The clinical utility of the prostate specific membrane antigen reverse-transcription/polymerase chain reaction to detect circulating prostate cells: an analysis in healthy men and women

Abstract

Objective: To evaluate the overall specificity of nested reverse transcriptase-polymerase chain reaction (RT-PCR) to detect prostate-specific membrane antigen (PSM) mRNA in peripheral blood samples of healthy donors.

Subjects and methods: Peripheral blood samples were taken from 60 healthy blood-donors (30 men and 30 women aged < 50 years) and analysed for PSM-mRNA using nested RT-PCR (in 'hot-start' conditions and confirmed using nested EcoRI restriction enzyme). Intron-spanning primer pairs specific for human PSM were deduced from the GenBank sequence (M99487) using gene software. The outer primer pair for PSM was: fwd: 1368 5'-TCACCGGGACTCATGGGTGT-3'; reverse: 1860 5'-GCCTGAAGCAATTCCAAGTCGG-3'. Inner primer pair for PSM was: fwd: 1480 5'-AAGGAAGGGTGGAGACCTAG-3'; reverse: 5-ACTGAACTCTGGGGAAGGAC-3'. The integrity of cDNAs was checked using primer pairs specific for the housekeeping gene beta-actin. The specificity and false-positive rate were calculated assuming that the underlying prostate cancer incidence was nil.

Results: The first PCR was negative for all samples (100% specificity; 0% false-positive rate). The nested PCR detected 23 positive samples (23/60, 38%) with an overall specificity of 62% (false positive rate, 38%).

Conclusion: Nested RT-PCR of PSM-mRNA in peripheral blood is highly unspecific. Its clinical utility in the management of prostate cancer must be low. Further development is needed of quantitative RT-PCR, primers that identify prostatic PSM or another prostate-specific marker gene to differentiate PSM mRNA from circulating prostate cells and from non-prostatic tissues.