BMC Cancer 2005, 5:111 http://www.biomedcentral.com/1471-2407/5/111 ibFAnyeintgneaRuirlgyTnrns-aeiPslpC3crooRfnsgttuearsntoienclgiednxβiswp-earhacesotseislnei(oyBbniPleolHedov)ededlfsrao7mf4p01570b3aosSfe,2-pp05a0pir4aSbtiaeannddtsapws5i1t0hS Analysis of gene expression levels of p503S, p504S and p510S by RT-PCR using β-actin yielded a 740 base-pair band as internal control in whole blood from 17 of 20 patients with benign prostatic disease (BPH). i(1pbFAnlaly6aintadgn)esaRr;uertleyPTinrcn1sCa-eai13P(ls3l)C4aco;gno(RlfuLlaenagStnsntue1dre2srn4oi,(en71ll4gaeD7ie,n)xβca;e(fpen-lraar7doncnev)dzt5;iesxiesn)hLn1ie(opNy4labrni)Creone;tlsneaSdt(ivPCeag1leadtn2O(nlilsc)ea(Vlonasta7inefns384dspe10u(t5s8hloea0b)1u;n3a1,mnesS03oe,a)1,-np;p5a5mpna)c0;iredyr4MololS6bsmClt)aiaannFenated7ntidcsrd:ai(putlsSnia5msVne1s4eou08eS- Analysis of gene expression levels of p503S, p504S and p510S by RT-PCR using β-actin yielded a 740 base-pair band as internal control in frozen benign (lanes 1, 3, and 6) and neo- plastic (lanes 2, 4, and 5) prostatic tissue; non prostatic tissue (adrenal gland (lane 7); heart (lanes 8 to 10); myometrium (lane 11); uterine cervix (lane 12) and human cell lines: SV48 (lane 13); LS147D (lane 14); SCOV 3 (lane 15); MCF7 (lane 16); PC3 (lane 17); and LNCaP (lane 18). Negative control no RNA (lane 19); MW (molecular weight). Discussion These findings argue against the usefulness of the p503S, p504S and p510S genes as diagnostic and probably against their usefulness as prognostic prostate-specific markers for monitoring disease progression and recur- rence in PB from patients with CaP. Our findings obtained by RT-PCR assay contrast with Zelie et al [11] et al who found the RT-PCR-AMACR test non-invasive and useful in predicting the presence of CaP in prostatic secretions. When we compared our findings for p503S, p504S and p510S with our earlier study investigating prostatic spe- cific antigen (PSA) [3], as potential markers for prostate tumors, all these three genes were more frequently expressed than PSA in PB from patients with treated CaP (63%, 63% and 100% vs 18%) and untreated CaP (18%, 81% and 100% vs 9%) and in PB from BPH (25%, 55% and 75% vs 0). In benign and malignant prostate tissue, p510S was more strongly expressed than PSA, or p503S and p504S (100% vs 66% vs 66%), whereas in prostatic cell lines, PSA and p503S, p504S and p510S were simi- larly expressed (100%). In non prostatic tissue and cell lines, PSA was not expressed; in contrast, in non prostatic samples and cell lines, p503S was more expressed than p510S and p504S (100% vs 33% vs 16%) and cell lines (100% vs 50% vs 50%). Hence, the increased expression of PSA mRNA in prostate tissue suggests that PSA is more specific than p503S, p504S and p510S as a marker of can- cer progression and dissemination. The ubiquitous presence of p503S, p504S and p510S in PB of patients with BPH, in non prostatic tissue and cell lines therefore argues against their potential value as biomarkers of prostate disease. Compared with PSA [13], none of the three newly-identified putative CaP markers we studied were sufficiently sensitive or specific to distin- guish patients in complete clinical remission from patients with occult metastatic disease at high-risk for relapse. The numerous false-positive findings in our assay and the reported presence of p503S, p504S and p510S in normal hematopoietic tissues also from patients without prostate malignancies [20], could depend on illegitimate expression of gene transcripts. This question awaits an answer from studies designed to investigate the whole p503S, p504S and p510S gene sequence. For RT-PCR analysis of PB cells in this study we used primers for three new putative genes (p503S, p504S, and p510S) whose prostate cancer specificity has already been confirmed by Northern blot, real-time PCR and immunohistochemical assay on prostatic tissue [3-8]. Although AMACR may be potentially useful as a tissue biomarker for prostate cancer [3,4], the limitations of this assay make it impossible to use an autoimmune response against AMACR as a means of detecting this antigen in the serum [10]. Nor did a novel approach to predict the presence of CaP from pros- tatic secretions prove predictive of CaP [9]. Whether these new markers have potential clinical utility as sensitive and specific indicators of CaP progression therefore warrants further research using quantitative RT-PCR technique to study the entire gene sequences. Conclusion We provide the first evidence of the ubiquitous presence of p503S, p504S and p510S in PB of patients with BPH, in non prostatic tissue and cell lines. These findings suggest that the investigated genes are poorly specific and proba- bly of little use as diagnostic or prognostic prostatic markers in peripheral blood for monitoring disease pro- gression and recurrence. Page 5 of 7 (page number not for citation purposes)