A Multiplexed Amplicon Approach for Detecting Gene Fusions by Next-Generation Sequencing

Carol Beadling, Abigail I. Wald, Andrea Warrick, Tanaya L. Neff, Shan Zhong, Yuri E. Nikiforov, Christopher Corless, Marina N. Nikiforova

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

Chromosomal rearrangements that result in oncogenic gene fusions are clinically important drivers of many cancer types. Rapid and sensitive methods are therefore needed to detect a broad range of gene fusions in clinical specimens that are often of limited quantity and quality. We describe a next-generation sequencing approach that uses a multiplex PCR-based amplicon panel to interrogate fusion transcripts that involve 19 driver genes and 94 partners implicated in solid tumors. The panel also includes control assays that evaluate the 3′/5′ expression ratios of 12 oncogenic kinases, which might be used to infer gene fusion events when the partner is unknown or not included on the panel. There was good concordance between the solid tumor fusion gene panel and other methods, including fluorescence in situ hybridization, real-time PCR, Sanger sequencing, and other next-generation sequencing panels, because 40 specimens known to harbor gene fusions were correctly identified. No specific fusion reads were observed in 59 fusion-negative specimens. The 3′/5′ expression ratio was informative for fusions that involved ALK, RET, and NTRK1 but not for BRAF or ROS1 fusions. However, among 37 ALK or RET fusion-negative specimens, four exhibited elevated 3′/5′ expression ratios, indicating that fusions predicted solely by 3′/5′ read ratios require confirmatory testing.

Original languageEnglish (US)
Pages (from-to)165-175
Number of pages11
JournalJournal of Molecular Diagnostics
Volume18
Issue number2
DOIs
StatePublished - Mar 1 2016

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Gene Fusion
Neoplasms
Multiplex Polymerase Chain Reaction
Fluorescence In Situ Hybridization
Real-Time Polymerase Chain Reaction
Phosphotransferases
Genes

ASJC Scopus subject areas

  • Molecular Medicine
  • Pathology and Forensic Medicine

Cite this

Beadling, C., Wald, A. I., Warrick, A., Neff, T. L., Zhong, S., Nikiforov, Y. E., ... Nikiforova, M. N. (2016). A Multiplexed Amplicon Approach for Detecting Gene Fusions by Next-Generation Sequencing. Journal of Molecular Diagnostics, 18(2), 165-175. https://doi.org/10.1016/j.jmoldx.2015.10.002

A Multiplexed Amplicon Approach for Detecting Gene Fusions by Next-Generation Sequencing. / Beadling, Carol; Wald, Abigail I.; Warrick, Andrea; Neff, Tanaya L.; Zhong, Shan; Nikiforov, Yuri E.; Corless, Christopher; Nikiforova, Marina N.

In: Journal of Molecular Diagnostics, Vol. 18, No. 2, 01.03.2016, p. 165-175.

Research output: Contribution to journalArticle

Beadling, Carol ; Wald, Abigail I. ; Warrick, Andrea ; Neff, Tanaya L. ; Zhong, Shan ; Nikiforov, Yuri E. ; Corless, Christopher ; Nikiforova, Marina N. / A Multiplexed Amplicon Approach for Detecting Gene Fusions by Next-Generation Sequencing. In: Journal of Molecular Diagnostics. 2016 ; Vol. 18, No. 2. pp. 165-175.
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