Development and evaluation of a secondary reference panel for BCR-ABL1 quantification on the International Scale

Area of Science:

• Hematology
• Molecular Diagnostics
• Oncology

Background:

• Accurate BCR-ABL1 molecular monitoring standardized to the International Scale (IS) is crucial for managing chronic myeloid leukemia (CML), particularly for treatment cessation decisions.
• Current IS calibration relies on time-consuming sample exchange with reference labs, and access to the World Health Organization (WHO) BCR-ABL1 reference panel is limited.

Purpose of the Study:

• To develop the first cell-based secondary reference panel for BCR-ABL1 testing, traceable to the WHO panel.
• To provide a more accessible and reliable tool for International Scale (IS) calibration and assay validation worldwide.
• To assess the performance of BCR-ABL1 assays using the developed secondary reference panel.

Main Methods:

• Development of a cell-based secondary reference panel with levels from MR(1) to MR(4.5).
• Calibration of the panel to IS using digital PCR with ABL1, BCR, and GUSB as reference genes.
• Evaluation of the panel by 44 laboratories globally, assessing assay optimization, IS accuracy, precision, and sensitivity.

Main Results:

• >40% of BCR-ABL1 assays exhibited inadequate optimization (e.g., poor linearity, suboptimal PCR efficiency).
• When using optimized inputs, >60% of assays showed satisfactory IS accuracy, precision, and/or MR(4.5) sensitivity.
• 58% of labs obtained IS conversion factors from the secondary panel concordant with their existing values; assay configuration did not significantly alter %BCR-ABL1 results.

Conclusions:

• The developed secondary reference panel is traceable to the WHO panel and replicates its performance.
• Significant assay optimization issues were identified, highlighting the need for improved IS calibration mechanisms.
• The panel demonstrates potential for enhancing the accuracy and reliability of BCR-ABL1 testing in global molecular diagnostics.