Single-cell mutational profiling and clonal phylogeny in cancer.
The development of cancer is a dynamic evolutionary process in which intraclonal, genetic diversity provides a substrate for clonal selection and a source of therapeutic escape. The complexity and topography of intraclonal genetic architectures have major implications for biopsy-based prognosis and for targeted therapy. High-depth, next-generation sequencing (NGS) efficiently captures the mutational load of individual tumors or biopsies. But, being a snapshot portrait of total DNA, it disguises the fundamental features of subclonal variegation of genetic lesions and of clonal phylogeny. Single-cell genetic profiling provides a potential resolution to this problem, but methods developed to date all have limitations. We present a novel solution to this challenge using leukemic cells with known mutational spectra as a tractable model. DNA from flow-sorted single cells is screened using multiplex targeted Q-PCR within a microfluidic platform allowing unbiased single-cell selection, high-throughput, and comprehensive analysis for all main varieties of genetic abnormalities: chimeric gene fusions, copy number alterations, and single-nucleotide variants. We show, in this proof-of-principle study, that the method has a low error rate and can provide detailed subclonal genetic architectures and phylogenies.
The Institute of Cancer Research, London, SM2 5NG, United Kingdom;
Nicola E
NE
Potter
Luca
L
Ermini
Elli
E
Papaemmanuil
Giovanni
G
Cazzaniga
Gowri
G
Vijayaraghavan
Ian
I
Titley
Anthony
A
Ford
Peter
P
Campbell
Lyndal
L
Kearney
Mel
M
Greaves
D045744:Cell Line, Tumor; D060965:Clonal Evolution; D056915:DNA Copy Number Variations; D014644:Genetic Variation; D023281:Genomics / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction; D009154:Mutation; D009369:Neoplasms / Q000235:genetics*; D010802:Phylogeny; D020641:Polymorphism, Single Nucleotide; D059010:Single-Cell Analysis