Imaging flow cytometry-based multiplex FISH for three IGH translocations in multiple myeloma

Area of Science:

• Hematology
• Molecular Biology
• Oncology

Background:

• Specific chromosomal translocations, including t(4;14), t(14;16), and t(11;14), are critical prognostic markers in multiple myeloma (MM).
• Accurate and sensitive detection of these translocations is essential for guiding therapeutic strategies and improving patient outcomes.

Purpose of the Study:

• To develop and validate a novel diagnostic method, Immunophenotyped-Suspension-Multiplex (ISM)-FISH, for the simultaneous detection of three major IGH translocations in MM.
• To assess the sensitivity, specificity, and concordance of ISM-FISH compared to standard FISH techniques.

Main Methods:

• Developed ISM-FISH by combining immunophenotyping (anti-CD138) with multiplex FISH (IGH, FGFR3, MAF, CCND1 probes) in suspension.
• Utilized imaging flow cytometry (MI-1000) and FISH spot counting for simultaneous analysis of translocations in CD138-positive cells.
• Validated the method on bone marrow nucleated cells from 70 patients with MM or monoclonal gammopathy of undetermined significance.

Main Results:

• ISM-FISH successfully detected t(11;14), t(4;14), and t(14;16) in CD138-positive MM cells with high sensitivity (down to 0.1%).
• The method demonstrated superior sensitivity compared to standard double-color (DC) FISH, detecting translocations in over 2.5 x 10^4 cells.
• ISM-FISH showed high concordance with standard DC-FISH (positive: 96.6%, negative: 98.8%).

Conclusions:

• ISM-FISH is a rapid, reliable, and sensitive diagnostic tool for simultaneous detection of three critical IGH translocations in multiple myeloma.
• This technique has the potential to enhance risk-adapted, individualized therapy selection for MM patients.
• The ISM-FISH assay offers improved diagnostic capabilities for MM stratification.