Diagnostic Efficacy of PSMA and PSCA mRNAs Combined to PSA in Prostate Cancer Patients

Background: Serum Prostate-specific antigen (PSA) has been used for screening and diagnosis of prostate cancer (PCa) but it is burdened by its low accuracy, creating a need for reliable diagnostic markers. Despite prostate-specific membrane antigen (PSMA) and prostate stem cell antigen (PSCA) being widely expressed in the tissue of PCa, no definite conclusion regarding their use as clinical biomarkers due to their lacking organ specificity. Therefore, this study aimed to evaluate the peripheral blood levels of PSMA and PSCA mRNAs and examine their diagnostic significance as non-invasive integrated markers.Materials and Methods: 125 subjects were enrolled in this study. They were divided into 25 healthy controls, 25 BPH patients, and 75 PCa patients. The expression levels of PSMA and PSCA were determined using quantitative RT- PCR, in addition to measuring serum PSA.Results: Levels of PSMA and PSCA were over-expressed in PCa patients compared to controls and BPH patients and were found to be associated with increased susceptibility to PCa. Moreover, the diagnostic values of PSMA and PSCA to distinguish PCa patients from BPH patients and controls were inferior to that of PSA. However, the combination of PSMA and PSCA with PSA enhanced the efficacy of the latter.Conclusion: This study suggests that these genes were associated with malignant susceptibility. Concerning the duality of PSMA-PSA or PSCA-PSA, this implies the significance of their investigation together in peripheral blood of prostate patients.     

Survival analysis of genome-wide gene expression profiles of prostate cancers identifies new prognostic targets of disease relapse

Current models of prostate cancer classification are poor at distinguishing between tumors that have similar histopathological features but vary in clinical course and outcome. Here, we applied classical survival analysis to genome-wide gene expression profiles of prostate cancers and preoperative prostate-specific antigen (PSA) levels from each patient, to identify prognostic markers of disease relapse that provide additional predictive value relative to PSA concentration. Three of approximately 200 probesets showing strongest correlation with relapse were identified as the gene for the putative calcium channel protein, trp-p8, with loss of trp-p8 mRNA expression associated with a significantly shorter time to PSA relapse-free survival. We observed subsequently that trp-p8 is lost in the transition to androgen independence in a prostate cancer xenograft model and in prostate cancer tissue from patients treated preoperatively with antiandrogen therapy, suggesting that trp-p8 is androgen regulated, and its loss may be associated with more advanced disease. The identification of trp-p8 and other proteins implicated in the phosphatidylinositol signal transduction pathway that are associated with prostate cancer outcome, both here and in other published work, suggests an integral role for this pathway in prostate carcinogenesis. Thus, our findings demonstrate that multivariable survival analysis can be applied to gene expression profiles of prostate cancers with censored follow-up data and used to identify molecular markers of prostate cancer relapse with strong predictive power and relevance to the etiology of this disease.     

前列腺特异性抗原人类激肽释放酶2和前列腺特异的膜抗原在前列腺癌患者外周血表达的临床意义

目的　探讨检测前列腺癌微转移的灵敏和特异性指标。方法　从 5 1例前列腺癌、33例前列腺增生 (BPH)患者及 32名正常人的外周血中分离单个核细胞 ,用巢式RT PCR方法检测其中前列腺上皮细胞前列腺特异性抗原 (PSA)、人类激肽释放酶 2 (hK2 )和前列腺特异的膜抗原 (PSMA)的表达。结果　PSA、hK2和PSMA在前列腺癌患者外周血中检出的阳性率分别为 5 2 .9%、4 3.1%和6 4 .7% ;正常人和BPH患者假阳性率分别为 6 .2 %、7.7%和 4 .6 % ,3项指标差异均有显著性 (P <0 .0 1)。各临床分期 (局限癌、侵袭性癌和转移癌 )间 ,PSA和hK2的阳性检出率差异无显著性 ;PSMA在各期前列腺癌中阳性检出率均较PSA和hK2高 ,且随临床分期进展 ,其阳性检出率亦增加 (P <0 .0 5 )。结论PSMA对前列腺癌诊断、治疗方案的选择及预后评估较PSA和hK2有更大的价值     

Comparison of human prostate specific glandular kallikrein 2 and prostate specific antigen gene expression in prostate with gene amplification and overexpression of prostate specific glandular kallikrein 2 in tumor tissue.

BACKGROUND: There is a need for specific markers that can indicate the different stages of prostate carcinoma. There is ongoing speculation concerning the value of prostate specific glandular kallikrein (hK2) in this regard. METHODS: The expression levels of both hK2 and human prostate specific antigen (hPSA) were compared at the mRNA and protein levels by using in situ hybridization and immunohistochemistry techniques in tissue specimens from patients with benign prostatic hyperplasia and malignant prostate carcinoma. The respective gene copy numbers were analyzed by a competitively differential polymerase chain reaction-based method. RESULTS: In situ hybridization revealed that hK2 was expressed at significantly higher levels in malignant prostate tissue compared with benign prostate tissue (P < 0.0005), whereas hPSA expression levels were the reverse (P = 0.06). In benign tissue, the mean level of hK2 mRNA was 82% of the respective value of hPSA (P < 0.003), whereas, in tumor tissue, the mean hK2 expression level was 21% higher than that of hPSA (P < 0.01). The results at the protein level supported the mRNA findings: hPSA expression was lower in malignant tissues compared with benign tissues (17 of 20 specimens), whereas an increase in hK2 expression was detected in 17 of 19 specimens. The authors report that the hK2 gene (hKLK2) was amplified in prostate carcinoma tissue, whereas the hPSA gene was not. There was a correlation between hPSA and hK2 mRNA levels in both benign tissue (correlation coefficient [r] = 0.735; P < 0.01) and malignant tissue (r = 0.767; P < 0.01). CONCLUSIONS: Gene amplification of hKLK2 may be one of the factors leading to higher expression of hK2 in prostate carcinoma. The correlation between hK2 and hPSA expression levels indicates coordinated expression of the genes in both normal and abnormal prostate gland. The results suggest the potential value of hK2 in the diagnosis of prostate carcinoma through mRNA analyses and gene amplification.     

Prostate stem cell antigen (PSCA) expression in human prostate cancer tissues: implications for prostate carcinogenesis and progression of prostate cancer

OBJECTIVE: Prostate stem cell antigen (PSCA) is a recently defined homolog of the Thy-1/Ly-6 family of glycosylphosphatidylinositol (GPI)-anchored cell surface antigens. The objective of the present study was to examine the expression status of PSCA protein and mRNA in clinical specimens of human prostate cancer (PCa) and to validate it as a potential molecular target for diagnosis and treatment of PCa. METHODS: Immunohistochemical (IHC) and in situ hybridization (ISH) analyses of PSCA expression were simultaneously performed on paraffin-embedded sections of 20 benign prostatic hyperplasia (BPH), 20 prostatic intraepithelial neoplasm (PIN) and 48 prostate cancer (PCa) tissues, including 9 androgen-independent prostate cancers. The level of PSCA expression was semiquantitatively scored by assessing both the percentage and intensity of PSCA-positive staining cells in the specimens. We then compared the PSCA expression between BPH, PIN and PCa tissues and analyzed the correlations of PSCA expression level with pathological grade, clinical stage and progression to androgen-independence in PCa. RESULTS: In BPH and low grade PIN, PSCA protein and mRNA staining were weak or negative and less intense and uniform than that observed in high grade PIN (HGPIN) and PCa. Moderate to strong PSCA protein and mRNA expression were noted in 8 of 11 (72.7%) HGPIN and in 40 of 48 (83.4%) PCa specimens examined by IHC and ISH analyses, and their statistical significance was compared with BPH (20%) and low-grade PIN (22.2%) specimens (P < 0.05). The expression level of PSCA increased with a higher Gleason grade, advanced stage and progression to androgen-independence (P < 0.05). In addition, IHC and ISH staining revealed a high degree of correlation between PSCA protein and mRNA overexpression. CONCLUSIONS: Our data demonstrate that PSCA as a new cell surface marker is overexpressed in a majority of cases of human PCa. PSCA expression correlates positively with adverse tumor characteristics, such as increasing pathological grade (poor cell differentiation), worsening clinical stage and androgen-independence and speculatively with prostate carcinogenesis. PSCA may possess prognostic utility and may be a promising molecular target for diagnosis and treatment of PCa.     

Free PSA isoforms and intact and cleaved forms of urokinase plasminogen activator receptor in serum improve selection of patients for prostate cancer biopsy

Clinicians currently use simple cut-points, such as serum prostate-specific antigen (PSA) >or=4 ng/ml, to decide whether to recommend further work-up for prostate cancer (PCa). As an alternative strategy, we evaluated multivariable models giving probabilities of a PCa diagnosis based on PSA and several circulating novel biomarkers. We measured total PSA, free PSA (fPSA), fPSA subfractions (single-chain fPSA-I and multichain fPSA-N), total human glandular kallikrein 2 (hK2) and full-length and cleaved forms of soluble urokinase plasminogen activator receptor (suPAR) in pretreatment serum from 355 men referred for prostate biopsy. Age and total PSA were combined in a "base" regression model to predict biopsy outcome. We then compared this base model to models supplemented by various combinations of circulating markers, using concordance index (AUC) to measure diagnostic discrimination. PCa prediction was significantly enhanced by models supplemented by measurements of suPAR fragments and fPSA isoforms. Addition of these markers improved bootstrap-corrected AUC from 0.611 for a cut-point and 0.706 for the base model to 0.754 for the full model (p=0.005). This improved diagnostic accuracy was also seen in subanalysis of patients with PSA 2-9.99 ng/ml and normal findings on DRE (0.652 vs. 0.715, p=0.039). In this setting, hK2 did not add diagnostic information. Measurements of individual forms of suPAR and PSA isoforms contributed significantly to discrimination of men with PCa from those with no evidence of malignancy.     

Real-time quantitative RT-PCR assessment of PIM-1 and hK2 mRNA expression in benign prostate hyperplasia and prostate cancer

To investigate the expressions of PIM-1 and hK2 mRNA in normal prostate, benign prostatic glandular hyperplasia (BPH), and prostate cancer (PCa), and to explore the association of PIM-1 and hK2 expressions with PCa progression. The samples were harvested from 37 patients with BPH, 23 patients with PCa, and three with normal prostate tissues. Total RNA was extracted from their prostate tissues and analyzed for PIM-1 and hK2 mRNA levels using SYBR green I-based quantitative real-time RT-PCR (QRT-PCR) assays and Southern blot analysis. The differences of gene expressions were calculated based on standard curve. Quantitative expressions of PIM-1 and hK2 mRNA in normal prostate, BPH, and PCa were 1.05 ± 0.04, 2.57 ± 0.74, 4.45 ± 0.63, and 1.02 ± 0.03, 2.264 ± 0.46, 5.905 ± 0.78, respectively. PIM-1 and hK2 were expressed higher in PCa than those in BPH and normal prostate tissues, the differences among which had statistic significance (P < 0.05). Our results support the hypothesis that PIM-1 and hK2 play a significant role in the growth of PCa and the detection of PIM-1 and hK2 mRNA expressions by QRT-PCR provided more reliable and helpful information on diagnosis, treatment, and prognosis of PCa. Hui-chan He and Xue-cheng Bi contributed equally to this article.     

Tissue specific expression and serum levels of human tissue factor in patients with urological cancer

Human tissue factor (TF) is involved in tumor angiogenesis and metastasis. However, little is known about the distribution of TF in urological cancer. In this study we investigated the TF expression in tumor tissue and autologous non-malignant tissue as well as in serum of patients with renal cell carcinoma (RCC), bladder cancer, and prostate cancer (PCa). To study the distribution of TF in tumor tissue and in the surrounding non-malignant tissue, we measured TF protein by ELISA in tissue specimens obtained intraoperatively from 18 RCC, seven bladder cancer and six PCa patients. Differences in TF expression were found between tumor tissue and nonmalignant tissue for the three tumor types at the protein level (in the order RCC < bladder cancer < PCa). In all but one of the 18 RCC patients (94 %) higher TF protein level was observed in non-malignant tissue as compared to the tumor tissue. In addition, the relative TF mRNA expression analyzed by a quantitative RT-PCR assay in the same RCC tissue sample pairs was higher in 78% of non-malignant tissues in comparison to the tumor tissue specimens. Moreover, using enzyme linked immunosorbent assay the TF protein content was measured in serum samples of 66 patients with bladder cancer, 75 RCC patients and 157 PCa patients, and was compared with the TF serum level of 92 healthy volunteers. Whereas no differences were detected between normal volunteers and patients with PCa or RCC, patients with bladder cancer showed a significantly increased level of serum TF (P=0.0076). However, no causal association between TF levels in serum and TF content in tissue extracts for all three tumor types of urological tumors was found. Our results suggest that TF in non-malignant renal tissues was expressed at a higher level compared to the supposed de novo TF expression in RCC tissue specimens. This indicates a tumor-associated induction of TF expression in the TF-negative RCC progenitor cells. The increased serum TF levels are alike the reported higher urinary TF levels found in bladder cancer patients. The potential clinical relevance of this finding should be further elucidated.     

Human Kallikrein-2, Prostate Specific Antigen and Free- Prostate Specific Antigen in Combination to Discriminate Prostate Cancer from Benign Diseases in Syrian Patients

Background: The high incidence of prostate cancer as the most common malignancy in males in many countriesraises the question of developing reliable detection tests. The prostate specific antigen (PSA) test is the mostwidely used for screening for prostate cancer; however, its low specificity elevates the number of unnecessarilybiopsies. Serum human kallikrein-2 (hK2) is considered as a promising marker, and especially its ratio to fPSA,for predicting the presence of malignancy to select the best choice referring to biopsy or surveillance. In this study,we investigated the role of hK2 and its combinations with other markers to discriminate prostate cancer frombenign diseases in Syrian patients. Materials and Methods: In this prospective oriented cross-sectional cohortstudy, serum samples were collected from patients referred to many Hospitals in Damascus, Syria, between May2011 and March 2012, and diagnosed with biopsy proven benign prostate hyperplasia (BPH) or prostate cancer(PCa). Serum was analyzed for hK2, PSA and fPSA, and the ratios of fPSA/PSA and hK2/fPSA were calculated.Results: We found that mean hK2/fPSA ratios were significantly higher (P=0.01) in prostate cancer patientsthan in the BPH or control groups. Also the ratio hk2/fPSA gave the largest area under the curve (AUC:0.96)which was significantly larger than for fPSA/PSA (AUC:0.41) indicative of higher specificity. Conclusions: Ourresults demonstrate that the ratio of hK2/fPSA might be superior to the use of fPSA/PSA alone. The hK2 couldbe shown to enhance the early detection of prostate cancer; especially the ratio hK2/fPSA improves specificityand hence may reduce the number of negative biopsies.