Abstract
Aims: Assessing whether Next Generation DNA Sequencing (NGS) can be used to screen prostate cancer for multiple gene alterations in men routinely diagnosed with this disease and/or who are entered into clinical trials. Previous studies are limited and have reported only low success rates.
Methods: We marked areas of cancer on H&E stained sections from formalin fixed-needle biopsys, and used these as templates to dissect cancer rich tissue from adjacent unstained sections. DNA was prepared using a Qiagen protocol modified to maximise DNA yield. The DNA was screened simultaneously for mutations in 365 cancer-related genes using an Illumina HiSeq 2000 NGS platform.
Results: From 63 prostate cancers examined (59/63, 94%) of the samples yielded at least 30ng of DNA, the minimum amount of DNA considered suitable for NGS analysis. Patients in the D’Amico high-risk group yielded an average of 1033ng; intermediate-risk patients 401ng; and low risk patients 97ng. NGS of 8 samples selected from high and intermediate risk groups gave a median exon read depth of 962 and detected TMPRRS2-ERG fusions, as well as a variety of mutations including those in the SPOP, TP53, ATM, MEN1, NBPF10, NCOR2, PIK3CB, and MAP2K5 (MEK5) genes.
Conclusions: Using the methods presented here NGS technologies can be used to screen a high proportion of prostate cancer patients for mutations in cancer-related genes in tissue samples opening up its general use in the context of clinical trials or routine diagnosis.
Methods: We marked areas of cancer on H&E stained sections from formalin fixed-needle biopsys, and used these as templates to dissect cancer rich tissue from adjacent unstained sections. DNA was prepared using a Qiagen protocol modified to maximise DNA yield. The DNA was screened simultaneously for mutations in 365 cancer-related genes using an Illumina HiSeq 2000 NGS platform.
Results: From 63 prostate cancers examined (59/63, 94%) of the samples yielded at least 30ng of DNA, the minimum amount of DNA considered suitable for NGS analysis. Patients in the D’Amico high-risk group yielded an average of 1033ng; intermediate-risk patients 401ng; and low risk patients 97ng. NGS of 8 samples selected from high and intermediate risk groups gave a median exon read depth of 962 and detected TMPRRS2-ERG fusions, as well as a variety of mutations including those in the SPOP, TP53, ATM, MEN1, NBPF10, NCOR2, PIK3CB, and MAP2K5 (MEK5) genes.
Conclusions: Using the methods presented here NGS technologies can be used to screen a high proportion of prostate cancer patients for mutations in cancer-related genes in tissue samples opening up its general use in the context of clinical trials or routine diagnosis.
Original language | English |
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Pages (from-to) | 212-217 |
Number of pages | 6 |
Journal | Journal of Clinical Pathology |
Volume | 68 |
Early online date | 13 Jan 2015 |
DOIs | |
Publication status | Published - 2015 |
Profiles
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Daniel Brewer
- Norwich Medical School - Professor in Medical Bioinformatics and Genomics
- Metabolic Health - Member
- Cancer Studies - Member
Person: Research Group Member, Research Centre Member, Academic, Teaching & Research
-
Colin Cooper
- Norwich Medical School - Professor of Cancer Genetics
- Cancer Studies - Member
Person: Research Group Member, Academic, Teaching & Research
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Marcus Flather
- Norwich Medical School - Emeritus Professor
- Cardiovascular and Metabolic Health - Member
Person: Honorary, Research Group Member