Author of

• 11543

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  P2.06 - Poster Session 2 - Prognostic and Predictive Biomarkers (ID 165)

  • Event: WCLC 2013
  • Type: Poster Session
  • Track: Biology
  • Presentations: 1
  • Moderators:
  • Coordinates: 10/29/2013, 09:30 - 16:30, Exhibit Hall, Ground Level

  • 11544

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    P2.06-001 - Introducing a novel RT-PCR assay for the detection of ROS1 fusion genes in NSCLC (ID 53)

    09:30 - 09:30  |  Author(s): C.L. Stephens
    • Abstract

    Background
    ROS1 is a receptor tyrosine kinase that regulates pathway signaling related to cell proliferation, growth and survival. Chromosomal rearrangements involving ROS1 have recently been identified as potential driver mutations in non-small-cell lung cancers (NSCLC) and preclinical and clinical studies indicate a significant response to crizotinib in tumor cells harboring this mutation. We developed an RT-PCR assay for all ROS1 fusion gene transcripts described in NSCLC to date which is suitable for use with FFPE tissue.

    Methods
    Primers and probes have been specifically designed to detect 14 known fusion transcripts from seven different fusion partners with the ROS1 gene using six multiplexed reactions. These fusion gene pairs include: CD74-ROS1, SLC34A2-ROS1, GOPC (FIG)-ROS1, SDC4-ROS1, EZR-ROS1, TPM3-ROS1, and LRIG3-ROS1. The 14 different ROS1 fusion products were represented by specifically designed and manufactured pCR 2.1-TOPO and pZErO 2.1 plasmids. After screening and identifying ROS1 rearrangements in clinical samples, the specific variants were further differentiated by gel electrophoresis and sequencing.

    Results
    All 14 ROS1 fusion genes could be detected using this RT-PCR assay. We screened 578 FFPE lung cancer samples that were negative for EGFR, KRAS and ALK mutations and detected 12 ROS1 rearrangements in this cohort (2.08%). All positive tumors were adenocarcinomas of the lung, 75% of the patients were female and median age was 53 years. The ROS1 rearrangements that were identified are: SLC34A2 (Exon 13)-ROS1 (Exon 32 and 34), CD74 (Exon 6)-ROS1 (Exon 34), EZR (Exon 10)-ROS1 (Exon 34) and SDC4 (Exon 2)-ROS1 (Exon 32). Messenger RNA expression of ROS1 was significantly higher in ROS1 positive samples than wild-type samples (median 5.2 vs.1.0; p<0.001). An inter-assay comparison of 31 samples containing four ROS1 positives using RT-PCR and FISH showed 100% concordance.

    Conclusion
    The clinical relevance of ROS1 testing in NSCLC and potentially other solid tumors as a predictive biomarker for sensitivity to crizotinib is rapidly increasing. We developed a multiplexed RT-PCR assay that enables the detection of ROS1 fusion genes using FFPE tissue and provide a rapid and cost effective screening tool.