Histological grade, perineural infiltration, tumour-infiltrating lymphocytes and apoptosis as determinants of long-term prognosis in prostatic adenocarcinoma.

A series of 325 prostatic adenocarcinomas with a long-term clinical follow-up were subjected to light microscopic analysis of histological prognostic factors, including three grading systems (Gleason score, WHO grade, nuclear grade), perineural infiltration of the tumour (PNI), tumour infiltrating lymphocytes (TIL) and the presence of apoptotic cells (APO). All three histological classifications correlated significantly with the prognosis, but in multivariate analysis, the Gleason score was superior to the WHO grading or nuclear grading in predicting patient survival. PNI was significantly related to poor differentiation of the tumour, its progression and ominous disease outcome, particularly in T1-2MO tumours. The density of TIL was independent of the tumour differentiation, and absent or weak TIL were signs of a high risk of tumour progression and of a fatal disease. Apoptotic cells were commonly detected in poorly differentiated tumours and apoptosis was related to disease progression and low survival probability. The results suggest that the Gleason score, PNI and the density of TIL should be included in routine pathology reports, to be used by clinicians while making therapeutic decisions in prostatic cancer.

Expression of retinoblastoma gene (Rb) protein in T12M0 prostatic adenocarcinoma.

The expression of Rb protein was analysed by immunocytochemical methods in 118 patients with T12M0 prostatic adenocarcinoma who were followed-up for more than 12 years. Rb protein was expressed in 117/118 (99%) tumours. The fraction of positive nuclei was (mean +/- SD, 92 +/- 19%) and only one tumour was completely negative for Rb protein. Abnormal expression of the Rb protein (in fewer than 90% of cells) was independent of the T category while there was a significant relationship between DNA ploidy (P = 0.0138), S-phase fraction (P = 0.0042) and expression of the Rb protein. Abnormal expression of the Rb protein had no prognostic value while T category (P = 0.0017), Gleason score (P = 0.0097), DNA ploidy (P = 0.0034), S-phase fraction (P = 0.0179) and mitotic index (P = 0.0001) were significant prognostic factors. In multivariate analysis the only independent predictor was the mitotic index [Risk ratio (RR) (95%CI) = 7.4(2.4-22.5), P = 0.004]. The results show that Rb protein immunohistochemistry has hardly any prognostic significance in prostatic adenocarcinoma whereas direct measurement of cell proliferative activity contains significant prognostic information.

A prognostic score for prostatic adenocarcinoma based on clinical, histological, biochemical and cytometric data from the primary tumour.

The aim of this study was to create a multivariate prognostic score for prostatic adenocarcinoma. A retrospective analysis of clinical, histological and cytometric prognostic factors in 325 cases of prostatic adenocarcinoma followed up on average over 13 years was performed. A multivariate prognostic score was built by using the independent prognostic factors. M-category, T-category, Gleason score and patient age were independent prognostic factors in the entire cohort. In MO tumors T-category, density of tumor infiltrating lymphocytes, the presence of apoptotic cells, and patient age were independent prognostic factors. In T1-2MO tumors the density of tumour infiltrating lymphocytes, mitotic index, standard deviation of maximum nuclear diameter and the serum level of acidic phosphatase were independent prognostic factors. By combining the coefficients of the regression model, a prognostic score was built for each of the tumour categories. The subsequent survival analysis based on the prognostic score indicated that it is a highly significant prognostic factor superior to individual prognostic parameters. The results show that the combination of prognostic data is a valuable tool in assessing the correct prognostic category for prostatic adenocarcinoma.

Tumor vascularity and basement membrane structure as prognostic factors in T1-2MO prostatic adenocarcinoma.

The basement membrane (BM) structure and vascular density (VD) were analysed using collagen IV immunostaining in a series of 88 T1-2MO prostatic adenocarcinomas. The results were correlated with the established prognostic factors and patient survival based on a follow-up of > 11 years. Prostatic cancer cells retained their capacity to synthesize BM components, although defects in the BM structures were associated with T2 category and high Gleason score. Progression of the tumours was related to the defects in BM continuity, and defective BMs were also a sign of poor outcome. High grade tumours were occupied by dense vascular network and dense VD was a sign of a poor outcome. Gleason score was a significant predictor of progression and patient survival, as was the T-category. Multivariate analysis of the prognostic factors indicated that Gleason score included all the available independent prognostic information. The results show that all prostatic adenocarcinomas continue to synthesize BM components, although the structure and continuity is disturbed in poorly differentiated tumours. A rich vascular network is a sign of highly malignant phenotype in prostatic adenocarcinoma. However, an assessment of the BM and VD is of limited independent prognostic value.

Results of the primary treatment in T1-3M0 prostatic adenocarcinoma are dependent on tumour biology.

The results of the primary treatment were analysed in relation to biological prognostic factors in a series of 215 males with T1-3M0 prostatic adenocarcinoma. T-category, Gleason score and DNA ploidy were significantly related to progression-free survival and overall survival. Incidental tumours usually had a favorable prognosis, 86% of them being diploid, whereas the incidental aneuploid tumours (14%) ran an unfavorable course. In clinically malignant prostates (TPR), neither Gleason score nor DNA ploidy had prognostic significance. The treatment had prognostic significance only in diploid tumours or in tumours with Gleason score value < or = 7, while aneuploid tumours or tumours with Gleason score value > 7 always had unfavorable prognosis irrespective of treatment. Tumours treated by radical prostatectomy or by radical radiation treatment were associated with a favorable prognosis, as contrasted to the tumours treated by endocrine treatment or those followed-up only, which had a high rate of mortality. Transurethral resection was weakly related to increased risk of death from prostatic adenocarcinoma, although the basic mechanisms behind this observation remain to be explained.

Expression of the apoptosis suppressing protein bcl-2 in prostatic adenocarcinoma is related to tumor malignancy.

BACKGROUND: The expression of bcl-2 protein has been related to histopathological features and prognosis in several epithelial tumors. In prostatic adenocarcinoma the prognostic significance of bcl-2 expression is mainly unexplored. METHODS: The expression of bcl-2 protein was assessed in 235 prostatic adenocarcinomas by using monoclonal bcl-2 protein antibody after microwave pretreatment of the cancer sections. The results of immunohistochemistry were related to histopathological features and prognosis of the patients. RESULTS: 71% of the tumors were bcl-2 negative, in 18% of cases the expression was weak, and in 11% of cases strong. The fraction of bcl-2 positive cells was variable (mean [SE]: 15 [2]%). The expression of bcl-2 protein was positively correlated to high T-category, metastatic disease, and poor histological differentiation of the tumor. Tumors that were aneuploid and rapidly proliferating frequently expressed bcl-2 protein, while tumors we densely infiltrated by inflammatory cells rarely expressed bcl-2. The expression of bcl-2 protein was related to lowered survival probability in univariate analysis, while in multivariate analysis the expression of bcl-2 had no independent prognostic value. CONCLUSIONS: The expression of bcl-2 protein in prostatic adenocarcinoma is related to tumor malignancy, but the prognostic significance of the expression requires further analyses, particularly in localized tumors.

Mitotic activity and prognosis in prostatic adenocarcinoma.

Mitotic figures were quantitated by two different methods (number of mitotic figures in 10 high power fields, MI; number of mitotic figures/mm2 of neoplastic epithelium, M/V) in a series of 303 prostatic adenocarcinomas, and the results were related to clinical and histological features as well as to prognosis. Gleason score and mitotic indices were significantly interrelated. Invasive T3-4 tumors showed higher mitosis counts than did the local ones, and metastasis at the time of diagnosis was related to mitotic indices as well. Progression and progression-free survival were related significantly to Gleason score and mitotic indices. In univariate survival analysis, T-category, M-category, Gleason score, MI, and M/V were significant prognostic factors. In multivariate analysis, independent predictors of survival were M-category, T-category, Gleason score, patient age, and the M/V index. If the Gleason score was not included, M/V included all the available prognostic information in T1-2M0 carcinomas. The results show that, in addition to the conventional Gleason grading system, mitotic indices are useful prognostic parameters while evaluating the long-term prognosis in prostatic adenocarcinoma.

Biochemical parameters as prognostic factors in prostatic adenocarcinoma.

The serum levels of creatinine (CR), alkaline phosphatase (ALP), acid phosphatase (ACP) and tartrate inhibitable acid phosphatase (TIAP) were related to Gleason score, TM-category, disease progression and survival in 325 prostatic adenocarcinoma patients followed up for over 12 years. Elevated serum levels of CR, ALP, ACP and TIAP were related to invasive and metastatic disease as well as with a high Gleason score. Elevated serum levels of CR, ALP, ACP and TIAP, all significantly predicted prognosis in a univariate analysis. In the M0 tumours, ACP and TIAP and TIAP had prognostic value, as they did in the T1-2M0 tumours respectively. Cox's multivariate analysis showed that serum creatinine level at diagnosis had independent prognostic value additional to the TM-classification, Gleason score and patient age. In the M0 tumours, ALP had independent prognostic significance additional to the T-category, Gleason score and patient age. In the T1-2M0 tumours, TIAP had independent prognostic value supplementary to the Gleason score, T-category and patient age, whereas in the T1M0 tumours, the gleason score was an independent prognostic parameter. The results indicate that these simple laboratory tests give important prognostic information in prostatic adenocarcinoma.

Value of Ki-67 immunolabelling as a prognostic factor in prostate cancer.

OBJECTIVE(S): The aim of the study was to analyze the results of Ki-67 immunostaining in prostatic adenocarcinoma and to assess its prognostic value. METHODS: Clinical follow-up data was reviewed in 190 prostatic adenocarcinomas and the results of Ki-67 immunolabelling were correlated to standard prognostic factors and survival data of the patients. All stage and grade categories were included. RESULTS: Ki-67 expression correlated significantly with histological differentiation of tumors, indicators of cell proliferation, perineural growth, density of tumor-infiltrating lymphocytes and TM classification. In survival analysis, Ki-67 expression was able to discriminate patients into different prognostic groups (p = 0.0035). In a separate analysis including T1-2M0 tumors, Ki-67 immunolabelling was a significant predictor of survival (p = 0.0258). In Cox's multivariate analysis Ki-67 was an independent prognostic factor in the entire cohort. CONCLUSIONS: The results show that Ki-67 expression is a potentially useful prognostic factor in prostatic adenocarcinoma and it could be used as an additional criterion in defining a correct prognostic category in this malignancy.

Effects of some putative amino acid neurotransmitters on the stimulated TSH secretion in male rats.

The importance of several amino acids (glycine, L-glutamic acid, L-serine, taurine and beta-alanine) in the regulation of the stimulated secretion of TSH was studied in male rats using both peripheral and central administration of the amino acids. Glycine (10-200 mg/kg i.p.), L-glutamic acid (10-500 mg/kg i.p.) and L-serine (500 mg/kg i.p.) decreased significantly the cold-induced TSH secretion whereas beta-alanine (1-500 mg/kg i.p.) and taurine (10-100 mg/kg i.p.) were not effective. The effect of L-glutamic acid (100 mg i.p.) was partially antagonized by bicuculline (1 mg/kg i.p.) but not by picrotoxin (1 or 2 mg/kg i.p.). Only glycine (50 and 100 mg/kg i.p.) inhibited the TRH-stimulated TSH secretion. When the intracerebroventricular route was used, L-serine (50 micrograms/rat) decreased the TSH could response whereas glycine and L-glutamic acid (1-50 micrograms/rat) had no clear effect. We conclude that glycine, glutamate and serine inhibit the cold-induced TSH secretion in the male rat. The action of serine and glycine is possibly mediated through the periventricular hypothalamus and the anterior pituitary, respectively. The inhibition caused by glutamate seems to be partially mediated through the bicuculline-sensitive GABA receptors in the hypothalamus. Taurine and beta-alanine play no role in the control of rat TSH secretion.

Progression and survival in prostatic adenocarcinoma: a comparison of clinical stage, Gleason grade, S-phase fraction and DNA ploidy.

Clinical data were reviewed in 325 patients with prostatic adenocarcinoma followed up for a mean of 13 years. Paraffin-embedded tumour biopsy specimens from the primary tumours were available for flow cytometry (FCM) in 273 cases. Intra-tumour heterogeneity in DNA index (DI) was found in 4% of the tumours (54 cases were analysed). S-phase fraction (SPF) and DNA ploidy were significantly interrelated. Aneuploidy and high SPF were significantly related to both a high T category and high Gleason score. The progression in T1-2M0 tumours was related to Gleason score (P = 0.009), DNA ploidy (P = 0.006) and SPF (P = 0.007), while the Gleason score (P = 0.0013), DNA ploidy (P = 0.002) and SPF (P < 0.001) had prognostic value in univariate survival analysis. In the entire cohort, the T category (P < 0.001), M category (P < 0.001), Gleason score (P < 0.001), DNA ploidy (P < 0.001) and SPF (P < 0.001) were significant prognostic factors. In Cox's analysis, the M category (P < 0.001), Gleason score (P < 0.001), T category (P = 0.003), age (P = 0.001) and SPF (P = 0.087) were independently related to prognosis. In the T1-2M0 tumours, Gleason score (P < 0.001), T category (P = 0.022) and SPF (P = 0.058) were independent predictors. A novel classification system in which the DNA ploidy or SPF and the Gleason score were combined was found to be of significant prognostic value in all M0 tumours (P < 0.001). The results suggest that FCM can be used as an adjunct to conventional histological assessments for determination of the correct prognostic category in prostatic adenocarcinoma.

Measurement of the complex between prostate-specific antigen and alpha1-protease inhibitor in serum.

BACKGROUND: Prostate-specific antigen (PSA) occurs in serum both free and in complex with protease inhibitors. The complex with alpha1-antichymotrypsin (ACT) is the major form in serum, and the proportion of PSA-ACT is higher in prostate cancer (PCa) than in benign prostatic hyperplasia (BPH). PSA also forms a complex with alpha1-protease inhibitor (API) in vitro, and the PSA-ACT complex has been detected in serum from patients with prostate cancer. The aim of the present study was to develop a quantitative method for the determination of PSA-API and to determine the serum concentrations in patients with PCa and BPH. METHODS: The assay for PSA-API utilizes a monoclonal antibody to PSA as capture and a polyclonal antibody to API labeled with a Eu-chelate as a tracer. For calibrators, PSA-API formed in vitro was used. Serum samples were obtained before treatment from 82 patients with PCa, from 66 patients with BPH, and from 22 healthy females. RESULTS: The concentrations of PSA-API are proportional to the concentrations of total PSA. PSA-API comprises 1.0-7.9% (median, 2.4%) of total immunoreactive PSA in PCa and 1.3-12.2% (median, 3.6%) in BPH patients with serum PSA concentrations >4 microgram/L. In patients with 4-20 microgram/L total PSA, the proportion of PSA-API serum is significantly higher in BPH (median, 4.1%) than in PCa (median, 3. 2%; P = 0.02). CONCLUSIONS: The proportion of PSA-API in serum is lower in patients with PCa than in those with BPH. These results suggest that PSA-API is a potential adjunct to total and free PSA in the diagnosis of prostate cancer.

Nuclear morphometry is of independent prognostic value only in T1 prostatic adenocarcinomas.

A series of 325 patients with prostatic adenocarcinoma followed-up for over 13 years was subjected to histomorphometric analysis for the following prognostic factors: the Gleason score and 10 nuclear morphometric factors (mean nuclear area, nuclear perimeter, shortest and longest nuclear axis, form factor and their SDs), using the IBAS 1&2 image analyzer. Nuclear factors, Gleason score (P = 0.0013-0.0148), and T-category (P = 0.004-0.0107) were significantly interrelated, while the M-category was independent of the morphometric parameters. Nuclear factors (P = 0.0003-0.5), Gleason score (P < 0.0001), T-category (P < 0.0001) and M-category (P < 0.0001) predicted the disease outcome. In T1-2M0 tumors, the T-category (P = 0.0001), Gleason score (P < 0.0001), SD of nuclear area (P = 0.057), SD of nuclear perimetry (P = 0.039), mean shortest nuclear axis (P = 0.030), SD of the shortest nuclear axis (P = 0.0045), SD of the longest nuclear axis (P = 0.009), and nuclear form factor (P = 0.0123) were significant prognostic factors. In the multivariate survival analysis, the nuclear area had independent prognostic significance only in T1 tumors, whereas in other subcategories, the clinical stage, Gleason score, and patient age included all the available prognostic information. The results indicate that nuclear morphometric measurements are of borderline significance only in evaluating the intrinsic malignancy of prostatic adenocarcinoma.

Characterization and immunological determination of the complex between prostate-specific antigen and alpha2-macroglobulin.

Prostate-specific antigen (PSA) rapidly forms a complex with alpha2-macroglobulin (A2M) in vitro; however, PSA complexed with A2M (PSA-A2M) is not detected by conventional immunoassays for PSA because it is encapsulated by the A2M. In this study, we show that denaturation of PSA-A2M at high pH renders PSA immunoreactive. Part of the complexed PSA is released in free form and part remains bound to denatured A2M. These forms can be measured by a conventional immunoassay for PSA. This finding enabled us to design a dissociation assay for the detection of PSA-A2M, which was based on the removal of immunoreactive PSA in serum by immunoadsorption, denaturation of PSA-A2M at high pH, and measurement of the released PSA immunoreactivity by a conventional PSA immunoassay. This PSA-A2M assay was calibrated with PSA-A2M formed in vitro. The detection limit of the assay was 0.14 microg/L. Inter- and intraassay coefficients variation were 4-9% and 8-14%, respectively. When purified PSA was incubated with A2M, the loss of PSA immunoreactivity was highly correlated with the PSA-A2M formed, as measured by the dissociation assay for PSA-A2M (r = 0.99; P <0.0001). The concentration of PSA-A2M in serum correlated with that of total PSA both in prostate cancer (PCa) and benign prostatic hyperplasia (BPH); however, the ratio of PSA-A2M in relation to total PSA was significantly higher in BPH than in PCa (P <0.0003). ROC curve analysis suggested that measurement of the ratio of PSA-A2M to total PSA in serum improves the diagnostic accuracy for PCa compared with assays for total PSA only.

Quantification of prostate specific antigen mRNA levels in circulation after prostatic surgery and endocrine treatment by quantitative reverse transcription-polymerase chain reaction.

Various methods to detect prostatic cells in circulation have given conflicting results. This is probably because qualitative rather than quantitative methods have been used to detect mRNA from prostatic cells. A quantitative method has been developed based on reverse transcription-polymerase chain reaction (RT-PCR) for detection of prostate specific antigen (PSA) mRNA in peripheral blood. A competitive internal mRNA standard was used for quantification of absolute amounts of PSA mRNA. The detection limit of the assay was 7 copies of mRNA, and the highest level of circulating PSA mRNA in 88 control subjects was 25 copies per milliliter of blood. This method was used to study the influence of prostatic surgery and endocrine treatment on prostatic cells in the circulation of 56 patients undergoing biopsy, radical prostatectomy, transurethral resection of the prostate (TURP), orchiectomy, or androgen blockade. Blood samples were drawn before, during and up to 26 weeks after these procedures had been carried out. The highest level of PSA mRNA in controls was 25 copies per milliliter of blood. After RP, TURP or orchiectomy, PSA mRNA levels increased above this level in 27%, 29%, and 25% of the samples, respectively. After prostate biopsy, two out of 15 patients became positive. PSA mRNA levels that were elevated by surgery became undetectable within 1-3 days. No significant correlation was found between PCR positivity and the clinical characteristics of the patients. It is concluded that the level of PSA mRNA in peripheral blood increases after prostatic surgery, indicating temporary dissemination of prostatic cells. However, preoperative levels do not correlate with serum PSA, stage or grade.

Proliferating cell nuclear antigen and p53 expression as prognostic factors in T1-2M0 prostatic adenocarcinoma.

The expression of proliferating cell nuclear antigen and p53 protein was analysed by immunocytochemical methods (PC10, CM1 antisera) in 139 patients with T1-2M0 prostatic adenocarcinomas followed-up for > 12 years. p53 protein was expressed in 21 (15%) tumours (15%), the fraction of positive nuclei being very low (mean SE, 1% +/- 0.7%). Accumulation of p53 protein in epithelial cells was independent of tumour stage and Gleason score, and had no effect on prognosis. In 4 cases, p53 protein was expressed only in stromal cells. The fraction of PCNA-positive nuclei (evaluable in 116 cases) was higher in T2 than in T1 tumours (p < 0.001); furthermore, high Gleason score was positively correlated with PCNA positivity (p < 0.001). A finding of over 5% of PCNA-positive nuclei predicted progression in T and M categories and were a sign of poor outcome. The fraction of PCNA-positive stromal-cell nuclei was related to T-category with a borderline significance (p = 0.06). In a multivariate analysis of the prognostic factors, independent predictors of survival included Gleason score (p < 0.001), fraction of PCNA-positive nuclei (p = 0.013), observation before therapy (p = 0.05), and T-category (p = 0.07) in that order of significance. The results suggest that overexpression of p53 protein is of marginal prognostic value in local prostatic adenocarcinomas, whereas direct measurement of cell proliferation by PCNA immunolabelling provides important prognostic information in T1-2M0 tumours, in addition to the Gleason score.