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

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DOI

  • Lasse Kjaer, Zealand University Hospital
  • ,
  • Vibe Skov, Zealand University Hospital
  • ,
  • Morten T. Andersen, Rigshospitalet
  • ,
  • Anni Aggerholm
  • Philippe Clair, Université de Montpellier
  • ,
  • Michal Gniot, Poznan University of Medical Sciences
  • ,
  • Karen Soeby
  • Lene Udby, Zealand University Hospital
  • ,
  • Mikkel H. Dorff, Zealand University Hospital
  • ,
  • Hans Hasselbalch, Zealand University Hospital
  • ,
  • Niels Pallisgaard, Zealand University Hospital

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.

OriginalsprogEngelsk
BogserieEuropean Journal of Haematology
Vol/bind103
Nummer1
Sider (fra-til)26-34
Antal sider9
ISSN0902-4441
DOI
StatusUdgivet - jul. 2019

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