Detection of prostate-specific antigen RNA before and after radical retropubic prostatectomy and transurethral resection of the prostate using "Light-Cycler"-based quantitative real-time polymerase chain reaction.
OBJECTIVE
To report our initial experience gained in establishing real-time reverse transcriptase-polymerase chain reaction (RT-PCR) detection of prostate-specific antigen (PSA) mRNA using the quantitative online PCR system LightCycler. Many studies have thus far failed to provide the desired proof that the detection of circulating PSA-expressing tumor cells with RT-PCR in the blood samples of patients with prostate cancer (PCa) is a highly sensitive prognostic and course marker. One of the possible reasons is the lack of reliable quantification methods.


METHODS
Blood samples before and after surgery were obtained from 87 patients who underwent radical prostatectomy for locally confined PCa and 27 patients who underwent transurethral resection of the prostate for benign prostatic hyperplasia. Eight days postoperatively, additional blood samples were obtained from the patients with PCa. Quantitative no-nested RT-PCR for PSA mRNA (291 bp) was performed using the LightCycler system applying the SYBR Green protocol. The number of circulating LNCaP tumor cell-equivalents per sample was estimated from the mean amplification value measured in a given number of LNCaP cells.


RESULTS
PSA mRNA was detected preoperatively in 19 patients with Stage pT2 tumor (40%) and in 28 patients with tumor greater than Stage pT2 (72%), but in only 2 patients with benign prostatic hyperplasia (8%; analysis of variance, P <0.001). Significant quantitative differences were observed among Stage pT2 disease (1034 LNCaP tumor cell-equivalents/mL), greater than Stage pT2 disease (7830 cells/mL), and benign prostatic hyperplasia (58 cells/mL; analysis of variance for all groups, P <0.001). The correlation between the detection of PSA expression by RT-PCR and the Gleason score and serum PSA value was statistically significant.


CONCLUSIONS
Our results show that the initial experience with the LightCycler system for PSA-assisted detection of circulating PSA mRNA in PCa by RT-PCR may be a promising preoperative prognostic marker for organ-confined or locally advanced PCa. Long-term follow-up of these patients with PCa must demonstrate the clinical value of molecular diagnostics with quantitative RT-PCR systems.
Straub
B
B
Department of Urology, Universitätsklinikum Benjamin Franklin, Freie Universität Berlin, Berlin, Germany.
Müller
M
M
Krause
H
H
Schrader
M
M
Goessl
C
C
Heicappell
R
R
Miller
K
K
D000704:Analysis of Variance; D006801:Humans; D008297:Male; D008875:Middle Aged; D009367:Neoplasm Staging; D017430:Prostate-Specific Antigen / Q000097:blood*; D011468:Prostatectomy / Q000379:methods; D011470:Prostatic Hyperplasia / Q000097:blood* / Q000473:pathology / Q000601:surgery; D011471:Prostatic Neoplasms / Q000097:blood* / Q000473:pathology / Q000601:surgery; D012333:RNA, Messenger / Q000097:blood*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D017430:Prostate-Specific Antigen