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Abstract
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. 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. 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. 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.
Highlights
Multiplex reverse transcription PCR (RT-PCR) was confirmed to be a reliable technique for detection of ALK fusion transcripts
We propose that diagnostic tools for echinoderm microtubule-associated protein–like 4 gene (EML4)-ALK should be selected in a manner dependent on the available specimen types
FISH and sensitive immunohistochemistry should be applied to formalinfixed, paraffin-embedded tissue, but multiplex reverse transcription (RT)-PCR is appropriate for other specimen types
Summary
Plastic lymphoma kinase gene (ALK) was discovered by functional screening with a non–small cell lung cancer (NSCLC) specimen [1]. EML4 and ALK are located within a short distance ($12 Mbp) of each other on the short arm of human chromosome 2, and a small inversion involving the 2 loci is responsible for generation of the EML4-ALK fusion in lung cancer. Transgenic mice expressing EML4-ALK in lung alveolar cells develop multiple adenocarcinoma nodules soon after birth, but treatment with an ALK inhibitor results in the rapid clearance of such nodules, confirming the addiction of EML4-ALK–positive tumors to the kinase activity of the fusion protein [4]. The therapeutic efficacy of ALK inhibitors has been confirmed in other transgenic mice expressing EML4-ALK [5]
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