domingo, 27 de março de 2011

 NATURE | ARTICLE OPEN

Initial genome sequencing and analysis of multiple myeloma

Nature
 
471,
 
467–472
 
(24 March 2011)
 
doi:10.1038/nature09837
Received
 
 
Accepted
 
 
Published online
 

Abstract

Multiple myeloma is an incurable malignancy of plasma cells, and its pathogenesis is poorly understood. Here we report the massively parallel sequencing of 38 tumour genomes and their comparison to matched normal DNAs. Several new and unexpected oncogenic mechanisms were suggested by the pattern of somatic mutation across the data set. These include the mutation of genes involved in protein translation (seen in nearly half of the patients), genes involved in histone methylation, and genes involved in blood coagulation. In addition, a broader than anticipated role of NF-κB signalling was indicated by mutations in 11 members of the NF-κB pathway. Of potential immediate clinical relevance, activating mutations of the kinase BRAF were observed in 4% of patients, suggesting the evaluation of BRAF inhibitors in multiple myeloma clinical trials. These results indicate that cancer genome sequencing of large collections of samples will yield new insights into cancer not anticipated by existing knowledge.

Subject terms:

Figures at a glance

left
right

Introduction

Multiple myeloma is an incurable malignancy of mature B-lymphoid cells, and its pathogenesis is only partially understood. About 40% of cases harbour chromosome translocations resulting in overexpression of genes (including CCND1, CCND3, MAF, MAFB, WHSC1 (also called MMSET) andFGFR3) via their juxtaposition to the immunoglobulin heavy chain (IgH) locus1. Other cases exhibit hyperdiploidy. However, these abnormalities are probably insufficient for malignant transformation because they are also observed in the pre-malignant syndrome known as monoclonal gammopathy of uncertain significance. Malignant progression events include activation of MYC, FGFR3, KRAS andNRAS and activation of the NF-κB pathway1, 2, 3. More recently, loss-of-function mutations in the histone demethylase UTX (also called KDM6A) have also been reported4.
A powerful way to understand the molecular basis of cancer is to sequence either the entire genome or the protein-coding exome, comparing tumour to normal from the same patient to identify the acquired somatic mutations. Recent reports have described the sequencing of whole genomes from a single patient5, 6, 7, 8, 9. Although informative, we hypothesized that a larger number of cases would permit the identification of biologically relevant patterns that would not otherwise be evident.

Discussion

The analysis of multiple myeloma genomes reveals that mechanisms previously suspected to have a role in the biology of multiple myeloma (for example, NF-κB activation and HMT dysfunction) may have broad roles by virtue of mutations in multiple members of these pathways. In addition, potentially new mechanisms of transformation are suggested, including mutations in the RNA exonuclease DIS3 and other genes involved in protein translation and homeostasis. Whether these mutations are unique to multiple myeloma or are common to other cancers remains to be determined. Furthermore, frequent mutations in the oncogenic kinase BRAF were observed—a finding that has immediate clinical translational implications.
Importantly, most of these discoveries could not have been made by sequencing only a single multiple myeloma genome—the complex patterns of pathway dysregulation required the analysis of multiple genomes. Whole-exome sequencing revealed the substantial majority of the significantly mutated genes. However, we note that half of total protein-coding mutations occurred via chromosomal aberrations such as translocations, most of which would not have been discovered by sequencing of the exome alone. Similarly, the recurrent point mutations in non-coding regions would have been missed with sequencing directed only at coding exons.
The analysis described here is preliminary. Additional multiple myeloma genomes will be required to establish the definitive genomic landscape of the disease and determine accurate estimates of mutation frequency in the disease. The sequence data described here will be available from the dbGaP repository (http://www.ncbi.nlm.nih.gov/gap) and we have created a multiple myeloma Genomics Portal (http://www.broadinstitute.org/mmgp) to support data analysis and visualization.

Methods

Informed consent from multiple myeloma patients was obtained in line with the Declaration of Helsinki. DNA was extracted from bone marrow aspirate (tumour) and blood (normal). WGS libraries (370–410-bp inserts) and WES libraries (200–350-bp inserts) were constructed and sequenced on an Illumina GA-II sequencer using 101- and 76-bp paired-end reads, respectively. Sequencing reads were processed with the Firehose pipeline, identifying somatic point mutations, indels and other structural chromosomal rearrangements. Structural rearrangements affecting protein-coding regions were then subjected to manual review to exclude alignment artefacts. True positive mutation rates were estimated by Sequenom mass spectrometry genotyping of randomly selected mutations. HOXA9 short hairpin (sh)RNAs were introduced into multiple myeloma cell lines using lentiviral infection using standard methods.
A complete description of the materials and methods is provided in the Supplementary Information.

Accession codes

Primary accessions

Gene Expression Omnibus

Author information

Affiliations

  1. The Eli and Edythe L. Broad Institute, 7 Cambridge Center, Cambridge, Massachusetts 02412, USA

    • Michael A. Chapman,
      •  
    • Michael S. Lawrence,
      •  
    • Kristian Cibulskis,
      •  
    • Carrie Sougnez,
      •  
    • Christina L. Harview,
    • Jean-Philippe Brunet,
      •  
    • Mazhar Adli,
      •  
    • Kristin G. Ardlie,
      •  
    • Bradley E. Bernstein,
      •  
    • Yotam Drier,
      •  
    • Stacey B. Gabriel,
      •  
    • Ted Liefeld,
      •  
    • Scott Mahan,
      •  
    • Stefano Monti,
      •  
    • Robb Onofrio,
      •  
    • Trevor J. Pugh,
      •  
    • Alex H. Ramos,
    • Andrey Sivachenko,
      •  
    • Douglas Voet,
      •  
    • Wendy Winckler,
      •  
    • William C. Hahn,
      •  
    • Levi A. Garraway,
      •  
    • Matthew Meyerson,
      •  
    • Eric S. Lander,
      •  
    • Gad Getz &
      •  
    • Todd R. Golub

  2. Mayo Clinic Arizona, 13400 East Shea Boulevard, Scottsdale, Arizona 85259, USA

    • Jonathan J. Keats,
      •  
    • Gregory J. Ahmann,
      •  
    • P. Leif Bergsagel,
      •  
    • Rafael Fonseca &
      •  
    • A. Keith Stewart

  3. The Multiple Myeloma Research Consortium, 383 Main Avenue, 5th Floor, Norwalk, Connecticut 06581, USA

    • Jonathan J. Keats,
      •  
    • Gregory J. Ahmann,
      •  
    • Kenneth C. Anderson,
      •  
    • Daniel Auclair,
      •  
    • P. Leif Bergsagel,
      •  
    • Rafael Fonseca,
      •  
    • Craig C. Hofmeister,
      •  
    • Sundar Jagannath,
      •  
    • Andrzej J. Jakubowiak,
      •  
    • Amrita Krishnan,
      •  
    • Joan Levy,
    • Sagar Lonial,
      •  
    • Bunmi Mfuko,
      •  
    • Louise M. Perkins,
      •  
    • S. Vincent Rajkumar,
      •  
    • David S. Siegel,
      •  
    • A. Keith Stewart,
      •  
    • Suzanne Trudel,
      •  
    • Ravi Vij &
      •  
    • Todd Zimmerman

  4. Dana-Farber Cancer Institute, 44 Binney Street, Boston, Massachusetts 02115, USA

    • Anna C. Schinzel,
      •  
    • Kenneth C. Anderson,
      •  
    • William C. Hahn,
      •  
    • Levi A. Garraway,
      •  
    • Matthew Meyerson &
    • Todd R. Golub

  5. Massachusetts General Hospital, 55 Fruit Street, Boston, Massachusetts 02114, USA

    • Mazhar Adli &
      •  
    • Bradley E. Bernstein

  6. Multiple Myeloma Research Foundation, 383 Main Avenue, 5th Floor, Norwalk, Connecticut 06581, USA

    • Daniel Auclair,
      •  
    • Joan Levy,
      •  
    • Bunmi Mfuko &
      •  
    • Louise M. Perkins

  7. The Translational Genomics Research Institute, 445 North Fifth Street, Phoenix, Arizona 85004, USA

    • Angela Baker,
      •  
    • John Carpten &
      •  
    • Jeff Trent

  8. Harvard Medical School, Boston, Massachusetts 02115, USA

    • Bradley E. Bernstein,
      •  
    • William C. Hahn,
      •  
    • Matthew Meyerson,
      •  
    • Eric S. Lander &
      •  
    • Todd R. Golub

  9. Howard Hughes Medical Institute, Chevy Chase, Maryland 20815, USA

    • Bradley E. Bernstein &
      •  
    • Todd R. Golub

  10. Department of Physics of Complex Systems, Weizmann Institute of Science, Rehovot 76100, Israel

    • Yotam Drier

  11. The Ohio State University Medical Center, 320 West 10th Avenue, Columbus, Ohio 43210, USA

    • Craig C. Hofmeister

Postado por às

Nenhum comentário :

Postar um comentário

Assinar: Postar comentários ( Atom )