Author of

• 15382

  +

  P3.04 - Poster Session/ Biology, Pathology, and Molecular Testing (ID 235)

  • Event: WCLC 2015
  • Type: Poster
  • Track: Biology, Pathology, and Molecular Testing
  • Presentations: 1
  • Moderators:
  • Coordinates: 9/09/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)

  • 15390

    +

    P3.04-008 - Detection of EML4-ALK Fusion Gene by Using Nested Long-Ranged Polymerase Chain Reaction (ID 704)

    09:30 - 09:30  |  Author(s): K. Tsukamoto
    • Abstract

    Background:
    The fusion of the anaplastic lymphoma kinase (ALK) with the echinoderm microtubule-associated protein-like 4 (EML4) was identified in Non-small cell lung cancer (NSCLC). ALK tyrosine kinase inhibitors were proved to be superior to standard chemotherapies and it is important to detect EML4-ALK fusion gene accurately. Reverse transcriptase-polymerase chain reaction (RT-PCR), Fluorescence in situ hybridization (FISH) and Immunohistochemical (IHC) stain are performed clinically to detect the fusion gene. However, there are discrepancies among these methods for detection of EML4-ALK fusion gene and the best detection method remain unknown. The purpose of this study was to evaluate the new method for detection of the EML4-ALK fusion gene.
    
    Methods:
    The combination of nested polymerase chain reaction (PCR) and ｌong-ranged PCR (Nested long-ranged PCR) was used to detect EML4-ALK fusion gene. Genomic deoxyribonucleic acid (gDNA) was extracted from EML4-ALK positive lung cancer cell lines (NCI-H2228,NCI-H3122,ALKSFA8). It was verified whether the fusion gene was amplified. We evaluated the sensitivity of Nested long-ranged PCR for EML4-ALK fusion gene in various ratios. It was confirmed whether the amplification products were EML4-ALK fusion gene by using PCR direct Sequencing.
    
    Results:
    EML4-ALK fusion genes were detectable successfully in each of EML4-ALK positive lung cancer cell lines.NCI-H3122 had EML4-ALK variant 1. ALK-SFA8 and NCI-H2228 had EML4-ALK variant 3a/b. One fusion gene in the presence of 1×10[2] wild type genes was detectable in each cell line. Each PCR product was confirmed by sequencing from both ends.
    
    Conclusion:
    In this study, we were able to detect the fusion gene in vitro by Nested long-ranged PCR. This may become a new diagnostic method for EML4-ALK fusion gene.