Decoding the fine-scale structure of a breast cancer genome and transcriptome

Abstract

A comprehensive understanding of cancer is predicated upon knowledge of the structure of malignant genomes underlying its many variant forms and the molecular mechanisms giving rise to them. It is well established that solid tumor genomes accumulate a large number of genome rearrangements during tumorigenesis. End Sequence Profiling (ESP) maps and clones genome breakpoints associated with all types of genome rearrangements elucidating the structural organization of tumor genomes. Here we extend the ESP methodology in several directions using the breast cancer cell line MCF-7. First, targeted ESP is applied to multiple amplified loci, revealing a complex process of rearrangement and coamplification in these regions reminiscent of breakage/fusion/bridge cycles. Second, genome breakpoints identified by ESP are confirmed using a combination of DNA sequencing and PCR. Third, in vitro functional studies assign biological function to a rearranged tumor BAC clone, demonstrating that it encodes antiapoptotic activity. Finally, ESP is extended to the transcriptome identifying four novel fusion transcripts and providing evidence that expression of fusion genes may be common in tumors. These results demonstrate the distinct advantages of ESP including: (1) the ability to detect all types of rearrangements and copy number changes; (2) straightforward integration of ESP data with the annotated genome sequence; (3) immortalization of the genome; (4) ability to generate tumor-specific reagents for in vitro and in vivo functional studies. Given these properties, ESP could play an important role in a tumor genome project.

Assessing human germ‐cell mutagenesis in the Postgenome Era: A celebration of the legacy of William Lawson (Bill) Russell

“…Gray presented a list of technologies for assessing DNA and genomic defects, and related them in a matrix to the biological endpoints that could potentially be measured or analyzed with those technologies, and their detection capabilities (Table II). The technologies evaluated included PCR-based sequencing, PCR-based conformation analysis, sequencing by hybridization, ESP [Volik et al, 2003[Volik et al, ,2006, high-throughput sequencing, primer extension, FISH, CGH, high-throughput LOH, optical mapping, genome subtraction, expression arrays [Wang et al, 2005], Serial Analysis Gene Expression (SAGE, both RNA, and DNA), chromatin immunoprecipitation assays, protein lysate arrays, 1D and 2D gel electrophoresis, mass spectrometry, and computational biology. Many of the advantages and limitations of these technologies were presented by earlier speakers.…”

“…Colin Collins (UC, San Francisco) noted that ESP is a relatively new technology that can clone and map many kinds of chromosomal defects, e.g., deletions, insertions, translocations, inversions, and copy number changes, in a single step [Volik et al, 2006]. The resolution of ESP is relatively high (10 kb), and this powerful technology could be useful for identifying and characterizing environmentally induced germ-cell mutations.…”

Prognosis and Gene Expression Profiling of 20q13-Amplified Breast Cancers

“…Further analyses using tiling-path array comparative genomic hybridization on many cases should determine whether distinct 20q13 amplicons can be recurrently identified in type 2 tumors. The region may also be ''packaged'' in a single amplicon as to contain only amplified and overexpressed selected genes (43,44). The participation of Aurora A in type 2 amplification is highly suspected.…”

High Fidelity Patient-Derived Xenografts for Accelerating Prostate Cancer Discovery and Drug Development

“…Chromosomal aberration is a sentinel feature of many cancers, and the associated gene deregulation and genome instability is implicated in the development and progression of prostate cancer (5,6,(28)(29)(30). Combined genome copy-number analysis of each independent transplantable tumor line Table 1.…”