title: Non-invasive analysis of acquired resistance to cancer therapy by sequencing of plasma DNA.
abstract: Cancers acquire resistance to systemic treatment as a result of clonal evolution and selection. Repeat biopsies to study genomic evolution as a result of therapy are difficult, invasive and may be confounded by intra-tumour heterogeneity. Recent studies have shown that genomic alterations in solid cancers can be characterized by massively parallel sequencing of circulating cell-free tumour DNA released from cancer cells into plasma, representing a non-invasive liquid biopsy. Here we report sequencing of cancer exomes in serial plasma samples to track genomic evolution of metastatic cancers in response to therapy. Six patients with advanced breast, ovarian and lung cancers were followed over 1-2 years. For each case, exome sequencing was performed on 2-5 plasma samples (19 in total) spanning multiple courses of treatment, at selected time points when the allele fraction of tumour mutations in plasma was high, allowing improved sensitivity. For two cases, synchronous biopsies were also analysed, confirming genome-wide representation of the tumour genome in plasma. Quantification of allele fractions in plasma identified increased representation of mutant alleles in association with emergence of therapy resistance. These included an activating mutation in PIK3CA (phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha) following treatment with paclitaxel; a truncating mutation in RB1 (retinoblastoma 1) following treatment with cisplatin; a truncating mutation in MED1 (mediator complex subunit 1) following treatment with tamoxifen and trastuzumab, and following subsequent treatment with lapatinib, a splicing mutation in GAS6 (growth arrest-specific 6) in the same patient; and a resistance-conferring mutation in EGFR (epidermal growth factor receptor; T790M) following treatment with gefitinib. These results establish proof of principle that exome-wide analysis of circulating tumour DNA could complement current invasive biopsy approaches to identify mutations associated with acquired drug resistance in advanced cancers. Serial analysis of cancer genomes in plasma constitutes a new paradigm for the study of clonal evolution in human cancers.
authors.affiliation: Cancer Research UK Cambridge Institute and University of Cambridge, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK.
authors.forename: Muhammed
authors.initials: M
authors.lastname: Murtaza
authors.forename: Sarah-Jane
authors.initials: SJ
authors.lastname: Dawson
authors.forename: Dana W Y
authors.initials: DW
authors.lastname: Tsui
authors.forename: Davina
authors.initials: D
authors.lastname: Gale
authors.forename: Tim
authors.initials: T
authors.lastname: Forshew
authors.forename: Anna M
authors.initials: AM
authors.lastname: Piskorz
authors.forename: Christine
authors.initials: C
authors.lastname: Parkinson
authors.forename: Suet-Feung
authors.initials: SF
authors.lastname: Chin
authors.forename: Zoya
authors.initials: Z
authors.lastname: Kingsbury
authors.forename: Alvin S C
authors.initials: AS
authors.lastname: Wong
authors.forename: Francesco
authors.initials: F
authors.lastname: Marass
authors.forename: Sean
authors.initials: S
authors.lastname: Humphray
authors.forename: James
authors.initials: J
authors.lastname: Hadfield
authors.forename: David
authors.initials: D
authors.lastname: Bentley
authors.forename: Tan Min
authors.initials: TM
authors.lastname: Chin
authors.forename: James D
authors.initials: JD
authors.lastname: Brenton
authors.forename: Carlos
authors.initials: C
authors.lastname: Caldas
authors.forename: Nitzan
authors.initials: N
authors.lastname: Rosenfeld
mesh_terms: D000483:Alleles; D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D058534:Class I Phosphatidylinositol 3-Kinases; D004252:DNA Mutational Analysis; D004273:DNA, Neoplasm / Q000032:analysis* / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000235:genetics*; D066246:ErbB Receptors / Q000235:genetics; D019143:Evolution, Molecular; D059472:Exome / Q000235:genetics; D005260:Female; D015894:Genome, Human / Q000235:genetics; D023281:Genomics; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics; D008175:Lung Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D056913:Mediator Complex Subunit 1 / Q000235:genetics; D009369:Neoplasms / Q000188:drug therapy* / Q000235:genetics* / Q000473:pathology; D010051:Ovarian Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics; D010949:Plasma / Q000737:chemistry*; D016160:Retinoblastoma Protein / Q000235:genetics; D000099255:Growth Arrest-Specific Protein 6
chemical_list: D000970:Antineoplastic Agents; D058534:Class I Phosphatidylinositol 3-Kinases; D004273:DNA, Neoplasm; D066246:ErbB Receptors; D036341:Intercellular Signaling Peptides and Proteins; D056913:Mediator Complex Subunit 1; D019869:Phosphatidylinositol 3-Kinases; D016160:Retinoblastoma Protein; D000099255:Growth Arrest-Specific Protein 6; C538855:MED1 protein, human; C484760:PIK3CA protein, human
references.citation: Proc Natl Acad Sci U S A. 2005 Nov 8;102(45):16368-73
references.pmid: 16258065
references.citation: Mol Cancer Ther. 2004 Dec;3(12):1605-13
references.pmid: 15634654
references.citation: N Engl J Med. 2012 Mar 8;366(10):883-892
references.pmid: 22397650
references.citation: Clin Cancer Res. 2009 Mar 15;15(6):2076-84
references.pmid: 19276259
references.citation: Cancer Res. 2009 Sep 1;69(17):6871-8
references.pmid: 19671800
references.citation: Biochem Biophys Res Commun. 2011 Feb 11;405(2):168-72
references.pmid: 21216229
references.citation: Nucleic Acids Res. 2010 Sep;38(16):e164
references.pmid: 20601685
references.citation: Nat Genet. 2011 May;43(5):491-8
references.pmid: 21478889
references.citation: N Engl J Med. 2013 Feb 28;368(9):842-51
references.pmid: 23445095
references.citation: Nat Med. 2008 Sep;14(9):985-90
references.pmid: 18670422
references.citation: Stem Cells. 2012 Mar;30(3):372-8
references.pmid: 22252887
references.citation: Bioinformatics. 2009 Jul 15;25(14):1754-60
references.pmid: 19451168
references.citation: Sci Transl Med. 2010 Dec 8;2(61):61ra91
references.pmid: 21148127
references.citation: BMC Cancer. 2012 Mar 19;12:91
references.pmid: 22429801
references.citation: Clin Cancer Res. 2012 Apr 15;18(8):2391-401
references.pmid: 22492982
references.citation: Cancer Res. 2005 Dec 1;65(23):10992-1000
references.pmid: 16322248
references.citation: Nature. 2012 Jun 28;486(7404):537-40
references.pmid: 22722843
references.citation: Genes Chromosomes Cancer. 2010 Nov;49(11):1062-9
references.pmid: 20725990
references.citation: Clin Chem. 2013 Jan;59(1):211-24
references.pmid: 23065472
references.citation: Cancer Res. 2012 Nov 1;72(21):5625-34
references.pmid: 22964581
references.citation: Nat Genet. 2012 Jul 01;44(8):852-60
references.pmid: 22751098
references.citation: Nat Rev Cancer. 2008 Sep;8(9):714-24
references.pmid: 19143056
references.citation: Nature. 2012 Apr 04;486(7403):395-9
references.pmid: 22495314
references.citation: Breast Cancer Res Treat. 2011 Jan;125(1):55-63
references.pmid: 20224928
references.citation: Carcinogenesis. 2012 Apr;33(4):918-30
references.pmid: 22345290
references.citation: Sci Transl Med. 2012 Nov 28;4(162):162ra154
references.pmid: 23197571
references.citation: PLoS Med. 2005 Mar;2(3):e73
references.pmid: 15737014
references.citation: Bioinformatics. 2009 Aug 15;25(16):2078-9
references.pmid: 19505943
references.citation: EMBO J. 2012 Aug 29;31(17):3524-36
references.pmid: 22828868
references.citation: Sci Transl Med. 2012 May 30;4(136):136ra68
references.pmid: 22649089