Author of

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  P2.09 - Pathology (Not CME Accredited Session) (ID 958)

  • Event: WCLC 2018
  • Type: Poster Viewing in the Exhibit Hall
  • Track:
  • Presentations: 1
  • Moderators:
  • Coordinates: 9/25/2018, 16:45 - 18:00, Exhibit Hall

  • 27071

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    P2.09-15 - A Next Generation Sequencing (NGS) RNA-Scan Multiplex Panel (QIAseq) to Identify Gene-Rearranged Non-Small Cell Lung Cancer (ID 12777)

    16:45 - 18:00  |  Author(s): Shuwen Huang
    • Abstract
    • Slides

    Background
    

    Oncogenic fusion gene rearrangements are detected in up to 10% of advanced NSCLC with important therapeutic (eg. ALK, ROS1) or translational impact (eg. RET, NTRK1). The ‘QIAseq’ fusion panel is a 31-gene NGS multiplex assay that synchronously detects multiple oncogene fusion transcripts from formalin-fixed paraffin-embedded (FFPE) tissue derived RNA (Figure 1).

    a9ded1e5ce5d75814730bb4caaf49419 Method
    

    QIAseq analysis was undertaken in 33 samples (31 NSCLC samples, 2 commercial controls). 12/31 NSCLC samples were positive controls with a known fusion genotype identified by Quantide X NGS or FISH +/- RT-PCR. The remaining 19/31 fusion negative NSCLC controls included 6 samples with EGFR/KRAS/NRAS mutations. Analysis required a minimum 2x 5ųM thick FFPE scrolls with >30% neoplastic cell content. Manual RNA extraction was undertaken in all samples except n=6 (ExScale automation extraction). NGS fusion breakpoints, crossing and spanning reads were calculated in QIAseq fusion-detected samples. An additional validation cohort of 40 NSCLC samples, including 20 with unknown fusion status will optimise QIAseq thresholds for fusion detection.

    4c3880bb027f159e801041b1021e88e8 Result
    

    48 QIAseq sequencing experiments was undertaken in 33 samples with ≥2 sequencing runs in 12 samples. QIAseq analysis detected a corresponding NGS fusion breakpoint in 14/14 (100%) positive controls including EML4-ALK (n=8), CLTC-ALK (n=1), CD74-ROS1 (n=3), CCDC6-RET (n=1) and 5-fusion control (n=1). QIAseq analysis was negative in 17/19 (89.4%) negative controls samples including all KRAS (n=4), NRAS (n=1) and EGFR (n=1) mutation samples. QIAseq detected novel fusion gene CD74 Exon 6-CAMK2A Exon 2 in n=1 sample subsequently confirmed on Sanger sequencing. Two separate runs detected TPM3 Exon 8-S100A7A Exon 2 fusion in n=1 sample not identified with Sanger sequencing. Both fusions have uncertain clinical significance. Validation cohort results will be presented.

    8eea62084ca7e541d918e823422bd82e Conclusion
    

    QIAseq detects NSCLC oncogenic fusions with high sensitivity and specificity. Future applications include optimising use of small biopsy specimens for synchronous gene rearrangement screening and identification of novel gene fusion targets.

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