The quantitation of BCR-ABL1 mRNA is mandatory for chronic myeloid leukemia (CML) patients, and RT-qPCR is the most extensively used method in testing laboratories worldwide. Nevertheless, substantial variation in RT-qPCR results makes inter-laboratory comparability hard. To facilitate inter-laboratory comparative assessment, an international scale (IS) for BCR-ABL1 was proposed.
The laboratory-specific conversion factor (CF) to the IS can be derived from the World Health Organization (WHO) genetic reference panel; however, this material is limited to the manufacturers to produce and calibrate secondary reference reagents. Therefore, we developed secondary reference calibrators, as lyophilized cellular material, aligned to the IS. Our purpose was both to re-evaluate the CF in 18 previously harmonized laboratories and to propagate the IS to new laboratories.
Our field trial including 30 laboratories across Latin America showed that, after correction of raw BCR-ABL1/ABL1 ratios using CF, the relative mean bias was significantly reduced. We also performed a follow-up of participating laboratories by annually revalidating the process; our results support the need for continuous revalidation of CFs. All participating laboratories also received a calibrator to determine the limit of quantification (LOQ); 90% of them could reproducibly detect BCR-ABL1, indicating that these laboratories can report a consistent deep molecular response. In addition, aiming to investigate the variability of BCR-ABL1 measurements across different RNA inputs, we calculated PCR efficiency for each individual assay by using different amounts of RNA.
In conclusion, for the first time in Latin America, we have successfully organized a harmonization platform for BCR-ABL1 measurement that could be of immediate clinical benefit for monitoring the molecular response of patients in low-resource regions.
We thank Solange Staropoli (IMEX), Gustavo Blandón and Paola Rozo (Genetica Lab), Ruby Rios (UDHO), Ezequiel Zubillaga (CIBIC), Ofelia Berenguer (Hemagen), Gabriel Via (Biogen), Yaribeth Olmedo Pimentel (Instituto Oncologico Nacional), Vanessa Castillo (Caja de Seguro Social), Mariana Debus (Fundaleu), Margarita Bragos (Hospital Centenario), Lorena Zanella (LEB), Clara Pott Godoy (Hospital Dr. Humberto J. Notti), Laura Orellano (Hospital Sor Maria Ludovica), Juan Carlos Ruiz Cabezas (Hospital Juan Tanca Marengo – SOLCA) and Martín Zubieta (Hospital El Cruce).
Author contributions: M.B. and I.L. designed the program of harmonization. M.S.R. and M.B.S performed most of the experiments. M.S.R. and M.B. performed data analysis; all the authors contributed with technical support in RT-qPCR runs, reviewed the data, drafted parts of the manuscript and participated with helpful discussion. M.B. wrote the manuscript and supervised the entire work. All authors read and approved the final manuscript. All authors have accepted responsibility for the entire content of this manuscript and approved its submission.
Research funding: This work was supported by grants from Novartis Argentina, Fundación Mosoteguy and Fundación SALES. M.B., I.L. and J.M. are researchers from the Consejo Nacional de Investigaciones Científicas y Tecnológicas of Argentina (CONICET). M.S.R. and M.B.S. received CONICET fellowships.
Employment or leadership: None declared.
Honorarium: None declared.
Competing interests: Authors state no conflict of interest.
Ethical approval: The local Institutional Review Board deemed the study exempt from review.
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The online version of this article offers supplementary material (https://doi.org/10.1515/cclm-2019-1283).
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