Biomedical subjects
Publications and source records attributed to Carsten Stephan.
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Gene products of the A disintegrin and metalloprotease (ADAM) family are critically involved in carcinogenesis and tumor progression of various solid tumors. Little is known about ADAM8 in prostate cancer. In our quest for novel diagnostic tissue markers of prostate cancer, we aimed to evaluate the expression of ADAM8 in prostate cancer and to correlate it with clinicopathological parameters. One hundred twenty-eight clinicopathologically characterized prostate cancer patients, with available follow-up data, were immunostained for ADAM8. Additionally, ADAM8 mRNA expression was quantified by real-time reverse transcription polymerase chain reaction (n = 59). ADAM8 protein expression was significantly associated with higher pT status, positive nodal status, and higher Gleason scores. Still, a significant prognostic value for the prostate-specific antigen relapse-free survival of ADAM8 could not be demonstrated. The differentiality of ADAM8 expression on protein and on mRNA level was low and partially inconclusive. Therefore, despite of its significant association with conventional parameters of an unfavorable prognosis, ADAM8 adds only limited information to the conventional histopathological assessment of prostate cancer.
OBJECTIVE: The pro-forms of prostate specific antigen (-2,-5,-7 proPSA) and also %free PSA based artificial neural networks (ANN) have been suggested to enhance the discrimination between prostate cancer (PCa) and no evidence of malignancy (NEM). This study reports on the combined use of proPSA within a %free PSA based ANN to enhance specificity of PCa. METHODS: Serum samples from 898 patients with PCa (n=384) or NEM (n=514) within the PSA range 1-10 microg/l were analyzed for PSA, free PSA and (-5,-7) proPSA (Roche assays). Patient data from two centers - taken first from the Swiss site of the ERSPC (Aarau) and from a referral population (Berlin) have been analyzed. Leave-one-out ANN models with the variables PSA, %fPSA, proPSA, prostate volume and status of digital rectal examination (DRE) were constructed and compared by receiver-operating characteristic (ROC) curve analysis. RESULTS: (-5,-7) proPSA was only significantly different between NEM and PCa in the PSA range 4-10 microg/l. Within the PSA range 4-10 microg/l (Berlin group) the ANN including only the two variables %fPSA and proPSA could reach the same performance like the conventional ANN with PSA, %fPSA, age, prostate volume and DRE (both AUCs: 0.84) However, at 95% sensitivity all ANN could not improve specificity compared to %fPSA. CONCLUSIONS: ProPSA as single parameter did not improve specificity over %fPSA whereas proPSA and %fPSA within an ANN in the PSA range 4-10 microg/l substituted prostate volume and DRE. At 95% sensitivity only ANN with prostate volume and DRE perform significantly better than %fPSA.
BACKGROUND: We aimed to evaluate the value of macrophage inhibitory cytokine 1 (MIC-1), human kallikrein 11 (hK11) migration inhibitor factor (MIF) in comparison to prostate-specific antigen (PSA) and % fPSA and also to develop a % fPSA-based ANN with the new input factors to determine whether these additional markers can further eliminate unnecessary prostate biopsies. METHODS: Serum samples from 371 patients with prostate cancer (PCa, n=135) or benign prostate hyperplasia (BPH, n=236) within the PSA range 0.5-20 microg/L were analyzed for total PSA, free PSA, MIC-1, hK11, and MIF. 'Leave one out' ANN models with these variables and prostate volume were constructed and compared to logistic regression (LR) and all single parameters. RESULTS: The discriminatory power of MIC-1, hK11, and MIF was less than that for PSA despite significant differences in BPH compared to PCa patients. At 90% and 95% sensitivity, the artificial neural networks (ANNs) were only significantly better than % fPSA if prostate volume was included. CONCLUSIONS: ANNs with the novel input factors of MIC-1, MIF, and/or hK11 and additional use of prostate volume demonstrated significant advantage compared with % fPSA and tPSA and may lead to a reduction in unnecessary prostate biopsies.
PURPOSE: Current serum testing for the detection of prostate cancer (PCa) lacks specificity. On diagnosis, the optimal therapeutic pathway is not clear and tools for adequate risk assessment of localized PCa progression are not available. This leads to a significant number of men having unnecessary diagnostic biopsies and surgery. A search for novel tumor markers identified macrophage inhibitory cytokine 1 (MIC-1) as a potentially useful marker. Follow-up studies revealed MIC-1 overexpression in local and metastatic PCa whereas peritumoral interstitial staining for MIC-1 identified lower-grade tumors destined for recurrence. Consequently, we sought to assess serum MIC-1 measurement as a diagnostic tool. EXPERIMENTAL DESIGN: Using immunoassay determination of serum MIC-1 concentration in 1,000 men, 538 of whom had PCa, we defined the relationship of MIC-1 to disease variables. A diagnostic algorithm (MIC-PSA score) based on serum levels of MIC-1, total serum prostate-specific antigen, and percentage of free prostate-specific antigen was developed. RESULTS: Serum MIC-1 was found to be an independent predictor of the presence of PCa and tumors with a Gleason sum > or =7. We validated the MIC-PSA score in a separate population and showed an improved specificity for diagnostic blood testing for PCa over percentage of free prostate-specific antigen, potentially reducing unnecessary biopsies by 27%. CONCLUSIONS: Serum MIC-1 is an independent marker of the presence of PCa and tumors with a Gleason sum of > or =7. The use of serum MIC-1 significantly increases diagnostic specificity and may be a future tool in the management of PCa.
PURPOSE: Comparison of diagnostic performances of total and complexed prostate specific antigen (tPSA, cPSA) using ROC curve analysis resulted in controversial interpretations. To overcome the biases due to analysis of subgroups selected by concentration ranges of only 1 marker the novel approach named discordance analysis characteristics is presented. MATERIALS AND METHODS: DAC method includes only patients who are discordantly categorized by the tests under investigation using cutoffs with identical sensitivity. Within those patients parameters describing diagnostic accuracy are calculated. The systematic nature of the results is confirmed by varying the cutoffs. DAC method is demonstrated considering the example of cPSA and tPSA data from 2 prostate cancer studies with a total of 1,624 patients (study A and B, of 283 and 565 with as well as 417 and 359 men without prostate cancer, respectively). PSA concentrations were measured using the Bayer assays. RESULTS: The DAC method proved that cPSA outperformed tPSA regarding the criteria diagnostic specificity and positive predictive value. Among discordantly tested patients cPSA detected patients with an about 2-fold higher risk to have prostate cancer and with a 3.6 to 5.5-fold better specificity than tPSA. Using cPSA instead of tPSA more than 10% of unnecessary biopsies could be avoided in the tPSA range of 3 to 5 ng/ml. CONCLUSIONS: The superior diagnostic performance of cPSA in comparison with tPSA warrants the recommendation to use cPSA as initial test in prostate cancer diagnostics. The DAC method is generally recommended to replace comparative ROC analyses of subgroups to evaluate the diagnostic usefulness of markers.
BACKGROUND: Human glandular kallikrein (hK2) has been shown to add important information regarding the early detection and staging of prostate cancer. Preliminary analysis pointed out that hK2 may discriminate between pT2 and pT3 tumors, and that hK2 may predict Gleason grade 4/5 cancer volume, better than prostate-specific antigen (PSA) or percent free PSA (%fPSA). We investigated the role of hK2 serum values for predicting pathological stage, grade and Gleason score. METHODS: Prostate-specific antigen, free PSA and hK2 were measured on 222 untreated prostate cancer patients who had received radical prostatectomy at the Charité Hospital, Berlin, Germany. Pathological work up revealed pT2-cancer in 111 patients and pT3-cancer in 111 patients. Grade 2 was found in 118 patients whereas grade 3 tumors were found in 104 patients. RESULTS: For pT2 and pT3 patients, the %fPSA (P=0.006), the ratios hK2/fPSA (P=0.08) and hK2xtPSA/fPSA (P=0.002) were all significant different whereas hK2 (P=0.143) and PSA (P=0.1) did not differ. Between grade 2 and grade 3 tumors, the hK2 alone (P=0.27), the %fPSA (P=0.13), the ratios hK2/fPSA (P=0.94) and hK2xtPSA/fPSA (P=0.12) did not separate, whereas PSA (P=0.039) showed a difference. The same relationships were found between the two groups in Gleason score<7 and >or=7. Neither the hK2 ratio, nor % fPSA was different. CONCLUSION: Human glandular kallikrein was not different between pT2 and pT3, nor between G2 versus G3 or Gleason scores<7 and >or=7 prostate cancer. Together with %fPSA, hK2 may only help to distinguish preoperatively between pT2 and pT3 prostate cancer but cannot add further information.
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BACKGROUND: The comparability of total and free prostate-specific antigen (tPSA and fPSA) results among commercial PSA assays has been suggested to be improved by calibration to WHO PSA reference materials and the development of equimolar-response assays. To characterize the current situation, we assessed 5 frequently used commercial assay combinations for tPSA and fPSA regarding the interchangeability of the PSA values and the ratio of fPSA to tPSA (%fPSA), equimolar characteristics, and diagnostic accuracy. METHODS: Sera from 314 patients with prostate cancer (PCa) and 282 men with no evidence of prostate cancer (NPCa) were measured with tPSA and fPSA assays from Abbott (AxSYM), Beckman Coulter (Access), Diagnostic Products Corporation (Immulite 2000), and Roche (Elecsys 2010) and with tPSA and complexed PSA (cPSA) assays from Bayer (ADVIA Centaur). RESULTS: Method comparisons (Passing and Bablok regressions; Bland-Altman plots) showed assay-dependent results for tPSA, fPSA, and %fPSA. With the Access tPSA values taken as 100%, tPSA concentrations varied from 87% (AxSYM and ADVIA Centaur) to 115% (Immulite), leading to different numbers of patients classified according to the commonly recommended tPSA cutoffs for performing a biopsy. Different %fPSA values also led to assay-dependent ROC analysis results, a finding that shows the importance for the diagnostic accuracy. CONCLUSION: Interchangeability of tPSA, fPSA, and %fPSA values obtained by commercial PSA assays remains inadequate, but attention to this issue may minimize the misinterpretation of PSA results obtained by different assays.
The human kallikrein locus on chromosome 19q13.3-13.4 contains kallikrein 1--the tissue kallikrein--and 14 related serine proteases. Recent investigations into their function and evolution have indicated that the present nomenclature for these proteins is inadequate or insufficient. Here we present a new nomenclature in which proteins without proven kininogenase activity are denoted kallikrein-related peptidase. Names are also given to the unique rodent proteins that are closely related to kallikrein 1.
Human kallikrein 11 (hK11) was evaluated in a percentage free PSA-based artificial neural network (ANN) to reduce unnecessary prostate biopsies. Serum samples from 357 patients with (n=132) and without (n=225) prostate cancer (PCa) were analyzed and ANN models were constructed and compared to all parameters. The discriminatory power of hK11 was lower than that of PSA, but receiver operator characteristic (ROC) analyses demonstrated significantly larger areas under the curves for the ANN compared to all other parameters. ANNs with hK11 may lead to a further reduction in unnecessary prostate biopsies, especially when analyzing patients with less than 15% free PSA.
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Using quantitative reverse transcription-polymerase chain reaction (RT-PCR), reference genes are utilized as endogenous controls for relative quantification of target genes in gene profiling studies. The suitability of housekeeping genes for that purpose in prostate cancer tissue has not been sufficiently investigated so far. The objective of this study was to select from a panel of 16 potential candidate reference genes the most stable genes for gene normalization. Expression of mRNA encoding ACTB, ALAS1, ALB, B2M, G6PD, GAPD, HMBS, HPRT1, K-ALPHA-1, POLR2A, PPIA, RPL13A, SDHA, TBP, UBC, and YWHAZ was examined in matched, microdissected malignant and nonmalignant tissue specimens obtained from 17 nontreated prostate carcinomas after radical prostatectomy by real-time RT-PCR. The genes studied displayed a wide expression range with cycle threshold values between 16 and 37. The expression was not different between samples from pT2 and pT3 tumors or between samples with Gleason scores <7 and >or=7 (P>0.05). ACTB, RPL13A, and HMBS showed significant differences (P<0.02 at least) in expressions between malignant and nonmalignant pairs. All other genes did not differ between the matched pairs, and the software programs geNorm and NormFinder were used to ascertain the most suitable reference genes from these candidates. HPRT1, ALAS1, and K-ALPHA-1 were calculated by both programs to be the most stable genes covering a broad range of expression. The expression of the target gene RECK normalized with HRPT1 alone and with the normalization factors generated by the combination of these three reference genes as well as with the unstable genes ACTB or RPL13A is given. That example shows the significance of using suitable reference genes to avoid erroneous normalizations in gene profiling studies for prostate cancer. The use of HPRT1 alone as a reference gene shown in our study was sufficient, but the normalization factors generated from two (HRPT1, ALAS1) or all three genes (HRPT1, ALAS1, K-ALPHA-1) should be considered for an improved reliability of normalization in gene profiling studies of prostate cancer.
BACKGROUND: The objective of this study was to evaluate the prostate specific antigen (PSA) density (PSAD) (the quotient of PSA and prostate volume) compared with the percent free PSA (%fPSA) in different total PSA (tPSA) ranges from 2 ng/mL to 20 ng/mL. Possible cut-off levels depending on the tPSA should be established. METHODS: In total, 1809 men with no pretreatment of the prostate were enrolled between 1996 and 2004. Total and free PSA were measured with the IMMULITE PSA and Free PSA kits (Diagnostic Products, Los Angeles, CA). Prostate volume was determined by transrectal ultrasound. The diagnostic validity of tPSA, %fPSA, and PSAD was evaluated by receiver operation characteristic (ROC) curve analysis. RESULTS: The PSAD differed significantly (P < 0.0001) between patients with prostate carcinoma and patients with benign prostatic hyperplasia in all analyzed ranges of tPSA and prostate volume. At the 90% and 95% sensitivity levels and regarding the area under the ROC curve (AUC) within the tPSA range of 2-4 ng/mL, The PSAD was significantly better than tPSA and %fPSA. Within the tPSA range of 4-10 ng/mL, the PSAD did not perform better than %fPSA. CONCLUSIONS: PSAD showed a better performance than %fPSA at tPSA concentrations < 4 ng/mL for detecting prostate carcinoma, with a significantly larger AUC for PSAD (0.739) compared with %fPSA (0.667). PSAD did not perform better than %fPSA when the tPSA range of 4-10 ng/mL was analyzed. Different PSAD cut-off values of 0.05 at tPSA 2-4 ng/mL, 0.1 at tPSA 4-10 ng/mL, and 0.19 at 10-20 ng/mL were necessary to reach 95% sensitivity.
BACKGROUND: Human protectin (CD59) is a regulator of complement activation that inhibits complement-mediated cell lysis, and thus might confer immune resistance to tumor cells. CD59 expression has been described in a variety of human malignancies, including breast cancer. Since a comprehensive investigation of CD59 expression in prostate cancer has not been conducted yet, we aimed to determine the significance of CD59 expression in prostate cancer. METHODS: Eighty-six primary adenocarcinomas of the prostate were immunostained using a monoclonal CD59 antibody (clone MEM-43) and a standard detection system. The immunoreactivity of the tumor was evaluated as low versus high for statistical analysis. Additionally, CD59 mRNA levels were determined by real-time PCR in matched (tumor/normal) microdissected tissues from 26 cases. RESULTS: Cytoplasmic CD59 immunoreactivity was found in epithelia of prostate cancer, prostatic intraepithelial neoplasia, benign hyperplasia, atrophic, and normal glands. High rates of CD59 expression were noted in 36% of prostate cancer cases and were significantly associated with tumor pT stage (P = 0.043), Gleason grade (P = 0.013) and earlier biochemical (PSA) relapse in Kaplan-Meier analysis (P = 0.0013). On RNA level, we found an upregulation in 19.2% (five cases), although the general rate of CD59 transcript was significantly lower in tumor tissue (P = 0.03). CONCLUSION: CD59 protein is strongly expressed in 36% of adenocarcinomas of the prostate and and is associated with disease progression and adverse patient prognosis.
BACKGROUND: All human kallikrein (KLK) genes have at least one splice variant, some of which possess clinical utility in cancer diagnostics/prognostics. Given that introns <100 bp in length are retained in 95% of human genes and that splice variants of KLK3 and KLK4 retain intron III, we hypothesized that other proteins in this family, with a small intron III, may also retain it. METHODS: Variant-specific reverse transcription-PCRs (RT-PCRs) for KLK1, KLK2, KLK5, and KLK15 were used to identify and clone the full coding sequence of intron III-containing splice variants. In addition, variant-specific RT-PCRs for the cloned KLK3 and KLK4 variants as well as for the "classical" forms of the six genes were used to determine their expression profiles in healthy tissues, their regulation by steroids, and their differential expression in prostate cancer. RESULTS: KLK1, KLK2, KLK3, KLK4, KLK5, and KLK15 showed a common type of splice variant in which intron III is retained. Expression profiling of these splice variants revealed expression profiles similar to those of the classical mRNA forms, although the pattern of hormonal regulation was different. The KLK15 splice variant was up-regulated in 8 of 12 cancerous prostate tissues. All encoded variant proteins were predicted to be truncated and catalytically inactive because of a lack of the serine residue of the catalytic triad. CONCLUSIONS: The first six centromeric members of the KLK gene family have splice variants that retain intron III. Some variants show tissue-specific expression. The KLK15 splice variant appears to be a candidate biomarker for prostate cancer.
BACKGROUND: Recent studies suggest that macrophage migration inhibitory factor (MIF) in serum is of prognostic significance for prostate cancer. The aim of this study was to re-evaluate this hypothesis. METHODS: Serum MIF levels were measured in healthy men (n = 86), untreated patients with benign prostate hyperplasia (BPH; n = 50), prostate cancer (PCa; n = 163), and after radical prostatectomy for 3 days (n = 5). PCa patients were classified according to the TNM system and the WHO grading scale. Prostate specific antigen (PSA) and C-reactive protein (CRP) were additionally determined. RESULTS: The MIF concentrations of healthy men and BPH patients did not differ (mean +/- SD, 2.08 +/- 1.08 microg/L vs. 2.04 +/- 1.08 microg/L), whereas the mean value of MIF in PCa patients was significantly decreased (1.77 +/- 1.12 microg/L). There was no any correlation between MIF and PSA (r(s) = -0.049, P = 0.271). MIF concentrations in patients with T1 tumors were higher than in those with T2 tumors (2.29 +/- 1.26 vs. 1.67 +/- 1.11 microg/L; P = 0.044). No any effect of grading was observed. After prostatectomy, the changes of PSA and MIF were not always concordant as MIF partly increased while PSA continuously decreased. Analyses of receiver-operating curves and logistic regressions did not show that MIF alone or MIF related variables (MIF/tPSA; fPSA/(tPSA x MIF); fPSA x MIF/tPSA) could improve specificity or sensitivity to detect prostate cancer in comparison to total PSA. CONCLUSION: Serum MIF alone or MIF to PSA related variables did not seem suitable for providing additional information on PCa patients. That re-evaluated diagnostic validity of MIF was in contrast to results by another group shown previously.
In order to screen for differentially expressed genes that might be useful in diagnosis or therapy of prostate cancer we have used a custom made Affymetrix GeneChip containing 3950 cDNA fragments. Expression profiles were obtained from 42 matched pairs of mRNAs isolated from microdissected malignant and benign prostate tissues. Applying three different bioinformatic approaches to define differential gene expression, we found 277 differentially expressed genes, of which 98 were identified by all three methods. Fourteen per cent of these genes were not found in other expression studies, which were based on bulk tissue. Resultant candidate genes were further validated by quantitative RT-PCR, mRNA in situ hybridization and immunohistochemistry. AGR2 was over-expressed in 89% of prostate carcinomas, but did not have prognostic significance. Immunohistologically detected over-expression of MEMD and CD24 was identified in 86% and 38.5% of prostate carcinomas respectively, and both were predictive of PSA relapse. Combined marker analysis using MEMD and CD24 expression proved to be an independent prognostic factor (RR = 4.7, p = 0.006) in a Cox regression model, and was also superior to conventional markers. This combination of molecular markers thus appears to allow improved prediction of patient prognosis, but should be validated in larger studies.