Abstract
Introduction
Acute myeloid leukemia (AML) is a heterogeneous clonal disorder often associated with dismal overall survival. The clinical diversity of AML is reflected in the range of recurrent somatic mutations in several genes, many of which have a prognostic and therapeutic value. Targeted next-generation sequencing (NGS) of these genes has the potential for translation into clinical practice. In order to assess this potential, an inter-laboratory evaluation of a commercially available AML gene panel across three diagnostic centres in the UK and Ireland was performed.
Methods
DNA from six AML patient samples was distributed to each centre and processed using a standardised workflow, including a common sequencing platform, sequencing chips and bioinformatics pipeline. A duplicate sample in each centre was run to assess inter- and intra-laboratory performance.
Results
An average sample read depth of 2725X (range 629–5600) was achieved using six samples per chip, with some variability observed in the depth of coverage generated for individual samples and between centres. A total of 16 somatic mutations were detected in the six AML samples, with a mean of 2.7 mutations per sample (range 1–4) representing nine genes on the panel. 15/16 mutations were identified by all three centres. Allelic frequencies of the mutations ranged from 5.6 to 53.3 % (median 44.4 %), with a high level of concordance of these frequencies between centres, for mutations detected.
Conclusion
In this inter-laboratory comparison, a high concordance, reproducibility and robustness was demonstrated using a commercially available NGS AML gene panel and platform.
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References
Buccisano F, Maurillo L, Del Principe MI, Del Poeta G, Sconocchia G, Lo-Coco F, et al. Prognostic and therapeutic implications of minimal residual disease detection in acute myeloid leukemia. Blood. 2012;119(2):332–41.
Dohner H, Weisdorf DJ, Bloomfield CD. Acute myeloid leukemia. N Engl J Med. 2015;373(12):1136–52.
The Cancer Genome Atlas Network. Genomic and epigenomic landscapes of adult de novo acute myeloid leukemia. N Engl J Med. 2013;368(22):2059–74.
Welch JS, Ley TJ, Link DC, Miller CA, Larson DE, Koboldt DC, et al. The origin and evolution of mutations in acute myeloid leukemia. Cell. 2012;150(2):264–78.
Klco JM, Miller CA, Griffith M, Petti A, Spencer DH, Ketkar-Kulkarni S, et al. Association between mutation clearance after induction therapy and outcomes in acute myeloid leukemia. JAMA. 2015;314(8):811–22.
Debarri H, Lebon D, Roumier C, Cheok M, Marceau-Renaut A, Nibourel O, et al. IDH1/2 but not DNMT3A mutations are suitable targets for minimal residual disease monitoring in acute myeloid leukemia patients: a study by the Acute Leukemia French Association. Oncotarget. 2015;6(39):42345–53.
Ivey A, Hills RK, Simpson MA, Jovanovic JV, Gilkes A, Grech A, et al. Assessment of minimal residual disease in standard-risk AML. N Engl J Med. 2016;374(5):422–33.
Falini B, Sportoletti P, Brunetti L, Martelli MP. Perspectives for therapeutic targeting of gene mutations in acute myeloid leukaemia with normal cytogenetics. Br J Haematol. 2015;170(3):305–22.
Kohlmann A, Grossmann V, Nadarajah N, Haferlach T. Next-generation sequencing: feasibility and practicality in haematology. Br J Haematol. 2014;160(6):736–53.
McCourt CM, McArt DG, Mills K, Catherwood MA, Maxwell P, Waugh DJ, et al. Validation of next generation sequencing technologies in comparison to current diagnostic gold standards for BRAF, EGFR, KRAS mutational analysis. PLoS One. 2013;8(7):e69604.
Ivanova M, Shivarov V, Pavlov I, Lilakos K, Naumova E. Clinical evaluation of a novel nine-gene panel for Ion Torrent PGM sequencing of myeloid malignancies. Mol Diagn Ther. 2016;20(1):27–32.
Robinson JT, Thorvaldsdóttir H, Winckler W, Guttman M, Lander ES, Getz G, et al. Integrative genomics viewer. Nat Biotechnol. 2011;29(1):24–6.
Kohlmann A, Klein HU, Weissmann S, Bresolin S, Chaplin T, Cuppens H, et al. The Interlaboratory Robustness of Next-generation sequencing (IRON) study: a deep sequencing investigation of TET2, CBL, and KRAS mutations by an international consortium involving 10 laboratories. Leukemia. 2011;25(12):1840–8.
Grossmann V, Roller A, Klein HU, Weissmann S, Kern W, Haferlach C, et al. Robustness of amplicon deep sequencing underlines its utility in clinical applications. J Mol Diagn. 2014;15(4):473–84.
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Karl Haslam, Mark A. Catherwood, Edwina Dobbin, Anne Sproul, Stephen E. Langabeer and Ken I. Mills declare that they have no competing interests.
Funding
Ken I. Mills was employed by Queen’s University Belfast, and part of this work was supported from the development Grant (R2536CNR) awarded from Leukaemia and Lymphoma Northern Ireland.
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All procedures performed involving human participants were in accordance with the ethical standards of the institution and the 1964 Helsinki declaration or comparable ethical standards.
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Haslam, K., Catherwood, M.A., Dobbin, E. et al. Inter-Laboratory Evaluation of a Next-Generation Sequencing Panel for Acute Myeloid Leukemia. Mol Diagn Ther 20, 457–461 (2016). https://doi.org/10.1007/s40291-016-0222-3
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DOI: https://doi.org/10.1007/s40291-016-0222-3