe set in APL patients.
Main Methods:
- A commercially available D-FISH probe set was used to analyze 82 bone marrow specimens.
- Specimens included normal donors, untreated APL, treated APL, and APL with variant translocations.
- 500 interphase nuclei per specimen were analyzed in a blinded study to determine signal patterns and abnormal nuclei percentages.
Main Results:
- The D-FISH method demonstrated high clinical sensitivity (98%) and specificity (100%) for APL detection.
- It successfully identified the classic t(15;17) translocation, variant forms, and alternate RARA translocations.
- Established normal cutoffs were >0.6% for t(15;17) and >1.6% for t(17;var).
Conclusions:
- The new D-FISH probe set is a reliable tool for the accurate diagnosis of APL.
- This FISH method is effective in detecting all forms of PML/RARA translocations and other RARA anomalies.
- It can be utilized for monitoring minimal residual disease in treated APL patients.
