title: Mutational analysis of pulmonary tumours with neuroendocrine features using targeted massive parallel sequencing: a comparison of a neglected tumour group.
abstract: BACKGROUND
  Lung cancer is the leading cause of cancer-related deaths worldwide. The typical and atypical carcinoid (TC and AC), the large-cell neuroendocrine carcinoma (LCNEC) and the small-cell lung cancers (SCLC) are subgroups of pulmonary tumours that show neuroendocrine differentiations. With the rising impact of molecular pathology in routine diagnostics the interest for reliable biomarkers, which can help to differentiate these subgroups and may enable a more personalised treatment of patients, grows.
  
  
  METHODS
  A collective of 70 formalin-fixed, paraffin-embedded (FFPE) pulmonary neuroendocrine tumours (17 TCs, 17 ACs, 19 LCNECs and 17 SCLCs) was used to identify biomarkers by high-throughput sequencing. Using the Illumina TruSeq Amplicon-Cancer Panel on the MiSeq instrument, the samples were screened for alterations in 221 mutation hot spots of 48 tumour-relevant genes.
  
  
  RESULTS
  After filtering >26 000 detected variants by applying strict algorithms, a total of 130 mutations were found in 29 genes and 49 patients. Mutations in JAK3, NRAS, RB1 and VHL1 were exclusively found in SCLCs, whereas the FGFR2 mutation was detected in LCNEC only. KIT, PTEN, HNF1A and SMO were altered in ACs. The SMAD4 mutation corresponded to the TC subtype. We prove that the frequency of mutations increased with the malignancy of tumour type. Interestingly, four out of five ATM-mutated patients showed an additional alteration in TP53, which was by far the most frequently altered gene (28 out of 130; 22%). We found correlations between tumour type and IASLC grade for ATM- (P=0.022; P=0.008) and TP53-mutated patients (P<0.001). Both mutated genes were also associated with lymph node invasion and distant metastasis (P⩽0.005). Furthermore, PIK3CA-mutated patients with high-grade tumours showed a reduced overall survival (P=0.040) and the mutation frequency of APC and ATM in high-grade neuroendocrine lung cancer patients was associated with progression-free survival (PFS) (P=0.020).
  
  
  CONCLUSIONS
  The implementation of high-throughput sequencing for the analysis of the neuroendocrine lung tumours has revealed that, even if these tumours encompass several subtypes with varying clinical aggressiveness, they share a number of molecular features. An improved understanding of the biology of neuroendocrine tumours will offer the opportunity for novel approaches in clinical management, resulting in a better prognosis and prediction of therapeutic response.
authors.affiliation: Institute of Pathology, University Hospital Cologne, Cologne, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Vollbrecht
authors.affiliation: Institute of Pathology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Robert
authors.initials: R
authors.lastname: Werner
authors.affiliation: Institute of Pathology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.|Ruhrlandklinik, West German Lung Centre, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Robert Fred Henry
authors.initials: RF
authors.lastname: Walter
authors.affiliation: Department of Oncology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Daniel Christian
authors.initials: DC
authors.lastname: Christoph
authors.affiliation: Institute of Pathology, University Hospital Cologne, Cologne, Germany.
authors.forename: Lukas Carl
authors.initials: LC
authors.lastname: Heukamp
authors.affiliation: Department of Translational Genomics, Cologne Center of Genomics, University of Cologne, Cologne, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Peifer
authors.affiliation: Institute of Pathology, Molecular Diagnostics, Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Hirsch
authors.affiliation: Institute of Pathology, Molecular Diagnostics, Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Lina
authors.initials: L
authors.lastname: Burbat
authors.affiliation: Institute of Pathology, Helios Klinikum Emil von Behring, Berlin, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Mairinger
authors.affiliation: Institute of Pathology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Kurt Werner
authors.initials: KW
authors.lastname: Schmid
authors.affiliation: Institute of Pathology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Jeremias
authors.initials: J
authors.lastname: Wohlschlaeger
authors.affiliation: Institute of Pathology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Fabian Dominik
authors.initials: FD
authors.lastname: Mairinger
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009154:Mutation; D018358:Neuroendocrine Tumors / Q000235:genetics* / Q000473:pathology; D016612:Paraffin Embedding; D055815:Young Adult