Variant-specific discrepancy when quantitating BCR-ABL1 e13a2 and e14a2 transcripts using the Europe Against Cancer qPCR assay.

Autor: Kjaer L; Department of Hematology, Zealand University Hospital, Roskilde, Denmark., Skov V; Department of Hematology, Zealand University Hospital, Roskilde, Denmark., Andersen MT; Department of Clinical Genetics, Rigshospitalet, Copenhagen, Denmark., Aggerholm A; Hemodiagnostic Laboratory, Aarhus University Hospital, Aarhus, Denmark., Clair P; Plateforme PCR, Université de Montpellier, Montpellier, France., Gniot M; Department of Hematology and Stem Cell Transplantation, Poznan University of Medical Sciences, Poznan, Poland., Soeby K; Department of Clinical Biochemistry, Zealand University Hospital, Roskilde, Denmark., Udby L; Department of Hematology, Zealand University Hospital, Roskilde, Denmark., Dorff MH; Department of Hematology, Zealand University Hospital, Roskilde, Denmark., Hasselbalch H; Department of Hematology, Zealand University Hospital, Roskilde, Denmark., Pallisgaard N; Department of Surgical Pathology, Zealand University Hospital, Roskilde, Denmark.
Jazyk: angličtina
Zdroj: European journal of haematology [Eur J Haematol] 2019 Jul; Vol. 103 (1), pp. 26-34. Date of Electronic Publication: 2019 May 01.
DOI: 10.1111/ejh.13238
Abstrakt: Objective: Molecular monitoring of treatment response in patients with chronic myelogenous leukemia is performed using the Europe Against Cancer (EAC) qPCR assay using the International Scale (IS). The assay amplifies both e13a2 and e14a2 BCR-ABL1 transcript variants. Observing distinct variant-dependent amplification curves during qPCR, we aimed to determine if this affected quantitation of BCR-ABL1.
Methods: We investigated the qPCR efficiency at three Danish diagnostic centers (Zealand University Hospital [ZUH], Aarhus University Hospital [AU], and Rigshospitalet [RH]) on cell lines expressing either the e13a2 or e14a2 BCR-ABL1 transcript variants and compared %IS values from 219 chronic myeloid leukemia patients from the centers with either the e13a2 (n = 113) or e14a2 (n = 106) transcript variants obtained by qPCR with absolute quantitation by droplet digital PCR (ddPCR).
Results: Although no significant differences were found in amplification efficiencies of the transcript variants, Bland-Altman analysis of qPCR vs ddPCR values for patient samples revealed a significant average difference in the bias between variants (e3a2/e14a2) of 4.6-, 6.5-, and 1.8-fold for ZUH, AU, and RH, respectively. Furthermore, qPCR %IS values of diagnostic patient samples revealed a significant 4.7-fold difference between the e13a2 and e14a2 variants.
Conclusion: Our findings suggest that the EAC qPCR assay may underestimate the e14a2 variant compared to the e13a2 variant.
(© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)
Databáze: MEDLINE
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