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Published on: April 5, 2018
A prospective PCR-based screening for the EML4-ALK oncogene in non-small cell lung cancer
Manabu Soda1, Kazutoshi Isobe, Akira Inoue
1Division of Functional Genomics, Jichi Medical University, Shimotsukeshi, Tochigi, Japan.
Purpose:
EML4-ALK is a lung cancer oncogene, and ALK inhibitors show marked therapeutic efficacy for tumors harboring this fusion gene. It remains unsettled, however, how the fusion gene should be detected in specimens other than formalin-fixed, paraffin-embedded tissue. We here tested whether reverse transcription PCR (RT-PCR)-based detection of EML4-ALK is a sensitive and reliable approach.
Experimental Design:
We developed a multiplex RT-PCR system to capture ALK fusion transcripts and applied this technique to our prospective, nationwide cohort of non-small cell lung cancer (NSCLC) in Japan.
Results:
During February to December 2009, we collected 916 specimens from 853 patients, quality filtering of which yielded 808 specimens of primary NSCLC from 754 individuals. Screening for EML4-ALK and KIF5B-ALK with our RT-PCR system identified EML4-ALK transcripts in 36 samples (4.46%) from 32 individuals (4.24%). The RT-PCR products were detected in specimens including bronchial washing fluid (n = 11), tumor biopsy (n = 8), resected tumor (n = 7), pleural effusion (n = 5), sputum (n = 4), and metastatic lymph node (n = 1). The results of RT-PCR were concordant with those of sensitive immunohistochemistry with ALK antibodies.
Conclusions:
Multiplex RT-PCR was confirmed to be a reliable technique for detection of ALK fusion transcripts. We propose that diagnostic tools for EML4-ALK should be selected in a manner dependent on the available specimen types. FISH and sensitive immunohistochemistry should be applied to formalin-fixed, paraffin-embedded tissue, but multiplex RT-PCR is appropriate for other specimen types.
Insights
Reverse transcription PCR (RT-PCR) reliably detects EML4-ALK fusion genes in various non-small cell lung cancer specimens. This method offers a sensitive alternative for detecting this lung cancer oncogene when formalin-fixed, paraffin-embedded tissue is unavailable.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- EML4-ALK is a key oncogene in lung cancer.
- ALK inhibitors are effective treatments for tumors with this fusion gene.
- Detection methods for EML4-ALK in non-formalin-fixed tissues are needed.
Purpose of the Study:
- To evaluate the sensitivity and reliability of reverse transcription PCR (RT-PCR) for detecting EML4-ALK fusion genes.
- To assess RT-PCR's utility in specimens beyond formalin-fixed, paraffin-embedded tissue.
Main Methods:
- Development of a multiplex RT-PCR system to detect ALK fusion transcripts.
- Application of the RT-PCR system to a nationwide cohort of Japanese non-small cell lung cancer (NSCLC) patients.
- Analysis of various specimen types including bronchial washing fluid, tumor biopsy, sputum, and pleural effusion.
Main Results:
- EML4-ALK transcripts were identified in 4.46% of 808 primary NSCLC specimens using RT-PCR.
- Detection was successful across diverse sample types, including liquid biopsies.
- RT-PCR results showed concordance with sensitive immunohistochemistry.
Conclusions:
- Multiplex RT-PCR is a reliable method for detecting ALK fusion transcripts.
- RT-PCR is suitable for EML4-ALK detection in various specimen types.
- Diagnostic strategies should consider specimen availability, with RT-PCR being appropriate for non-FFPE samples.
