Pharmacological ascorbic acid suppresses syngeneic tumor growth and metastases in hormone-refractory prostate cancer.

AIM: The aim of this study was to test for the influence of ascorbic acid on tumorigenicity and metastases of implanted PAIII prostate cancer adenocarcinoma cells in syngeneic LW rats. MATERIALS AND METHODS: Hormone-refractory prostate cancer PAIII cells were implanted subcutaneously into immunologically intact, Lobund-Wistar (LW) rats. Intraperitoneal pharmacological doses of ascorbic acid were administered each day for the ensuing 30 days. On the 40th day, animals were sacrificed. Local tumor weights were measured, and metastases were counted. RESULTS: At the end of the 40 day experimental period, the primary tumors were found to be significantly reduced in weight (p=0.026). In addition, sub-pleural lung metastases were even more profoundly reduced in number and size (p=0.009). Grossly enlarged ipsilateral lymph node metastases declined from 7 of 15 rats to 1 of 15 rats. CONCLUSION: Pharmacological doses of ascorbic acid suppress tumor growth and metastases in hormone-refractory prostate cancer.

Rat prostate tumors express cancer procoagulant, an activator of coagulation factor X.

Two common procoagulant activities associated with tumors are tissue factor and cancer procoagulant (CP), an activator of coagulation factor X. We have identified a convenient source of CP in transplanted Lobund-Wistar rat PA3 prostate tumors. CP activity was purified from 5 independent transplanted prostate tumors by column chromatography. The protein activated factor X in the absence of TF and factor VII. An antihuman CP antibody recognized rat CP in an ELISA and inactivated CP activity in a chromogenic assay. Lobund-Wistar prostate tumors may provide a convenient animal model useful in determining the role of CP in cancer development.

Dietary prevention of hormone refractory prostate cancer in Lobund-Wistar rats: a review of studies in a relevant animal model.

Lobund-Wistar (LW) rats, which have high testosterone levels, are predisposed to develop hormone-refractory prostate cancer (HRPC) spontaneously and by methylnitrosourea (MNU) induction, and the development of HRPC progresses through 2 stages. This paper reviews several studies in which LW rats were placed on soy-containing diets and were evaluated for development of either spontaneous or MNU-induced prostate cancer. The premalignant, testosterone-dependent stage is inhibited by testosterone deprivation. In the absence of testosterone deprivation, tumorigenesis progresses spontaneously to the testosterone-independent refractory stage. In LW rats: moderate caloric restriction prevented development of spontaneous prostate cancer; dietary 4-hydroxyphenylretinamide prevented MNU-induced prostate cancer; and dietary supplementation with soy protein isolate with high isoflavones prevented spontaneous and induced tumors and led to moderate reduction of serum testosterone. In rats 12 mo of age and younger, changing from the control diet to the soy+isoflavone diet significantly prevented progression of spontaneous tumors to the refractory stage of disease. Tumors that developed spontaneously and after MNU induction showed similar developmental stages and morphology, but MNU-induced tumors had shorter latency periods before development. The accumulated data indicate that soy-based diets are effective in the prevention of prostate cancer.

Hormone-refractory prostate cancer in the Lobund-Wistar rat.

Research on cancer prevention and therapy must focus on the refractory disease, the fatal end-stage of cancer that develops in patients with organ-related solid tumors. Refractory cancers develop spontaneously in advanced-stage tumors or in relapsed cases after failed therapy. Because neither prevention nor therapy is currently feasible, refractory cancer is a major impediment to survival. There is a great need for an animal model of prostate cancer (PC), one that develops cancer from initial premalignant to the terminal refractory stages. We describe here a model of hormone-refractory prostate cancer (HRPC) that develops spontaneously through two stages by endogenous mechanisms in the Lobund-Wistar (LW) rat. The early premalignant, testosterone (T)-dependent stage is promoted by high levels of endogenous T, and up to age 12 months is reversible by T deprivation; without this intervention, the tumorigenic process progresses to the refractory stage, which is highly aggressive and does not respond to T deprivation or to a wide range of therapies. Initial refractory tumors are palpable at approximately 18 months of age. As they continue to grow, the tumors express characteristics seen in refractory cancers in humans (i.e., hypoxia, expression of hypoxia-inducible factors, and metastasis). Chemically induced HRPCs in LW rats manifest the same two developmental stages, but with shorter latency periods. A transplantable, metastasizing cell line (PAIII) was derived from a germfree LW rat with advanced-stage cancer. Both spontaneous and chemically induced autochthonous HRPC model systems serve as outstanding models for studies on the prevention and therapy of refractory cancer.

Cross-species global and subset gene expression profiling identifies genes involved in prostate cancer response to selenium.

BACKGROUND: Gene expression technologies have the ability to generate vast amounts of data, yet there often resides only limited resources for subsequent validation studies. This necessitates the ability to perform sorting and prioritization of the output data. Previously described methodologies have used functional pathways or transcriptional regulatory grouping to sort genes for further study. In this paper we demonstrate a comparative genomics based method to leverage data from animal models to prioritize genes for validation. This approach allows one to develop a disease-based focus for the prioritization of gene data, a process that is essential for systems that lack significant functional pathway data yet have defined animal models. This method is made possible through the use of highly controlled spotted cDNA slide production and the use of comparative bioinformatics databases without the use of cross-species slide hybridizations. RESULTS: Using gene expression profiling we have demonstrated a similar whole transcriptome gene expression patterns in prostate cancer cells from human and rat prostate cancer cell lines both at baseline expression levels and after treatment with physiologic concentrations of the proposed chemopreventive agent Selenium. Using both the human PC3 and rat PAII prostate cancer cell lines have gone on to identify a subset of one hundred and fifty-four genes that demonstrate a similar level of differential expression to Selenium treatment in both species. Further analysis and data mining for two genes, the Insulin like Growth Factor Binding protein 3, and Retinoic X Receptor alpha, demonstrates an association with prostate cancer, functional pathway links, and protein-protein interactions that make these genes prime candidates for explaining the mechanism of Selenium's chemopreventive effect in prostate cancer. These genes are subsequently validated by western blots showing Selenium based induction and using tissue microarrays to demonstrate a significant association between downregulated protein expression and tumorigenesis, a process that is the reverse of what is seen in the presence of Selenium. CONCLUSIONS: Thus the outlined process demonstrates similar baseline and selenium induced gene expression profiles between rat and human prostate cancers, and provides a method for identifying testable functional pathways for the action of Selenium's chemopreventive properties in prostate cancer.

The anti-ischemia agent ranolazine promotes the development of intestinal tumors in APC(Min/+) mice.

Ranolazine was shown to improve exercise parameters in patients with chronic angina. It works by switching myocardial energy metabolism from fatty acids to glucose, thus increasing the efficiency of ATP production under hypoxic conditions. Tumors are hypoxic and may also respond to ranolazine. We found that ranolazine caused a dose-dependent increase in tumor number in APC(Min/+) mice, a model of spontaneous intestinal tumorigenesis. Tumors from drug-treated mice were also more dysplastic and invasive than those from untreated mice. These findings have implications for the use of ranolazine in patients with a history of malignant neoplasms or adenomatous polyps.

Vaccination against prostate cancer using a live tissue factor deficient cell line in Lobund-Wistar rats.

Reducing expression of the tissue factor gene in prostate adenocarcinoma cells (PAIII) results in a cell line that, in vivo, mimics the growth of wildtype (wt) PAIII. However, instead of continuing to grow and metastasize as wt PAIII tumors do, tissue factor deficient PAIII (TFD PAIII) masses spontaneously regress after several weeks. Although whole cell vaccines are typically inactivated prior to administration to prevent proliferation within the host, numerous studies have suggested that exposure to live, attenuated, whole tumor cells, and the extracellular microenvironment they recruit, increases immunotherapeutic potential. Here, we provide support for this notion, and a strategy through which to implement it, by demonstrating that subcutaneous vaccinations with the TFD PAIII protect the Lobund-Wistar rat against subsequent wt PAIII cell challenge. TFD PAIII immunized rats suffered significantly less metastasis of wt PAIII challenge tumors compared to unvaccinated naïve controls rats. These results offer the intriguing possibility that the TFD PAIII vaccine is an effective system for the prevention and, possibly, the treatment of prostate cancer.

Prevention of de novo prostate cancer by immunization with tumor-derived vaccines.

Since advanced prostate cancer is difficult to treat, we have chosen a very different approach: the development of vaccines to prevent initial de novo tumor formation. To test the hypothesis that prostate cancer can be prevented by vaccination, Lobund-Wistar (LW) rats were vaccinated subcutaneously with complete Freund's adjuvant (CFA) plus glutaraldehyde-fixed (GFT) whole cell or potassium thiocyanate extract (PTE) preparations derived from in vivo tumors, or with media and CFA (media-vaccinated). Rats were vaccinated each month substituting incomplete Freund's adjuvant for CFA, from age 3 to 12 months, and methylnitrosourea (30 mg/kg) was administered intravenously at 4 months of age. Groups of 30 GFT cell-vaccinated rats showed a 90% reduction, and PTE-vaccinated rats, a 50% reduction in the occurrence of de novo prostate tumors compared with media-vaccinated controls. When splenocytes from vaccinated rats were incubated with tumor cells prior to subcutaneous implantation, PTE-vaccinated rats showed a 80% reduction, and GFT cell-vaccinated rats showed a 40% reduction in the occurrence of tumors, demonstrating a role for the spleen in the protective response. The inflammatory responses in tumors from GFT cell-vaccinated rats and PTE-vaccinated rats were distinguished by an influx of eosinophils compared with the responses in tumors from media-vaccinated rats. These results demonstrate the possibility that prostate cancer can be prevented by immunization with vaccines based on whole tumor-derived vaccines.

Susceptibility of Lobund-Wistar x Copenhagen hybrid rats to autochthonous prostate carcinogenesis.

BACKGROUND: Transplantation of the Dunning R-3327 prostate tumor cell line is a common model of prostate cancer, though the rat strain (Copenhagen) from which the cell line was derived is resistant to spontaneous and chemically induced prostate cancer. METHODS: To determine if susceptibility to chemically induced prostate carcinogenesis can be transferred from the susceptible Lobund-Wistar (LW) rat strain to the resistant Copenhagen (COP) strain, male COP rats and LW x COP hybrids were administered an intravenous dose (30 mg/kg) of methylnitrosourea (MNU) and implanted three times with silastic capsules containing testosterone propionate (25 mg each) at a 2-month interval. Serum was sampled at 3, 6, 9, and 12 months of age and assayed for testosterone. RESULTS: Serum testosterone was significantly but transiently elevated following implantation of capsules with testosterone propionate, but then decreased by 12 months of age to a level which was significantly less (P < or = 0.001) than in LW rats but not significantly different from COP rats. Prostate cancer did not develop in COP rats, but 36% of LW x COP hybrids developed prostate-seminal vesicle tumors which expanded into the dorsolateral lobes within 10 months of MNU administration; this compares to 90% of LW rats which develop such tumors following this same induction protocol. CONCLUSIONS: COP rats lack inherent susceptibility to development of prostate cancer; susceptibility may be transferred from LW rats, or resistance from COP rats, to LW x COP hybrids and is present in the haploid state, consistent with the two-mutation event hypothesis of Knudson which holds that two mutations are required at a genetic locus for development of some cancers.

Using Comparative Genomics to Leverage Animal Models in the Identification of Cancer Genes. Examples in Prostate Cancer.

The identification of cancer biomarkers that will predict susceptibility to disease and subsequent clinical outcome are key components of future genomics-based tailored medical care. Animal models of disease provide a rich resource for the identification of potential cancer biomarkers. Animal models of prostate cancer in particular offer the potential to identify cancer genes associated with dietary and environmental factors. The key issue is the timely and efficient identification of candidate genes that are likely to impact on human prostate cancer. Here, we demonstrate comparative genomics-based methods for the identification of candidate genes in animal models that are associated with human chromosomal regions implicated in prostate cancer. Using publicly available bioinformatics tools, comparisons can be made between cancer-specific datasets, genomic sequencing data and cross-species comparative maps to identify potential cancer biomarkers. This process is demonstrated by using rat models of prostate cancer to identify candidate human prostate cancer genes. Genes identified through these techniques can be screened as biomarkers for response to chemopreventive agents, as well as being used in transgenic or knockout mice to engineer better animal models of human prostate cancer. The bioinformatics techniques outlined here can be used to leverage genomic data from any animal cancer model for use in the study and treatment of human cancer.

Fas ligand expression and its correlation with apoptosis and proliferation in Lobund-Wistar prostate carcinomas.

OBJECTIVE: The Fas (CD95) interaction with its receptor Fas ligand (FasL) is one of the main mechanisms of cell apoptosis. High expression of FasL has been consistently observed in a variety of human cancers. In this study, we evaluated FasL and its relationship with apoptosis and proliferation in Lobund-Wistar (L-W) cancers. The L-W rat strain develops spontaneous and induced adenocarcinomas in the anterior prostate and seminal vesicles. Although FasL expression has been observed in a subset of human prostate carcinomas, this multistage model allowed in vivo evaluation of subclones of malignant cells with a single genetic susceptibility. METHODS: Apoptosis was evaluated in spontaneous, induced and transplanted tumors as well as metastasis using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) technique and transmission electron microscopy. Proliferating cell nuclear antigen (PCNA) and FasL expression were detected using immunohistochemistry and analyzed according to the number of positive cells and intensity of staining using a semiquantitive method. RESULTS: Apoptotic indexes were significantly higher in spontaneous tumors compared to induced (p < 0.008), transplanted tumors (p < 0.0112) and metastases (p < 0.009). TUNEL-positive cells were frequently observed in the leukocytic infiltrate of the stroma in transplanted carcinomas and metastases. These findings were confirmed by electron microscopy. FasL expression was not uniformly localized in L-W carcinomas and its highest expression was observed in transplanted tumors and metastasis (p < 0.005). Moreover, PCNA indices were directly correlated with cancers showing high FasL total scores (Hscores). CONCLUSIONS: In this model, high FasL expression was associated with cells displaying low apoptotic indexes and high PCNA index. Therefore, analysis of FasL may have clinical relevance in detecting the malignant potential of prostate cancers.