Intraoperational Prostate Loge Biopsies (iPROLOGX) After Radical Prostatovesiculectomy (iPROLOGX)

Intraoperational Prostate Loge Biopsies (iPROLOGX) After Radical Prostatovesiculectomy (RPVE) in Prostate Cancer (PCA) Patients for Molecular Tumor Marker Analysis

This project is about the detection of occult tumor cells in surgical margins of radical prostatovesiculectomy by analysing the methylation status of Glutathione S-transferase P 1 (GSTP1). After gland excision specimens are obtained from 9 defined areas of the prostatic fossa. The biopsies are divided into two parts. One part used for histopathological analysis and the other part for moleculargenetic analysis. Results will be correlated e.g. with tumor stage, Gleason Score and prostate specific antigen (PSA). The prostate-cancer-negative control group with bladder cancer.

This project is about the detection of occult tumor cells in surgical margins of radical prostatovesiculectomy by analysing the methylation status of Glutathione S-transferase P 1 (GSTP1). After gland excision specimens are obtained from 9 defined areas of the prostatic fossa. The biopsies are divided into two parts. One part used for histopathological analysis and the other part for moleculargenetic analysis. Results will be correlated e.g. with tumor stage, Gleason Score and prostate specific antigen (PSA). The prostate-cancer-negative control group with bladder cancer. DNA ISOLATION DNA from biopsies stored by -80°C was isolated by using innuPREP DNA mini Kit (Analytik Jena, Jena, Germany) following protocol 1 of the manufacturer's instructions. DNA was eluted with 50 µl elution buffer. Concentration and purity were analysed by using Nanodrop 2000. DNA BISULFITE MODIFICATION DNA was modified by using EpiTect Bisulfite Kit (QIAGEN, Hilden, Germany) according to manufacturer's instructions. Samples were eluted once with 20 µl elution buffer. QUANTITATIVE METHYLATION SPECIFIC PCR Methylation status of GSTP1 is analysed by quantitative methylation-specific PCR (Q-MSP) using StepOnePlus Real-Time PCR System and StepOne Software v2.1 from Applied Biosystems (Darmstadt, Germany). Q-MSP was performed in duplicate analysing genes Actin and GSTP1. The primers' and testing probes' sequences used to amplify and detect hypermethylated GSTP1 were: 5'-AgTTgCgCggCgATTTC (forward primer), 5'-gCCCCAATACTAAATCACgACg (reverse primer) and 5'-CggTCgACgTTCggggTgTAgCg (taqman probe), labelled with fluorescence dye FAM. The primers' and testing probes' sequences used to amplify and detect Actin were: 5'-TggTgATggAggAggTTTAgTAAgT (forward primer), 5'-AACCAATAAAACCTACTCCTCCCTTAA (reverse primer),5'-ACCACCACCCAACACACAATAACAAACACA (taqman probe), labelled with fluorescence dye VIC. The Q-MSP was carried out at 50°C for 2 min., 95°C for 15 min. followed by 50 cycles of 95°C for 1s and 60°C for 1 min. As a positive control bisulfite-converted DNA of DU145 and LNCap were used. Blank reactions with destillated water, which replaced DNA, served as negative control (NTC).

Prostate cancer conducted for RPVE with curative Intention, biopsies of the prostatic fossa in Magdeburg

Conducted for CE in Male with bladder cancer or other indication for cystectomy but without prostate cancer in Magdeburg or Gronau, biopsies of the prostatic fossa in Gronau

intraoperative Open surgical biopsies of the prostatic fossa after prostatevesiculectomy in Magdeburg

intraoperative endoscopic robotassisted biopsies of the prostatic fossa after prostatevesiculectomy in Gronau

Criteria for patients with prostate adenocarcinoma: Inclusion Criteria sex: male diagnosis: prostate adenocarcinoma treatment: radical prostatovesiculectomy period of treatment: 11/30/2011 - 10/15/2013 Exclusion Criteria sex: female diagnosis: no prostate adenocarcinoma treatment: no radical prostatovesiculectomy period of treament: before 11/30/2011 or after 10/15/2013 Criteria for prostate adenocarcinoma negative control group: Inclusion Criteria sex: male diagnosis: urothelial carcinoma treatment: cysto-prostatectomy period of treament: 12/14/2011 - 02/18/2014 Exclusion Criteria sex: female diagnosis: incidental prostate adenocarcinoma treatment: no cysto-prostatectomy period of treatment: before 12/14/2011 or after 02/18/2014