title: Whole genome sequence analysis suggests intratumoral heterogeneity in dissemination of breast cancer to lymph nodes.
abstract: BACKGROUND
  Intratumoral heterogeneity may help drive resistance to targeted therapies in cancer. In breast cancer, the presence of nodal metastases is a key indicator of poorer overall survival. The aim of this study was to identify somatic genetic alterations in early dissemination of breast cancer by whole genome next generation sequencing (NGS) of a primary breast tumor, a matched locally-involved axillary lymph node and healthy normal DNA from blood.
  
  
  METHODS
  Whole genome NGS was performed on 12 µg (range 11.1-13.3 µg) of DNA isolated from fresh-frozen primary breast tumor, axillary lymph node and peripheral blood following the DNA nanoball sequencing protocol. Single nucleotide variants, insertions, deletions, and substitutions were identified through a bioinformatic pipeline and compared to CIN25, a key set of genes associated with tumor metastasis.
  
  
  RESULTS
  Whole genome sequencing revealed overlapping variants between the tumor and node, but also variants that were unique to each. Novel mutations unique to the node included those found in two CIN25 targets, TGIF2 and CCNB2, which are related to transcription cyclin activity and chromosomal stability, respectively, and a unique frameshift in PDS5B, which is required for accurate sister chromatid segregation during cell division. We also identified dominant clonal variants that progressed from tumor to node, including SNVs in TP53 and ARAP3, which mediates rearrangements to the cytoskeleton and cell shape, and an insertion in TOP2A, the expression of which is significantly associated with tumor proliferation and can segregate breast cancers by outcome.
  
  
  CONCLUSIONS
  This case study provides preliminary evidence that primary tumor and early nodal metastasis have largely overlapping somatic genetic alterations. There were very few mutations unique to the involved node. However, significant conclusions regarding early dissemination needs analysis of a larger number of patient samples.
authors.affiliation: Department of Cancer Studies and Molecular Medicine, Robert Kilpatrick Clinical Sciences Building, Leicester Royal Infirmary, Leicester, LE2 7LX, United Kingdom.
authors.forename: Kevin
authors.initials: K
authors.lastname: Blighe
authors.affiliation: Division of Cancer, Imperial College, Hammersmith Hospital Campus, London, W12 0NN, United Kingdom.
authors.forename: Laura
authors.initials: L
authors.lastname: Kenny
authors.affiliation: Division of Cancer, Imperial College, Hammersmith Hospital Campus, London, W12 0NN, United Kingdom.
authors.forename: Naina
authors.initials: N
authors.lastname: Patel
authors.affiliation: Department of Cancer Studies and Molecular Medicine, Robert Kilpatrick Clinical Sciences Building, Leicester Royal Infirmary, Leicester, LE2 7LX, United Kingdom.
authors.forename: David S
authors.initials: DS
authors.lastname: Guttery
authors.affiliation: Department of Cancer Studies and Molecular Medicine, Robert Kilpatrick Clinical Sciences Building, Leicester Royal Infirmary, Leicester, LE2 7LX, United Kingdom.
authors.forename: Karen
authors.initials: K
authors.lastname: Page
authors.affiliation: Division of Cancer, Imperial College, Hammersmith Hospital Campus, London, W12 0NN, United Kingdom.
authors.forename: Julian H
authors.initials: JH
authors.lastname: Gronau
authors.affiliation: Division of Cancer, Imperial College, Hammersmith Hospital Campus, London, W12 0NN, United Kingdom.
authors.forename: Cyrus
authors.initials: C
authors.lastname: Golshani
authors.affiliation: Division of Cancer, Imperial College, Hammersmith Hospital Campus, London, W12 0NN, United Kingdom.
authors.forename: Justin
authors.initials: J
authors.lastname: Stebbing
authors.affiliation: Division of Cancer, Imperial College, Hammersmith Hospital Campus, London, W12 0NN, United Kingdom.
authors.forename: R Charles
authors.initials: RC
authors.lastname: Coombes
authors.affiliation: Department of Cancer Studies and Molecular Medicine, Robert Kilpatrick Clinical Sciences Building, Leicester Royal Infirmary, Leicester, LE2 7LX, United Kingdom.
authors.forename: Jacqueline A
authors.initials: JA
authors.lastname: Shaw
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics; D000951:Antigens, Neoplasm / Q000235:genetics; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D056765:Cyclin B2 / Q000235:genetics; D004250:DNA Topoisomerases, Type II / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D005260:Female; D020690:GTPase-Activating Proteins / Q000235:genetics; D018740:Genetic Heterogeneity; D015894:Genome, Human; D018398:Homeodomain Proteins / Q000235:genetics; D006801:Humans; D008207:Lymphatic Metastasis; D000075223:Poly-ADP-Ribose Binding Proteins; D020641:Polymorphism, Single Nucleotide; D012097:Repressor Proteins / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: C446512:ARAP3 protein, human; D048868:Adaptor Proteins, Signal Transducing; D000951:Antigens, Neoplasm; C503758:CCNB2 protein, human; D056765:Cyclin B2; D004268:DNA-Binding Proteins; D020690:GTPase-Activating Proteins; D018398:Homeodomain Proteins; C118660:PDS5B protein, human; D000075223:Poly-ADP-Ribose Binding Proteins; D012097:Repressor Proteins; C414585:TGIF2 protein, human; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D004250:DNA Topoisomerases, Type II; C000618811:TOP2A protein, human