Genome-Wide Massively Parallel Sequencing Using SOLiD™ 4 of Formalin Fixed Paraffin Embedded Prostate Cancer Tissue.
Pavel Nikolov, Roopika Menon, Martin Brown, Veit Scheble, Falko Fend, Detlef Boehm, Saskia Biskup, Sven Perner. University Hospital Bonn, Germany; University Hospital Tuebingen, Germany; CeGaT GmbH, Tuebingen, Germany; University of Tuebingen Medical Center II, Germany
Background: Alternation in the exome is considered to be one of the instigators of cancer. The exome forms 1.5% of the human genome, consisting of the genetic blueprint for protein synthesis and other functional gene products. SOLiD-Sequencing by Oligonucleotide Ligation and Detection (Life Technologies) has an accuracy of 99.94%. This robust technique has proven to be helpful in detecting novel genes associated with prostate cancer genomic and chromosomal rearrangements, copy number variation, and nucleotide substitutions. The aim of this study was to compare whole exome sequencing of paired frozen/formalin fixed paraffin embedded (FFPE) tumor tissue with non-tumor FFPE tissue obtained from the same patient in order to detect genes involved in cancer progression.
Design: So far, we investigated one prostate sample. This sample was divided into FFPE tumor, fresh frozen tumor and FFPE non-tumor material, all derived from the same patient. DNA was isolated and purified from all three samples. These samples were then processed for library construction to carry out deep sequencing on the SOLiD 4 system.
Results: The high mapping stringency approach generated reliable values for the unique placed reads. Our results show highly similar ontarget reads within +/- 150 bp for each sample, including 81.91% for the FFPE non tumor, 78.47% for FFPE tumor and 82.44% for frozen tumor. We detectably captured 98.19% of targeted exons for FFPE non-tumor, compared to the 98.32% for FFPE tumor and 98.52% for the frozen tumor samples. Using QV-filtering, we mapped 20560 SNPs to regions of interest, for each dataset. By comparing the SNP profiles between tumor and non- tumor sets, 1297 SNPs were found to be highly specific for the tumor samples. Out of these, we could identify 341 non-synonymous SNPs and 289 synonymous SNPs that will be further analyzed.
Conclusions: This is the first study showing comparable exome sequencing results between the FFPE and corresponding frozen cancer tissues using SOLiD 4. Based on the promising results, the study will be continued on a much larger cohort exhibiting both localized and metastatized prostate cancer. The usage of FFPE blocks will open up the Pathology archives for exome-related studies that provide insight into the PCa genome, resulting in novel biomarkers and therapeutic targets discovery.
Category: Special Category - Pan-genomic/Pan-proteomic approaches to Cancer
Monday, February 28, 2011 1:00 PM
Poster Session II # 239, Monday Afternoon