title: Targeted deep sequencing of mucinous ovarian tumors reveals multiple overlapping RAS-pathway activating mutations in borderline and cancerous neoplasms.
abstract: BACKGROUND
  Mucinous ovarian tumors represent a distinct histotype of epithelial ovarian cancer. The rarest (2-4 % of ovarian carcinomas) of the five major histotypes, their genomic landscape remains poorly described. We undertook hotspot sequencing of 50 genes commonly mutated in human cancer across 69 mucinous ovarian tumors. Our goals were to establish the overall frequency of cancer-hotspot mutations across a large cohort, especially those tumors previously thought to be "RAS-pathway alteration negative", using highly-sensitive next-generation sequencing as well as further explore a small number of cases with apparent heterogeneity in RAS-pathway activating alterations.
  
  
  METHODS
  Using the Ion Torrent PGM platform, we performed next generation sequencing analysis using the v2 Cancer Hotspot Panel. Regions of disparate ERBB2-amplification status were sequenced independently for two mucinous carcinoma (MC) cases, previously established as showing ERBB2 amplification/overexpression heterogeneity, to assess the hypothesis of subclonal populations containing either KRAS mutation or ERBB2 amplification independently or simultaneously.
  
  
  RESULTS
  We detected mutations in KRAS, TP53, CDKN2A, PIK3CA, PTEN, BRAF, FGFR2, STK11, CTNNB1, SRC, SMAD4, GNA11 and ERBB2. KRAS mutations remain the most frequently observed alteration among MC (64.9 %) and mucinous borderline tumors (MBOT) (92.3 %). TP53 mutation occurred more frequently in carcinomas than borderline tumors (56.8 % and 11.5 %, respectively), and combined IHC and mutation data suggest alterations occur in approximately 68 % of MC and as many as 20 % of MBOT. Proven and potential RAS-pathway activating changes were observed in all but one MC. Concurrent ERBB2 amplification and KRAS mutation were observed in a substantial number of cases (7/63 total), as was co-occurrence of KRAS and BRAF mutations (one case). Microdissection of ERBB2-amplified regions of tumors harboring KRAS mutation suggests these alterations are occurring in the same cell populations, while consistency of KRAS allelic frequency in both ERBB2 amplified and non-amplified regions suggests this mutation occurred in advance of the amplification event.
  
  
  CONCLUSIONS
  Overall, the prevalence of RAS-alteration and striking co-occurrence of pathway "double-hits" supports a critical role for tumor progression in this ovarian malignancy. Given the spectrum of RAS-activating mutations, it is clear that targeting this pathway may be a viable therapeutic option for patients with recurrent or advanced stage mucinous ovarian carcinoma, however caution should be exercised in selecting one or more personalized therapeutics given the frequency of non-redundant RAS-activating alterations.
authors.affiliation: Molecular Oncology, BC Cancer Agency Research Centre, Vancouver, Canada. rmackenzie@bccrc.ca.
authors.forename: Robertson
authors.initials: R
authors.lastname: Mackenzie
authors.affiliation: Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. stefan.kommoss@med.uni-tuebingen.de.|Gynecology and Obstetrics, Tuebingen University Hospital, Tuebingen, Germany. stefan.kommoss@med.uni-tuebingen.de.
authors.forename: Stefan
authors.initials: S
authors.lastname: Kommoss
authors.affiliation: Laboratory Medicine and Pathology, Mayo Clinic, Rochester, USA. bwinterhoff@hotmail.com.
authors.forename: Boris J
authors.initials: BJ
authors.lastname: Winterhoff
authors.affiliation: Laboratory Medicine and Pathology, Mayo Clinic, Rochester, USA. Kipp.Benjamin@mayo.edu.
authors.forename: Benjamin R
authors.initials: BR
authors.lastname: Kipp
authors.affiliation: Laboratory Medicine and Pathology, Mayo Clinic, Rochester, USA. garcia.joaquin@mayo.edu.
authors.forename: Joaquin J
authors.initials: JJ
authors.lastname: Garcia
authors.affiliation: Laboratory Medicine and Pathology, Mayo Clinic, Rochester, USA. Voss.Jesse@mayo.edu.
authors.forename: Jesse
authors.initials: J
authors.lastname: Voss
authors.affiliation: Laboratory Medicine and Pathology, Mayo Clinic, Rochester, USA. Halling.Kevin@mayo.edu.
authors.forename: Kevin
authors.initials: K
authors.lastname: Halling
authors.affiliation: Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. akarnezis@bccrc.ca.
authors.forename: Anthony
authors.initials: A
authors.lastname: Karnezis
authors.affiliation: Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. Janine.Senz@vch.ca.
authors.forename: Janine
authors.initials: J
authors.lastname: Senz
authors.affiliation: Molecular Oncology, BC Cancer Agency Research Centre, Vancouver, Canada. wyang@bccrc.ca.
authors.forename: Winnie
authors.initials: W
authors.lastname: Yang
authors.affiliation: Applied Tumor Biology, Institute of Pathology, University of Heidelberg, Heidelberg, Germany. elena.prigge@med.uni-heidelberg.de.
authors.forename: Elena-Sophie
authors.initials: ES
authors.lastname: Prigge
authors.affiliation: Applied Tumor Biology, Institute of Pathology, University of Heidelberg, Heidelberg, Germany. miriam.reuschenbach@med.uni-heidelberg.de.
authors.forename: Miriam
authors.initials: M
authors.lastname: Reuschenbach
authors.affiliation: Applied Tumor Biology, Institute of Pathology, University of Heidelberg, Heidelberg, Germany. knebel@med.uni-heidelberg.de.
authors.forename: Magnus Von Knebel
authors.initials: MV
authors.lastname: Doeberitz
authors.affiliation: Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. Blake.Gilks@vch.ca.
authors.forename: Blake C
authors.initials: BC
authors.lastname: Gilks
authors.affiliation: Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. dhuntsma@bccancer.bc.ca.|Gynecology and Obstetrics, Division of Gynecologic Oncology, University of British Columbia, Vancouver, Canada. dhuntsma@bccancer.bc.ca.
authors.forename: David G
authors.initials: DG
authors.lastname: Huntsman
authors.affiliation: Gynecology and Obstetrics, Mayo Clinic, Rochester, USA. Bakkum.Jamie@mayo.edu.
authors.forename: Jamie
authors.initials: J
authors.lastname: Bakkum-Gamez
authors.affiliation: Gynecology and Obstetrics, Division of Gynecologic Oncology, University of British Columbia, Vancouver, Canada. Jessica.Mcalpine@vch.ca.
authors.forename: Jessica N
authors.initials: JN
authors.lastname: McAlpine
authors.affiliation: Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. manglesio@bccrc.ca.|Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada. manglesio@bccrc.ca.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Anglesio
mesh_terms: D002288:Adenocarcinoma, Mucinous / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D000077216:Carcinoma, Ovarian Epithelial; D005260:Female; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007150:Immunohistochemistry; D009154:Mutation; D059645:Mutation Rate; D009367:Neoplasm Staging; D009375:Neoplasms, Glandular and Epithelial / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D018631:ras Proteins / Q000235:genetics* / Q000378:metabolism*
chemical_list: D016159:Tumor Suppressor Protein p53; D018631:ras Proteins