A prospective PCR-based screening for the EML4-ALK oncogene in non-small cell lung cancer.
OBJECTIVE
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.


METHODS
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.
Division of Functional Genomics, Jichi Medical University, Shimotsukeshi, Tochigi, Japan.
Manabu
M
Soda
Kazutoshi
K
Isobe
Akira
A
Inoue
Makoto
M
Maemondo
Satoshi
S
Oizumi
Yuka
Y
Fujita
Akihiko
A
Gemma
Yoshihiro
Y
Yamashita
Toshihide
T
Ueno
Kengo
K
Takeuchi
Young Lim
YL
Choi
Hitoshi
H
Miyazawa
Tomoaki
T
Tanaka
Koichi
K
Hagiwara
Hiroyuki
H
Mano
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D007564:Japan; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011446:Prospective Studies
C522504:EML4-ALK fusion protein, human; D015514:Oncogene Proteins, Fusion