A tissue biopsy-based epigenetic multiplex PCR assay for prostate cancer detection

Leander Van Neste, Joseph Bigley, Adam Toll, Gaëtan Otto, James Clark, Paul Delrée, Wim Van Criekinge, Jonathan Ira Epstein

Research output: Contribution to journalArticle

Abstract

Background: PSA-directed prostate cancer screening leads to a high rate of false positive identifications and an unnecessary biopsy burden. Epigenetic biomarkers have proven useful, exhibiting frequent and abundant inactivation of tumor suppressor genes through such mechanisms. An epigenetic, multiplex PCR test for prostate cancer diagnosis could provide physicians with better tools to help their patients. Biomarkers like GSTP1, APC and RASSF1 have demonstrated involvement with prostate cancer, with the latter two genes playing prominent roles in the field effect. The epigenetic states of these genes can be used to assess the likelihood of cancer presence or absence. Results: An initial test cohort of 30 prostate cancer-positive samples and 12 cancer-negative samples was used as basis for the development and optimization of an epigenetic multiplex assay based on the GSTP1, APC and RASSF1 genes, using methylation specific PCR (MSP). The effect of prostate needle core biopsy sample volume and age of formalin-fixed paraffin-embedded (FFPE) samples was evaluated on an independent follow-up cohort of 51 cancerpositive patients. Multiplexing affects copy number calculations in a consistent way per assay. Methylation ratios are therefore altered compared to the respective singleplex assays, but the correlation with patient outcome remains equivalent. In addition, tissue-biopsy samples as small as 20 μm can be used to detect methylation in a reliable manner. The age of FFPE-samples does have a negative impact on DNA quality and quantity. Conclusions: The developed multiplex assay appears functionally similar to individual singleplex assays, with the benefit of lower tissue requirements, lower cost and decreased signal variation. This assay can be applied to small biopsy specimens, down to 20 microns, widening clinical applicability. Increasing the sample volume can compensate the loss of DNA quality and quantity in older samples.

Original languageEnglish (US)
Article number16
JournalBMC Urology
Volume12
DOIs
StatePublished - 2012

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Multiplex Polymerase Chain Reaction
Epigenomics
Prostatic Neoplasms
Methylation
Biopsy
Paraffin
Formaldehyde
Biomarkers
APC Genes
Large-Core Needle Biopsy
DNA
Tumor Suppressor Genes
Early Detection of Cancer
Genes
Prostate
Neoplasms
Physicians
Costs and Cost Analysis
Polymerase Chain Reaction

Keywords

  • APC
  • Diagnosis
  • Epigenetics
  • GSTP1
  • Methylation
  • MSP
  • Multiplex
  • Prostate cancer
  • RASSF1
  • Singleplex

ASJC Scopus subject areas

  • Urology
  • Reproductive Medicine

Cite this

A tissue biopsy-based epigenetic multiplex PCR assay for prostate cancer detection. / Van Neste, Leander; Bigley, Joseph; Toll, Adam; Otto, Gaëtan; Clark, James; Delrée, Paul; Van Criekinge, Wim; Epstein, Jonathan Ira.

In: BMC Urology, Vol. 12, 16, 2012.

Research output: Contribution to journalArticle

Van Neste, L, Bigley, J, Toll, A, Otto, G, Clark, J, Delrée, P, Van Criekinge, W & Epstein, JI 2012, 'A tissue biopsy-based epigenetic multiplex PCR assay for prostate cancer detection', BMC Urology, vol. 12, 16. https://doi.org/10.1186/1471-2490-12-16
Van Neste, Leander ; Bigley, Joseph ; Toll, Adam ; Otto, Gaëtan ; Clark, James ; Delrée, Paul ; Van Criekinge, Wim ; Epstein, Jonathan Ira. / A tissue biopsy-based epigenetic multiplex PCR assay for prostate cancer detection. In: BMC Urology. 2012 ; Vol. 12.
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AU - Delrée, Paul

AU - Van Criekinge, Wim

AU - Epstein, Jonathan Ira

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AB - Background: PSA-directed prostate cancer screening leads to a high rate of false positive identifications and an unnecessary biopsy burden. Epigenetic biomarkers have proven useful, exhibiting frequent and abundant inactivation of tumor suppressor genes through such mechanisms. An epigenetic, multiplex PCR test for prostate cancer diagnosis could provide physicians with better tools to help their patients. Biomarkers like GSTP1, APC and RASSF1 have demonstrated involvement with prostate cancer, with the latter two genes playing prominent roles in the field effect. The epigenetic states of these genes can be used to assess the likelihood of cancer presence or absence. Results: An initial test cohort of 30 prostate cancer-positive samples and 12 cancer-negative samples was used as basis for the development and optimization of an epigenetic multiplex assay based on the GSTP1, APC and RASSF1 genes, using methylation specific PCR (MSP). The effect of prostate needle core biopsy sample volume and age of formalin-fixed paraffin-embedded (FFPE) samples was evaluated on an independent follow-up cohort of 51 cancerpositive patients. Multiplexing affects copy number calculations in a consistent way per assay. Methylation ratios are therefore altered compared to the respective singleplex assays, but the correlation with patient outcome remains equivalent. In addition, tissue-biopsy samples as small as 20 μm can be used to detect methylation in a reliable manner. The age of FFPE-samples does have a negative impact on DNA quality and quantity. Conclusions: The developed multiplex assay appears functionally similar to individual singleplex assays, with the benefit of lower tissue requirements, lower cost and decreased signal variation. This assay can be applied to small biopsy specimens, down to 20 microns, widening clinical applicability. Increasing the sample volume can compensate the loss of DNA quality and quantity in older samples.

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