Relevance of Tri-color Fluorescence In situ Hybridization in Analysis and Interpretation of Various Atypical Signal Patterns for Characterization of BCR/ABL1 Fusion Gene in Chronic Myeloid Leukemia Patients in Alexandria University Hospitals

Abstract: BACKGROUND: Chronic myelogenous leukemia (CML) is characterized by the t(9;22)(q34;q11.2), which forms the BCR/ABL1 fusion gene. The t(9;22) may be linked to cryptic submicroscopic deletions on chromosomes 9 and/or 22, which maybe associated with unfavorable behavior. Fluorescence in situ hybridization (FISH) analysis is used to determine the number of interphase cells exhibiting BCR/ABL1 fusion with its typical dual-fusion pattern and also can detect cryptic deletions in the form of atypical signal patterns. Serial FISH testing is valuable for follow-up and monitoring therapy response. AIMS AND OBJECTIVES: This work evaluated the implementation of the BCR-ABL1 + 9q34 tri-color plus dual-fusion FISH (D-FISH) system in identifying fusion signal patterns, in comparison with dual-color D-FISH, and to explore its value in detecting BCR/ABL1 fusion and ASS1 gene deletion. MATERIALS AND METHODS: A total of 104 samples from CML patients were analyzed using commercially available BCR-ABL1 dual-color D-FISH and tri-color dual-fusion probes, and their signal profiles were compared. RESULTS: While D-FISH was effective for identifying typical fusion signals, the tri-color probe improved interpretation by distinguishing true fusions from overlapping signals through its aqua-labeled ASS1 component and additionally enabled direct recognition of ASS1 deletions. CONCLUSION: The tri-color dual-fusion probe allows broader detection of molecular variants, reliably distinguishing cryptic deletions from signal overlaps, reduces false positives, and enables direct ASS1 deletion detection, improving BCR/ABL1 fusion diagnosis and measurable residual disease monitoring.