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http://purl.uniprot.org/citations/22722791http://www.w3.org/1999/02/22-rdf-syntax-ns#typehttp://purl.uniprot.org/core/Journal_Citation
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Introduction

The presence of the transforming fusion gene echinoderm microtubule-associated protein-like 4 (EML4)-anaplastic lymphoma kinase (ALK) in non-small-cell lung cancer (NSCLC) is a predictive marker for the efficacy of anaplastic lymphoma kinase inhibitors. However, the currently available assays for the detection of the different variants of EML4-ALK have limitations.

Methods

We developed an assay system for the detection of EML4-ALK variants 1, 2, 3a, 3b, 4, 5a, 5b, 6, or 7 transcripts in total RNA obtained from formalin-fixed, paraffin-embedded (FFPE) specimens of NSCLC tissue. The assay is based on region-specific polymerase chain reaction amplification of EML4-ALK complementary DNA followed by specific single-base primer extension and analysis of the extension products by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. The assay was validated by fluorescence in situ hybridization and the results confirmed by subcloning and sequencing of polymerase chain reaction products.

Results

Evaluation of the analytic sensitivity of the assay with serial dilutions of plasmids containing EML4-ALK complementary DNA sequences revealed it to be capable of the reliable detection of one copy of each plasmid per reaction. The assay also detected EML4-ALK variants 1 or 3 in three FFPE samples of surgically resected NSCLC shown to be positive for anaplastic lymphoma kinase rearrangement by fluorescence in situ hybridization. Furthermore, the assay identified variant 1 of EML4-ALK in 3 of 20 FFPE biopsy samples from patients with advanced NSCLC. All positive samples were confirmed by subcloning and sequencing.

Conclusions

Our novel assay is highly sensitive and effective for the detection of EML4-ALK in FFPE specimens."xsd:string
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http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Fujita Y."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Matsumoto K."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Nakagawa K."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Kimura H."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Sakai K."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Takeda M."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Nishio K."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Arao T."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Okamoto I."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Kaneda H."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Hirashima T."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/author"Takezawa K."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/date"2012"xsd:gYear
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/name"J Thorac Oncol"xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/pages"913-918"xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/title"A novel mass spectrometry-based assay for diagnosis of EML4-ALK-positive non-small cell lung cancer."xsd:string
http://purl.uniprot.org/citations/22722791http://purl.uniprot.org/core/volume"7"xsd:string
http://purl.uniprot.org/citations/22722791http://www.w3.org/2004/02/skos/core#exactMatchhttp://purl.uniprot.org/pubmed/22722791
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http://purl.uniprot.org/uniprot/#_Q9HC35-mappedCitation-22722791http://www.w3.org/1999/02/22-rdf-syntax-ns#objecthttp://purl.uniprot.org/citations/22722791
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