John J. Grygiel

International Validation of a Preoperative Nomogram for Prostate Cancer Recurrence After Radical Prostatectomy

PURPOSE We evaluated the predictive accuracy of a recently published preoperative nomogram for prostate cancer that predicts 5-year freedom from recurrence. We applied this nomogram to patients from seven different institutions spanning three continents. METHODS Clinical data of 6,754 patients were supplied for validation, and 6,232 complete records were used. Nomogram-predicted probabilities of 60-month freedom from recurrence were compared with actual follow-up in two ways. First, areas under the receiver operating characteristic curves (AUCs) were determined for the entire data set according to several variables, including the institution where treatment was delivered. Second, nomogram classification-based risk quadrants were compared with actual Kaplan-Meier plots. RESULTS The AUC for all institutions combined was 0.75, with individual institution AUCs ranging from 0.67 to 0.83. Nomogram predictions for each risk quadrant were similar to actual freedom from recurrence rates: predicted probabilities of 87% (low-risk group), 64% (intermediate-low-risk group), 39% (intermediate-high-risk group), and 14% (high-risk group) corresponded to actual rates of 86%, 64%, 42%, and 17%, respectively. The use of neoadjuvant therapy, variation in the prostate-specific antigen recurrence definitions between institutions, and minor differences in the way the Gleason grade was reported did not substantially affect the predictive accuracy of the nomogram. CONCLUSION The nomogram is accurate when applied at international treatment institutions with similar patient selection and management strategies. Despite the potential for heterogeneity in patient selection and management, most predictions demonstrated high concordance with actual observations. Our results demonstrate that accurate predictions may be expected across different patient populations.

The Propeptide Mediates Formation of Stromal Stores of PROMIC-1: Role in Determining Prostate Cancer Outcome

The extracellular matrix (ECM) is a reservoir of cellular binding proteins and growth factors that are critical for normal cell behavior, and aberrations in the ECM invariably accompany malignancies such as prostate cancer. Carcinomas commonly overexpress macrophage inhibitory cytokine 1 (MIC-1), a proapoptotic and antitumorigenic transforming growth factor-beta superfamily cytokine. Here we show that MIC-1 is often secreted in an unprocessed propeptide containing form. It is variably processed intracellularly, with unprocessed forms being secreted from several tumor lines, including prostate carcinoma lines, PC-3 and LNCaP. Once secreted, only unprocessed proMIC-1 binds ECM, demonstrating for the first time the occurrence of extracellular stores of MIC-1. The propeptide mediates this association via its COOH-terminal 89 amino acids. Xenograft models bearing tumors secreting various engineered forms of MIC-1 show that the propeptide regulates the balance between ECM stores and circulating serum levels of mature MIC-1 in vivo. The absence of propeptide results in approximately 20-fold increase in serum MIC-1 levels. The significance of stromal MIC-1 stores was evaluated in prostate cancer tissue cores, which show major variation in stromal levels of MIC-1. Stromal MIC-1 levels are linked to prostate cancer outcome following radical prostatectomy, with decreasing stromal levels providing an important independent predictor of disease relapse. In low-grade localized prostate cancer (Gleason sum score < or = 6), the level of MIC-1 stromal stores was the best predictor of future relapse when compared with all other clinicopathologic variables. The secretion and ECM association of unprocessed proMIC-1 is likely to play a central role in modulating local bioavailability of MIC-1 which can affect patient outcome in prostate cancer and other epithelial tumors.

Prognostic significance of Gleason pattern in patients with Gleason score 7 prostate carcinoma

In the current study, the authors sought to further stratify the prognosis of patients with Gleason score (GS) 7 prostate carcinoma. They assessed the influence on outcome of a predominant poorly differentiated Gleason pattern (primary Gleason pattern [GP] 4) and/or a coincident small focus of poorly differentiated tumor of higher grade (tertiary GP 5).

Prognostic significance of preoperative factors in localized prostate carcinoma treated with radical prostatectomy

Predicting outcome for men with clinically localized prostate carcinoma treated with curative intent remains imprecise and further evaluation of accepted and potential predictive factors is needed.

Expression of the zinc transporter ZnT4 is decreased in the progression from early prostate disease to invasive prostate cancer.

We have utilized oligonucleotide microarrays to identify novel genes of potential clinical and biological importance in prostate cancer. RNA from 74 prostate cancers and 164 normal body samples representing 40 different tissues were analysed using a customized Affymetrix GeneChip® oligonucleotide microarray representative of over 90% of the expressed human genome. The gene for the zinc transporter ZnT4 was one of several genes that displayed significantly higher expression in prostate cancer compared to normal tissues from other organs. A polyclonal antipeptide antibody was used to demonstrate ZnT4 expression in the epithelium of all 165 elements of benign and 326 elements of localized prostate cancers examined and in nine of 10 advanced prostate cancer specimens by immunohistochemistry. Interestingly, decreased intensity of ZnT4 immunoreactivity occurred in the progression from benign to invasive localized prostate cancer and to metastatic disease. Immunofluorescence analysis and surface biotinylation studies of cells expressing ZnT4 localised the protein to intracellular vesicles and to the plasma membrane. These findings are consistent with a role for ZnT4 in vesicular transport of zinc to the cell membrane and potentially in efflux of zinc in the prostate.

Membranous Expression of Secreted Frizzled-Related Protein 4 Predicts for Good Prognosis in Localized Prostate Cancer and Inhibits PC3 Cellular Proliferation in Vitro

Purpose: Activation of the Wnt-signaling pathway is implicated in aberrant cellular proliferation in a variety of cancers. Secreted frizzled-related protein 4 (sFRP4) is a secreted protein with putative inhibitory activity of the Wnt-signaling cascade through binding and sequestering Wnt ligands. Because sFRP4 mRNA is overexpressed in prostate cancers (PCs), the aim of this study was to define the pattern of sFRP4 protein expression in normal and malignant human prostate tissue and to determine whether changes in expression were associated with disease progression and prognosis, as well as to define the phenotype of sFRP4-overexpression in an in vitro model of PC. Experimental Design: Polyclonal antibodies were raised against a COOH-terminal peptide of sFRP4, characterized and used to assess sFRP4 protein expression in benign prostate tissue and 229 patients with clinically localized PC (median follow-up 77 months, range 1–156). In vitro studies of the function of sFRP4 overexpression were performed using PC3 cells transfected with sFRP4. Results: Benign and malignant prostate tissue demonstrated cytoplasmic sFRP4 immunoreactivity, but there was a decrease in the expression of membranous sFRP4 in PCs compared with the hyperplastic lesions (P < 0.0001). Kaplan-Meier analysis revealed that patients whose PC expressed membranous sFRP4 in >20% of cells had improved relapse-free survival compared with those with ≤20% membranous expression (P = 0.002). Moreover, membranous sFRP4 expression (P = 0.04) was an independent predictor of relapse when modeled with Gleason score (P = 0.006), pathological stage (P = 0.002), and pre-operative prostate-specific antigen levels (P = 0.004). In addition, in vitro studies demonstrated a decrease in the proliferation rate of PC3 cells transfected with sFRP4 when compared with the control PC3-empty vector cells (P < 0.0001). Decreased levels of phosphorylated glycogen synthase kinase 3β in PC3-sFRP4 cells suggested that this phenotype is mediated by the “Wnt/β-catenin” pathway. Conclusions: These data suggest that sFRP4 expression may be prognostic for localized PC, potentially as a consequence of an inhibitory effect on PC cell proliferation.

Prognostic Significance of Pathologic Features in Localized Prostate Cancer Treated With Radical Prostatectomy: Implications for Staging Systems and Predictive Models

PURPOSE Although predicting outcome for men with clinically localized prostate cancer (PC) has improved, the staging system and nomograms used to do this are based on results from the North American health system. To be internationally applicable, these models require testing in cohorts from a variety of different health systems based on the predominant PC case identification methods used. PATIENTS AND METHODS We studied 732 men with localized PC treated with radical prostatectomy and no preoperative therapy between 1986 and 1999 at one Australian institution to determine the effect of clinicopathologic features on disease-free survival. RESULTS Preoperative serum prostate-specific antigen (PSA) concentration, Gleason score, pathologic stage, and year of surgery were independent predictors of outcome. Although margin status demonstrated only a trend toward significance in multivariate modeling overall, it proved to be independent in subgroups based on later year of surgery (1986 to 1994 v 1995 to 1998), preoperative PSA of less than 10 ng/mL, and Gleason score > or = 7. Adjuvant radiation therapy improved disease-free survival rates in patients with multiple surgical margin involvement. CONCLUSION This work confirms the prognostic significance of pathologic stage, Gleason score, and preoperative serum PSA. In the context of a contemporaneous screening effect in Australia, these findings may have implications for methods that predict outcome following surgery as screening becomes more prevalent in a population. The independent prognostic effect of margin status may alter with an increase in the proportion of screening-identified PCs. Staging systems and nomograms that predict outcome following surgery require validation in cohorts with different health practices before being universally applied.

Lower levels of nuclear β‐catenin predict for a poorer prognosis in localized prostate cancer

β‐catenin in its role as a nuclear signaling molecule has been implicated in prostate carcinogenesis primarily through modulation of androgen receptor activity. We defined the pattern of β‐catenin protein expression in the nuclei of normal, hyperplastic and malignant human prostate tissue and determined whether differences in expression were associated with disease progression and prognosis. Five normal prostates, 26 benign prostatic hypertrophy specimens, 232 radical prostatectomy specimens from patients with clinically localized prostate cancer (PC) and 20 cases of advanced PC were assessed for β‐catenin expression using immunohistochemistry. Nuclear β‐catenin expression in localized PC was significantly lower than that in benign prostate tissue (p < 0.001) and significantly higher than that in advanced PC tissue (p < 0.001). In addition, lower levels of nuclear β‐catenin expression (< 10% of cancer cells) predicted for a shorter biochemical relapse‐free survival in patients with localized PC (p = 0.008) and were inversely correlated with preoperative prostate‐specific antigen (PSA) levels (p = 0.01). Analysis of the low‐risk subgroup of patients with preoperative PSA levels < 10 ng/ml demonstrated that lower levels of nuclear β‐catenin expression (< 10% of cancer cells) again predicted for a poorer prognosis (p = 0.006). In conclusion, lower levels of nuclear β‐catenin expression are found in malignant compared to benign prostate tissue. In addition, lower nuclear β‐catenin expression is associated with a poorer prognosis in localized PC, in particular, in the low‐risk subgroup of patients with preoperative PSA levels < 10 ng/ml. Thus, the level of nuclear β‐catenin expression may be of clinical utility as a preoperative prognostic marker in low‐risk localized PC.

Aberrant Neuropeptide Y and Macrophage Inhibitory Cytokine-1 Expression Are Early Events in Prostate Cancer Development and Are Associated with Poor Prognosis

Studies to elucidate dysregulated gene expression patterns in premalignant prostate lesions have identified several candidate genes with the potential to be targeted to prevent the development and progression of prostate cancer and act as biomarkers of early disease. Herein, we explored the importance of two proteins, neuropeptide Y (NPY) and macrophage inhibitory cytokine-1 (MIC-1), as biomarkers of preinvasive prostate disease and investigated the relationship of expression to biochemical recurrence following treatment for localized prostate cancer. NPY and MIC-1 protein expression was determined by immunohistochemistry on tissue microarrays containing 1,626 cores of benign, low-grade prostatic intraepithelial neoplasia (PIN), high-grade PIN (HGPIN), and prostate cancer tissue from 243 radical prostatectomy patients. Both NPY and MIC-1 showed higher proportional immunostaining in HGPIN and prostate cancer compared with benign epithelium (P < 0.0001). NPY and MIC-1 immunostaining was higher in low-grade PIN compared with other benign tissues (both P < 0.0001) and was equivalent to immunostaining in HGPIN. NPY immunostaining of prostate cancer was independently associated with relapse, after adjusting for traditional prognostic factors, as a categorical variable in 20% intervals (P = 0.0449-0.0103) and as a continuous variable (P = 0.0017). Low MIC-1 immunostaining (20% categories) was associated with pathologic stage >2C after adjusting for predictors of pathologic stage (P = 0.3894-0.0176). This is the first study to show that altered NPY and MIC-1 expression are significantly associated with prostate cancer progression and suggests that these molecules be developed further as biomarkers in the management of prostate disease. (Cancer Epidemiol Biomarkers Prev 2006;15(4):711–6)

Nuclear beta-catenin as a marker of outcome in localized prostate cancer

9570 Background: Beta-catenin, in its role as a nuclear signaling molecule, has been implicated in prostate carcinogenesis through modulation of androgen receptor activity. Mice in whom beta-catenin is overexpressed in the nuclei of prostatic epithelium develop hyperplastic and dysplastic lesions. Hence this study focused on nuclear beta-catenin expression and aimed to define the pattern of beta-catenin protein expression in the nuclei of normal, hyperplastic and malignant human prostate tissue and to determine whether changes in expression were associated with disease progression and/or prognosis. METHODS Five normal prostates, 26 benign prostatic hypertrophy specimens, 232 radical prostatectomy specimens from patients with clinically localized prostate cancer (PC), 186 specimens of hyperplasia adjacent to PC and 20 cases of advanced PC were assessed for beta-catenin expression using immunohistochemistry. RESULTS There were significantly lower levels of nuclear beta-catenin expression in localized PC compared to benign prostate tissue (p<0.001). Nuclear beta-catenin expression was also lower in advanced compared to localized PC (p<0.001). In addition, lower levels of nuclear beta-catenin expression (immunoreactivity in <10% of malignant cells) predicted for a shorter biochemical relapse-free survival in patients with localized PC (HR 1.9, 95%CI 1.2-3.0; p=0.008) and were inversely correlated with pre-operative prostate-specific antigen (PSA) levels (p=0.01). Analysis of a low-risk subgroup of patients with pre-operative PSA levels <10 ng/ml (n=114), demonstrated that lower levels of nuclear beta-catenin (<10% of malignant cells) again predicted for a poorer prognosis (HR 3.2, 95%CI 1.4-7.3; p=0.006). CONCLUSIONS Lower levels of nuclear beta-catenin expression are found in malignant compared to benign prostate tissue. Lower nuclear beta-catenin expression is also associated with a poorer prognosis in localized PC, in particular in the low-risk subgroup of patients with pre-operative PSA levels <10 ng/ml. Consequently, nuclear beta-catenin expression may be of clinical utility as a pre-operative prognostic marker in patients with low-risk PC. No significant financial relationships to disclose.