Intratumor Heterogeneity and Branched Evolution Revealed by Multiregion Sequencing

Marco Gerlinger, Andrew J. Rowan, Stuart Horswell, James Larkin, David Endesfelder, Eva Gronroos, Pierre Martinez, Nicholas Matthews, Aengus Stewart, Patrick Tarpey, Ignacio Varela, Benjamin Phillimore, Sharmin Begum, Neil Q. McDonald, Adam Butler, David Jones, Keiran Raine, Calli Latimer, Claudio R. Santos, Mahrokh NohadaniAron Charles Eklund, Bradley Spencer-Dene, Graham Clark, Lisa Pickering, Gordon Stamp, Martin Gore, Zoltan Imre Szallasi, Julian Downward, P. Andrew Futreal, Charles Swanton

    Research output: Contribution to journalJournal articleResearchpeer-review

    Abstract

    BACKGROUND: Intratumor heterogeneity may foster tumor evolution and adaptation and hinder personalized-medicine strategies that depend on results from single tumor-biopsy samples.METHODSTo examine intratumor heterogeneity, we performed exome sequencing, chromosome aberration analysis, and ploidy profiling on multiple spatially separated samples obtained from primary renal carcinomas and associated metastatic sites. We characterized the consequences of intratumor heterogeneity using immunohistochemical analysis, mutation functional analysis, and profiling of messenger RNA expression.RESULTS: Phylogenetic reconstruction revealed branched evolutionary tumor growth, with 63 to 69% of all somatic mutations not detectable across every tumor region. Intratumor heterogeneity was observed for a mutation within an autoinhibitory domain of the mammalian target of rapamycin (mTOR) kinase, correlating with S6 and 4EBP phosphorylation in vivo and constitutive activation of mTOR kinase activity in vitro. Mutational intratumor heterogeneity was seen for multiple tumor-suppressor genes converging on loss of function; SETD2, PTEN, and KDM5C underwent multiple distinct and spatially separated inactivating mutations within a single tumor, suggesting convergent phenotypic evolution. Gene-expression signatures of good and poor prognosis were detected in different regions of the same tumor. Allelic composition and ploidy profiling analysis revealed extensive intratumor heterogeneity, with 26 of 30 tumor samples from four tumors harboring divergent allelic-imbalance profiles and with ploidy heterogeneity in two of four tumors.CONCLUSIONS: Intratumor heterogeneity can lead to underestimation of the tumor genomics landscape portrayed from single tumor-biopsy samples and may present major challenges to personalized-medicine and biomarker development. Intratumor heterogeneity, associated with heterogeneous protein function, may foster tumor adaptation and therapeutic failure through Darwinian selection. (Funded by the Medical Research Council and others.)
    Original languageEnglish
    JournalThe New England Journal of Medicine
    Volume366
    Issue number10
    Pages (from-to)883-892
    ISSN0028-4793
    Publication statusPublished - 2012

    Keywords

    • Primates Mammalia Vertebrata Chordata Animalia (Animals, Chordates, Humans, Mammals, Primates, Vertebrates) - Hominidae [86215] human common
    • human KDM5C gene [Hominidae]
    • human PTEN gene [Hominidae]
    • human SETD2 gene [Hominidae]
    • mammalian target of rapamycin kinase
    • messenger RNA mRNA
    • 03502, Genetics - General
    • 03508, Genetics - Human
    • 10062, Biochemistry studies - Nucleic acids, purines and pyrimidines
    • 15506, Urinary system - Pathology
    • 24004, Neoplasms - Pathology, clinical aspects and systemic effects
    • Molecular Genetics
    • Nephrology
    • Oncology
    • renal carcinoma Kidney Neoplasms (MeSH) Carcinoma (MeSH) urologic disease, neoplastic disease
    • branched evolution
    • intratumor heterogeneity
    • Biochemistry and Molecular Biophysics
    • Human Medicine, Medical Sciences
    • chromosome aberration analysis laboratory techniques, genetic techniques
    • exome sequencing laboratory techniques, genetic techniques
    • immunohistochemical analysis laboratory techniques, immunologic techniques
    • multiregion sequencing laboratory techniques, genetic techniques
    • mutation functional analysis laboratory techniques, genetic techniques
    • ploidy profiling laboratory techniques, genetic techniques

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