title: Targeted next-generation sequencing of cancer genes in advanced stage malignant pleural mesothelioma: a retrospective study.
abstract: BACKGROUND
  Malignant pleural mesothelioma (MPM) is a rare malignant disease, and the understanding of molecular pathogenesis has lagged behind other malignancies.
  
  
  METHODS
  A series of 123 formalin-fixed, paraffin-embedded tissue samples with clinical annotations were retrospectively tested with a commercial library kit (Ion AmpliSeq Cancer Hotspot Panel v.2, Life Technologies, Grand Island, NY) to investigate 50 genes plus other two, BRCA1-associated protein-1 (BAP-1) and neurofibromatosis-2 (NF2), frequently altered in MPM. DNA was obtained from tissues after manual microdissection and enriched for at least 50% cancer cells. Variations affecting protein stability or previously correlated to cancer, more frequently identified (≥ 25 patients with at least 10% of allelic frequency), were subsequently evaluated by Sanger sequencing. Immunohistochemistry staining for BAP1 and NF2 proteins was also performed.
  
  
  RESULTS
  The commonest genetic variations were clustered in two main pathways: the p53/DNA repair (TP53, SMACB1, and BAP1) and phosphatidylinositol 3-kinase-AKT pathways (PDGFRA, KIT, KDR, HRAS, PIK3CA, STK11, and NF2). PIK3CA:c.1173A>G mutation, STK11:rs2075606 (T>C), or TP53:rs1042522 (Pro/Pro) was significantly associated with time to progressive disease (TTPD; all p values < 0.01). Furthermore, the accumulation of genetic alterations correlated with shorter TTPD and reduced overall survival (TTPD p value = 0.02, overall survival p value = 0.04). BAP1 genetic variations identified were mainly located in exons 13 and 17, and BAP1 nonsynonymous variations were significantly correlated with BAP1 protein nuclear localization.
  
  
  CONCLUSIONS
  Next-generation sequencing was applied to a relatively large retrospective series of MPM using formalin-fixed, paraffin-embedded archival material. Our results indicate a complex mutational landscape with a higher number of genetic variations in the p53/DNA repair and phosphatidylinositol 3-kinase pathways, some of them with prognostic value.
authors.affiliation: *Department of Oncology, University of Turin, Orbassano, Italy; †Pathology & Oncology Divisions, Saint Antonio and Biagio General Hospital, Alessandria, Italy; and ‡Istituto Superiore di Sanità, Roma, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Lo Iacono
authors.forename: Valentina
authors.initials: V
authors.lastname: Monica
authors.forename: Luisella
authors.initials: L
authors.lastname: Righi
authors.forename: Federica
authors.initials: F
authors.lastname: Grosso
authors.forename: Roberta
authors.initials: R
authors.lastname: Libener
authors.forename: Simona
authors.initials: S
authors.lastname: Vatrano
authors.forename: Paolo
authors.initials: P
authors.lastname: Bironzo
authors.forename: Silvia
authors.initials: S
authors.lastname: Novello
authors.forename: Loredana
authors.initials: L
authors.lastname: Musmeci
authors.forename: Marco
authors.initials: M
authors.lastname: Volante
authors.forename: Mauro
authors.initials: M
authors.lastname: Papotti
authors.forename: Giorgio V
authors.initials: GV
authors.lastname: Scagliotti
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005500:Follow-Up Studies; D020869:Gene Expression Profiling; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D007124:Immunoenzyme Techniques; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D008297:Male; D008654:Mesothelioma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000086002:Mesothelioma, Malignant; D008875:Middle Aged; D009367:Neoplasm Staging; D010997:Pleural Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D011379:Prognosis; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D014408:Biomarkers, Tumor; D012333:RNA, Messenger