title: The Subclonal Architecture of Metastatic Breast Cancer: Results from a Prospective Community-Based Rapid Autopsy Program "CASCADE".
abstract: BACKGROUND
  Understanding the cancer genome is seen as a key step in improving outcomes for cancer patients. Genomic assays are emerging as a possible avenue to personalised medicine in breast cancer. However, evolution of the cancer genome during the natural history of breast cancer is largely unknown, as is the profile of disease at death. We sought to study in detail these aspects of advanced breast cancers that have resulted in lethal disease.
  
  
  RESULTS
  Three patients with oestrogen-receptor (ER)-positive, human epidermal growth factor receptor 2 (HER2)-negative breast cancer and one patient with triple negative breast cancer underwent rapid autopsy as part of an institutional prospective community-based rapid autopsy program (CASCADE). Cases represented a range of management problems in breast cancer, including late relapse after early stage disease, de novo metastatic disease, discordant disease response, and disease refractory to treatment. Between 5 and 12 metastatic sites were collected at autopsy together with available primary tumours and longitudinal metastatic biopsies taken during life. Samples underwent paired tumour-normal whole exome sequencing and single nucleotide polymorphism (SNP) arrays. Subclonal architectures were inferred by jointly analysing all samples from each patient. Mutations were validated using high depth amplicon sequencing. Between cases, there were significant differences in mutational burden, driver mutations, mutational processes, and copy number variation. Within each case, we found dramatic heterogeneity in subclonal structure from primary to metastatic disease and between metastatic sites, such that no single lesion captured the breadth of disease. Metastatic cross-seeding was found in each case, and treatment drove subclonal diversification. Subclones displayed parallel evolution of treatment resistance in some cases and apparent augmentation of key oncogenic drivers as an alternative resistance mechanism. We also observed the role of mutational processes in subclonal evolution. Limitations of this study include the potential for bias introduced by joint analysis of formalin-fixed archival specimens with fresh specimens and the difficulties in resolving subclones with whole exome sequencing. Other alterations that could define subclones such as structural variants or epigenetic modifications were not assessed.
  
  
  CONCLUSIONS
  This study highlights various mechanisms that shape the genome of metastatic breast cancer and the value of studying advanced disease in detail. Treatment drives significant genomic heterogeneity in breast cancers which has implications for disease monitoring and treatment selection in the personalised medicine paradigm.
authors.affiliation: Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Peter
authors.identifier: http://orcid.org/0000-0001-5999-428X
authors.initials: P
authors.lastname: Savas
authors.affiliation: Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Zhi Ling
authors.identifier: http://orcid.org/0000-0002-0748-0858
authors.initials: ZL
authors.lastname: Teo
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia.|Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Victoria, Australia.
authors.forename: Christophe
authors.identifier: http://orcid.org/0000-0002-2876-0648
authors.initials: C
authors.lastname: Lefevre
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia.|Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Victoria, Australia.
authors.forename: Christoffer
authors.initials: C
authors.lastname: Flensburg
authors.affiliation: Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Franco
authors.identifier: http://orcid.org/0000-0002-5086-2544
authors.initials: F
authors.lastname: Caramia
authors.affiliation: Cancer Genomics Program, Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Kathryn
authors.initials: K
authors.lastname: Alsop
authors.affiliation: Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Mariam
authors.initials: M
authors.lastname: Mansour
authors.affiliation: Division of Cancer Medicine, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Prudence A
authors.initials: PA
authors.lastname: Francis
authors.affiliation: Cancer Genomics Program, Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.|kConFab, Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Heather A
authors.initials: HA
authors.lastname: Thorne
authors.affiliation: Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Maria Joao
authors.initials: MJ
authors.lastname: Silva
authors.affiliation: UCL Cancer Institute, CRUK Lung Cancer Centre of Excellence, London, United Kingdom.
authors.forename: Nnennaya
authors.initials: N
authors.lastname: Kanu
authors.affiliation: UCL Cancer Institute, CRUK Lung Cancer Centre of Excellence, London, United Kingdom.
authors.forename: Michelle
authors.initials: M
authors.lastname: Dietzen
authors.affiliation: The Francis Crick Institute, London, United Kingdom.
authors.forename: Andrew
authors.identifier: http://orcid.org/0000-0003-1926-9707
authors.initials: A
authors.lastname: Rowan
authors.affiliation: University of Applied Sciences Koblenz, RheinAhrCampus Remagen, Department of Mathematics and Technology, Remagen, Germany.
authors.forename: Maik
authors.identifier: http://orcid.org/0000-0003-3977-9982
authors.initials: M
authors.lastname: Kschischo
authors.affiliation: Department of Pathology, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
authors.forename: Stephen
authors.identifier: http://orcid.org/0000-0002-7648-8896
authors.initials: S
authors.lastname: Fox
authors.affiliation: Cancer Genomics Program, Division of Research, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.|Sir Peter MacCallum Department of Oncology, the University of Melbourne, Victoria, Australia.
authors.forename: David D
authors.initials: DD
authors.lastname: Bowtell
authors.affiliation: Division of Cancer Medicine, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.|Sir Peter MacCallum Department of Oncology, the University of Melbourne, Victoria, Australia.
authors.forename: Sarah-Jane
authors.identifier: http://orcid.org/0000-0002-8276-0374
authors.initials: SJ
authors.lastname: Dawson
authors.affiliation: Bioinformatics Division, Walter & Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia.|Department of Mathematics and Statistics, University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Terence P
authors.initials: TP
authors.lastname: Speed
authors.affiliation: UCL Cancer Institute, CRUK Lung Cancer Centre of Excellence, London, United Kingdom.|The Francis Crick Institute, London, United Kingdom.
authors.forename: Charles
authors.initials: C
authors.lastname: Swanton
authors.affiliation: Division of Cancer Medicine, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.|Sir Peter MacCallum Department of Oncology, the University of Melbourne, Victoria, Australia.
authors.forename: Sherene
authors.initials: S
authors.lastname: Loi
mesh_terms: D000328:Adult; D001344:Autopsy; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology*; D055812:Community-Based Participatory Research; D059472:Exome; D005260:Female; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D009154:Mutation; D020641:Polymorphism, Single Nucleotide