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Advances in understanding cancer genomes through second-generation sequencing

Key Points

  • Analyses of cancer genome sequences and structures provide insights for understanding cancer biology, diagnosis and therapy.

  • The application of second-generation DNA sequencing technologies (also known as next-generation sequencing) is allowing substantial advances in cancer genomics. In recent years, it has become feasible to sequence the expressed genes ('transcriptomes'), known exons ('exomes'), and complete genomes of cancer samples.

  • There are particular challenges for the detection and diagnosis of cancer genome alterations. For example, some cancer genome alterations are prevalent at low frequency in clinical samples, often owing to substantial admixture with non-malignant cells.

  • The large quantity of data from second-generation sequencing provides statistical and computational challenges.

  • An impetus for studies of somatic genome alterations is the potential for therapies targeted against the products of these alterations.

Abstract

Cancers are caused by the accumulation of genomic alterations. Therefore, analyses of cancer genome sequences and structures provide insights for understanding cancer biology, diagnosis and therapy. The application of second-generation DNA sequencing technologies (also known as next-generation sequencing) — through whole-genome, whole-exome and whole-transcriptome approaches — is allowing substantial advances in cancer genomics. These methods are facilitating an increase in the efficiency and resolution of detection of each of the principal types of somatic cancer genome alterations, including nucleotide substitutions, small insertions and deletions, copy number alterations, chromosomal rearrangements and microbial infections. This Review focuses on the methodological considerations for characterizing somatic genome alterations in cancer and the future prospects for these approaches.

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Figure 1: Depth of coverage and physical coverage.
Figure 2: Sequence capture for cancer genomics.
Figure 3: Types of genome alterations that can be detected by second-generation sequencing.

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Acknowledgements

We thank M. Lawrence and G. Saksena for careful review of the manuscript. We acknowledge support from The Cancer Genome Atlas programme of the National Cancer Institute, U24CA143867 and U24CA143845, and from the National Human Genome Research Institute, U54HG003067.

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Correspondence to Matthew Meyerson.

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Matthew Meyerson receives research support from Genentech, is a consultant to and receives research support from Novartis, and is a founding advisor of and a consultant to Foundation Medicine.

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Glossary

Second-generation sequencing

Used in this Review to refer to sequencing methods that have emerged since 2005 that parallelize the sequencing process and produce millions of typically short sequence reads (50–400 bases) from amplified DNA clones. It is also often known as next-generation sequencing.

First-generation sequencing

(also known as Sanger sequencing or capillary sequencing). The standard sequencing methodology used to sequence the reference human (and other model organism) genomes. It uses radioactively or fluorescently labelled dideoxynucleotide triphosphates (ddNTPs) as DNA chain terminators. Various detection methods allow read-out of sequence according to the incorporation of each specific terminator (ddATP, ddCTP, ddGTP or ddTTP).

Whole-genome amplification

Various molecular techniques (including multiple displacement amplification, rolling circle amplification or degenerate oligonucleotide primed PCR) in which very small amounts (nanograms) of a genomic DNA sample can be multiplied in a largely unbiased fashion to produce suitable quantities for genomic analysis (micrograms).

Moore's law

The observation made in 1965 by Gordon Moore that the number of transistors per square inch on integrated circuits had doubled every other year since the integrated circuit was invented.

Chromatin immunoprecipitation

A technique used to identify the location of DNA-binding proteins and epigenetic marks in the genome. Genomic sequences containing the protein of interest are enriched by binding soluble DNA chromatin extracts (complexes of DNA and protein) to an antibody that recognizes the protein or modification.

Over-sampling

Reading the same stretch of DNA sequence many times to gain a confident sequence read-out.

Shotgun sequencing

Sequencing randomly derived fragments of the whole genome. The order and orientation of the sequences are determined by mapping individual reads back to a reference or through assembly of overlapping sequences into larger contigs of sequence.

Jumping library

A method of library construction in which the genome is divided into large fragments using a rare cutter enzyme. Fragments are circularized and DNA sequences are read from the ends of the fragment, without reading the intervening sequence.

Transformation assay

The measurement of cell phenotypes to assess oncogenic changes.

Digital karyotyping

A method to quantify DNA copy number. Short sequence-derived tags that cover the genome are used to read-out relative copy number.

Directed sequencing

Sequencing only subsets of the genome, for example, particular genes or regions of interest.

Free serum DNA

DNA that is cell-free and is circulating in the bloodstream. It typically refers to tumour DNA that can be isolated in the blood.

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Meyerson, M., Gabriel, S. & Getz, G. Advances in understanding cancer genomes through second-generation sequencing. Nat Rev Genet 11, 685–696 (2010). https://doi.org/10.1038/nrg2841

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