title: Inconsistent results in the analysis of ALK rearrangements in non-small cell lung cancer.
abstract: Identification of targetable EML4-ALK fusion proteins has revolutionized the treatment of a minor subgroup of non-small cell lung cancer (NSCLC) patients. Although fluorescence in situ hybridization (FISH) is regarded as the gold standard for detection of ALK rearrangements, ALK immunohistochemistry (IHC) is often used as screening tool in clinical practice. In order to unbiasedly analyze the diagnostic impact of such a screening strategy, we compared ALK IHC with ALK FISH in three large representative Swedish NSCLC cohorts incorporating clinical parameters and gene expression data.
  
  
  
  ALK rearrangements were detected using FISH on tissue microarrays (TMAs), including tissue from 851 NSCLC patients. In parallel, ALK protein expression was detected using IHC, applying the antibody clone D5F3 with two different protocols (the FDA approved Ventana CDx assay and our in house Dako IHC protocol). Gene expression microarray data (Affymetrix) was available for 194 patients.
  
  
  
  ALK rearrangements were detected in 1.7 % in the complete cohort and 2.0 % in the non-squamous cell carcinoma subgroup. ALK protein expression was observed in 1.8 and 1.4 % when applying the Ventana assay or the in house Dako protocol, respectively. The specificity and accuracy of IHC was high (> 98 %), while the sensitivity was between 69 % (Ventana) and 62 % (in house Dako protocol). Furthermore, only 67 % of the ALK IHC positive cases were positive with both IHC assays. Gene expression analysis revealed that 6/194 (3 %) tumors showed high ALK gene expression (≥ 6 AU) and of them only three were positive by either FISH or IHC.
  
  
  
  The overall frequency of ALK rearrangements based on FISH was lower than previously reported. The sensitivity of both IHC assays was low, and the concordance between the FISH and the IHC assays poor, questioning current strategies to screen with IHC prior to FISH or completely replace FISH by IHC.
authors: Department of Immunology, Genetics and Pathology, Uppsala University, 751 85, Uppsala, Sweden. johanna.mattsson@igp.uu.se.
authors: Johanna S M
authors: JS
authors: Mattsson
authors: Department of Clinical Sciences Lund, Division of Oncology and Pathology, Lund University, Lund, Sweden.|Department of Pathology, Regional Laboratories Region Skåne, SE-221 85, Lund, Sweden.
authors: Hans
authors: H
authors: Brunnström
authors: Department of Statistics, TU Dortmund University, Dortmund, Germany.
authors: Verena
authors: V
authors: Jabs
authors: Leibniz Research Centre for Working Environment and Human Factors (IfADo) at Dortmund TU, Dortmund, Germany.
authors: Karolina
authors: K
authors: Edlund
authors: Department of Clinical Sciences Lund, Division of Oncology and Pathology, Lund University, Lund, Sweden.
authors: Karin
authors: K
authors: Jirström
authors: Department of Medical Sciences, Respiratory, Allergy and Sleep Research, Uppsala University, Uppsala, Sweden.
authors: Stephanie
authors: S
authors: Mindus
authors: Department of Immunology, Genetics and Pathology, Uppsala University, 751 85, Uppsala, Sweden.
authors: Linnéa
authors: L
authors: la Fleur
authors: Department of Immunology, Genetics and Pathology, Uppsala University, 751 85, Uppsala, Sweden.
authors: Fredrik
authors: F
authors: Pontén
authors: Department of Research and Education, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
authors: Mats G
authors: MG
authors: Karlsson
authors: School of Health Sciences, Örebro University, Örebro, Sweden.
authors: Christina
authors: C
authors: Karlsson
authors: Department of Respiratory Medicine, Gävle hospital, Gävle; Centre for Research and Development, Uppsala University/County Council of Gävleborg, Gävle, Sweden.
authors: Hirsh
authors: H
authors: Koyi
authors: Department of Respiratory Medicine, Gävle hospital, Gävle; Centre for Research and Development, Uppsala University/County Council of Gävleborg, Gävle, Sweden.
authors: Eva
authors: E
authors: Brandén
authors: Department of Immunology, Genetics and Pathology, Uppsala University, 751 85, Uppsala, Sweden.
authors: Johan
authors: J
authors: Botling
authors: Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
authors: Gisela
authors: G
authors: Helenius
authors: Department of Immunology, Genetics and Pathology, Uppsala University, 751 85, Uppsala, Sweden.
authors: Patrick
authors: P
authors: Micke
authors: Clinical Research Center, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
authors: Maria A
authors: MA
authors: Svensson
mesh_terms: D000077548:Anaplastic Lymphoma Kinase; D014408:Biomarkers, Tumor / Q000032:analysis* / Q000235:genetics; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics*; D015331:Cohort Studies; D015321:Gene Rearrangement; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D008175:Lung Neoplasms / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics*; D012680:Sensitivity and Specificity; D046888:Tissue Array Analysis
chemical_list: D014408:Biomarkers, Tumor; C000626173:ALK protein, human; D000077548:Anaplastic Lymphoma Kinase; D020794:Receptor Protein-Tyrosine Kinases