Enforced epithelial expression of IGF-1 causes hyperplastic prostate growth while negative selection is requisite for spontaneous metastogenesis.

The insulin-like growth factor-1 (IGF-1) signaling axis is important for cell growth, differentiation and survival and increased serum IGF is a risk factor for prostate and other cancers. To study IGF-1 action on the prostate, we created transgenic (PB-Des) mice that specifically express human IGF-1(des) in prostate epithelial cells. This encodes a mature isoform of IGF-1 with decreased affinity for IGF binding proteins (IGFBP) due to a 3-amino acid deletion in the N terminus. Expression of IGF-1(des) was sufficient to cause hyperplastic lesions in all mice, however the well-differentiated lesions did not progress to adenocarcinoma within a year. Remarkably, crossing the PB-Des mice to an established model of prostate cancer delayed progression of organ-confined tumors and emergence of metastatic lesions in young mice. While dissemination of metastatic lesions was widespread in old bigenic mice we did not detect IGF-1(des) in poorly differentiated primary tumors or metastatic lesions. Expression of endogenous IGF-1 and levels of P-Akt and P-Erk were reduced independent of age. These data suggest that increased physiologic levels of IGF-1 facilitate the emergence of hyperplastic lesions while imposing a strong IGF-1-dependent differentiation block. Selection against IGF-1 action appears requisite for progression of localized disease and metastogenesis.

Conditional activation of FGFR1 in the prostate epithelium induces angiogenesis with concomitant differential regulation of Ang-1 and Ang-2.

The expression of fibroblast growth factor receptor (FGFR)-1 correlates with angiogenesis and is associated with prostate cancer (CaP) progression. To more precisely define the molecular mechanisms whereby FGFR1 causes angiogenesis in the prostate we exploited a transgenic mouse model, JOCK-1, in which activation of a conditional FGFR1 allele in the prostate epithelium caused rapid angiogenesis and progressive hyperplasia. By labeling the vasculature in vivo and applying a novel method to measure the vasculature in three dimensions, we were able to observe a significant increase in vascular volume 1 week after FGFR1 activation. Although vessel volume and branching both continued to increase throughout a 6-week period of FGFR1 activation, importantly, we discovered that continued activation of FGFR1 was not required to maintain the new vasculature. Exploring the molecular mediators of the angiogenic phenotype, we observed consistent upregulation of HIF-1alpha, vascular endothelial growth factor (VEGF) and angiopoietin 2 (Ang-2), whereas expression of Ang-1 was lost. Further analysis revealed that loss of Ang-1 expression occurred in the basal epithelium, whereas the increase in Ang-2 expression occurred in the luminal epithelium. Reporter assays confirmed that the Ang-2 promoter was regulated by FGFR1 signaling and a small molecule inhibitor of FGFR activity, PD173074, could abrogate this response. These findings establish a method to follow spontaneous angiogenesis in a conditional autochthonous system, implicate the angiopoietins as downstream effectors of FGFR1 activation in vivo, and suggest that therapies targeting FGFR1 could be used to inhibit neovascularization during initiation and progression of CaP.

Patterns of protease production during prostate cancer progression: proteomic evidence for cascades in a transgenic model.

Extracellular proteases are recognized as critical factors in the progression of a number of carcinomas, including prostate cancer. Matrix metalloproteases (MMP) are important in processes of tumor growth, invasion and dissemination, but other classes of proteases, such as serine and cysteine proteases, also contribute. We utilized the TRAMP model for prostate cancer to elucidate proteases involved in prostate cancer progression. General proteomic analysis was performed on normal murine prostate, early TRAMP tumors and advanced TRAMP tumors, as well as normal and involved lymph nodes. Zymography and antigenic analyses revealed increased expression of mainly pro-MMP in early TRAMP tumors but substantial elaboration of activated MMP only in late TRAMP tumors. Progressive increase in cysteine, serine and certain membrane-bound proteases from normal to early to advanced prostate tumors, was also seen. Our results implicate pericellular proteases as initiators of major proteolytic cascades during tumor progression and suggest targets for maximal therapeutic effect.

Models of metastatic prostate cancer: a transgenic perspective.

PURPOSE: Transgenic mouse models are proving to be invaluable in our effort to understand the molecular basis of metastatic prostate cancer (CaP). We review and discuss how current animal models have contributed to our understanding of the metastatic cascade and how transgenic technology is being used to develop the next generation of mouse models. Our goal is to provide a review of the recent advances and provide a framework for further studies. MATERIALS AND METHODS: We performed a MEDLINE search of the literature on CaP metastasis transgenic and animal models. RESULTS: We present a summary of the characteristics of nine different animal models of CaP. Each model is unique and provides valuable insight into the molecular mechanisms governing the progression of CaP. Our experience with transgenic models and all the new data from the literature predicts that we will be able to develop genetically engineered mice that accurately mimic the heterogeneity, androgen-independent growth, and metastatic spread seen in clinical disease. CONCLUSION: In order to elucidate the molecular mechanisms of CaP metastasis, it will be necessary to compare gene and protein expression patterns and biochemical analyses of clinical metastatic disease with data obtained from current models. We will also need to refine our ability to engineer and characterize genetic perturbation models. This type of integrative and iterative approach should facilitate better understanding of the molecular biology of CaP metastases.

Genistein in the diet reduces the incidence of poorly differentiated prostatic adenocarcinoma in transgenic mice (TRAMP).

Latent prostate tumors are commonly found with similar frequency in many countries and ethnic groups. In contrast, aggressive prostate cancer (PC) is significantly less prevalent among Asian men, where the intake of soy products is very high. High consumption of foods containing soy results in high plasma, urine, and prostatic fluid concentrations of phytoestrogens, including genistein. The objective of the present study was to test the hypothesis that dietary genistein might prevent PC progression in a transgenic mouse model of PC (TRAMP). By 28-30 weeks of age, all TRAMP mice in the study had developed prostate tumors, with about half of them displaying well-differentiated prostatic adenocarcinoma (WD, score 4), and the other half divided between moderately differentiated (MD, score 5) and poorly differentiated adenocarcinoma (PD, score 6). Two lines of evidence supported the possibility that prostates with PD may represent a more advanced stage of PC in TRAMP mice: (a) the weight of prostates with PD was two orders of magnitude higher than that of prostates with WD or MD; and (b) expression of androgen receptor transcripts was altered in PD as compared with WD and MD. To test the potential of genistein to prevent the incidence of mice with PD, starting at 5-6 weeks of age, transgenic males were fed a phytoestrogen-free diet (AIN-76A) containing 0, 100, 250, or 500 mg of genistein per kg AIN-76A (25, 10, 17, and 7 mice/group, respectively). Mice were on the diet until they were 28-30 weeks of age. Each mouse was weighed once a week throughout the study. At necropsy, selected organs were weighed and prepared for histopathological evaluation. Serum levels of genistein in mice on diet containing 0, 250, or 500 mg of genistein per kg AIN-76A were 52.4 +/- 32.7, 138.9 +/- 69.6, and 397.3 +/- 104.9 nM, respectively, comparable with those found in Asian men on regular soy diet (276 nM). Using body and organ weights as indicators, dietary genistein had no toxic effect on TRAMP mice. The percentage of transgenic males that developed PD was reduced in a dose-dependent manner by dietary genistein.

Down-regulation of prostasin serine protease: a potential invasion suppressor in prostate cancer.

BACKGROUND: Prostasin is a serine protease predominantly expressed in normal prostate epithelial cells. The biological function of prostasin has not been determined. METHODS: Western blot and RT-PCR analyses were used to examine the expression of prostasin in prostate cancer cell lines. Immunohistochemistry was used to evaluate prostasin protein expression in human prostate cancer. An in vitro Matrigel invasion assay was used to test the invasiveness of prostate cancer cell lines forced to express recombinant prostasin. RESULTS: Both prostasin protein and mRNA were found to be expressed in normal human prostate epithelial cells and a non-invasive human prostate cancer cell line, the LNCaP, but neither was found in invasive human prostate cancer cell lines DU-145 and PC-3. Prostasin mRNA expression was absent in invasive prostate cancer cell lines of a transgenic mouse model. Immunohistochemistry analysis showed that prostasin protein expression is down-regulated in high-grade prostate cancer. Transfection of DU-145 and PC-3 cells with a full-length human prostasin cDNA restored prostasin expression and reduced the in vitro invasiveness by 68 and 42%, respectively. CONCLUSIONS: Our data indicate that prostasin may be implicated in normal prostate biology and is able to suppress prostate cancer invasion in vitro.

Autochthonous mouse models for prostate cancer: past, present and future.

Prostate cancer continues to be the second leading cancer related death among men. In order to more fully develop effective prevention and intervention strategies for this prevalent disease, the underlying molecular mechanisms of initiation, progression and metastatic spread must be understood. To this end mouse models have an essential role in prostate cancer research in that they can closely mimic the pathological and biochemical features of the disease. In this review we discuss the history of autochthonous murine models of prostate cancer, the essentials of the idealized mouse model for prostate cancer and speculate on approaches towards this goal.

Collocation of androgen receptor gene mutations in prostate cancer.

Consistent with both the development of the normal prostate gland and prostate tumorigenesis being dependent on testicular androgens, targeting the androgen-signaling axis (i.e., androgen ablation therapy) remains the predominant treatment regime for patients with metastatic prostate cancer. Although there is a very good initial response to androgen ablation, these treatments are essentially palliative. Recent evidence suggests that treatment failure may not result from a loss of androgen signaling but, rather, from the acquisition of genetic changes that lead to aberrant activation of the androgen-signaling axis. A consistent finding is that androgen receptor (AR) gene mutations, present in metastatic prostate cancer and in human prostate cancer cell lines as well as in xenograft and other animal models, result in decreased specificity of ligand-binding and inappropriate receptor activation by estrogens, progestins, adrenal androgens, glucocorticoids and/or AR antagonists. Because a significant proportion of missense mutations in the AR gene reported in prostate cancer collocate to the signature sequence and AF-2, two discrete regions of the ligand-binding domain critical for androgen signaling, we recently proposed that collocation of mutations identified in prostate cancer would identify additional regions of the AR important in receptor function. This approach led to the identification of a four-amino acid region at the boundary of the hinge and ligand-binding domains of the receptor that forms half of a potential protein-protein binding site. AR gene mutations have also been identified that collocate to areas in the DNA-binding domain, to the NH(2)-terminal transactivation domain, and to the hinge region in prostate tumors. In nearly every case, missense mutations in the AR gene identified in prostate cancer that collocate to discrete regions of the receptor contribute to altered androgen signaling and provide a potential mechanism to explain the reemergence of tumor growth during the course of hormone ablation therapies.

Angiogenesis and prostate cancer: identification of a molecular progression switch.

To elucidate the sequence of molecular events intricate with angiogenesis and the initiation and progression prostate cancer, the temporal and spatial expression patterns of platelet endothelial cell adhesion molecule-1 (PECAM1/CD31), hypoxia-induced factor-1alpha (HIF-1alpha), vascular endothelial growth factor (VEGF), and the cognate receptors VEGFR1 and VEGFR2 were characterized. Immunohistochemical and in situ analyses of prostate tissue specimens derived from the spontaneous autochthonous transgenic adenocarcinoma of the mouse prostate (TRAMP) model identified a distinct early angiogenic switch consistent with the expression of PECAM-1, HIF-1alpha, and VEGFR1 and the recruitment of new vasculature to lesions representative of high-grade prostatic epithelial neoplasia (PIN). During progression of prostate cancer, the intraductal microvessel density (IMVD) was also observed to increase as a function of tumor grade. Immunoblot and in situ analyses further demonstrated a distinct late angiogenic switch consistent with decreased expression of VEGFR1, increased expression of VEGFR2, and the transition from a differentiated adenocarcinoma to a more poorly differentiated state. Analysis of clinical prostate cancer specimens validated the predictions of the TRAMP model. This resolution of prostate cancer-associated angiogenesis into distinct early and late molecular events establishes the basis for a "progression-switch" model to explain how the targets of antiangiogenic therapy might change as a function of tumor progression.

Mutations at the boundary of the hinge and ligand binding domain of the androgen receptor confer increased transactivation function.

The androgen receptor (AR), a member of the steroid receptor superfamily of nuclear transcription factors, mediates androgen signaling in diverse target tissues. Here we report AR gene mutations identified in human prostate cancer and the autochthonous transgenic adenocarcinoma of the mouse prostate model that colocate to residues (668)QPIF(671) at the boundary of the hinge and ligand-binding domain, resulting in receptors that exhibit 2- to 4-fold increased activity compared with wild-type AR in response to dihydrotestosterone, estradiol, progesterone, adrenal androgens, and the AR antagonist, hydroxyflutamide, without an apparent effect on receptor levels, ligand binding kinetics, or DNA binding. The expression of these or similar variants could explain the emergence of hormone refractory disease in a subset of patients. Homology modeling indicates that amino acid residues (668)QPIF(671) form a ridge bordering a potential protein-protein interaction surface. The naturally occurring AR gene mutations reported in this study result in decreased hydrophobicity of this surface, suggesting that altered receptor-protein interaction mediates the precocious activity of the AR variants.

Hormone status selects for spontaneous somatic androgen receptor variants that demonstrate specific ligand and cofactor dependent activities in autochthonous prostate cancer.

We have used the autochthonous transgenic adenocarcinoma of mouse prostate (TRAMP) model to investigate the relationship between somatic mutation in the androgen receptor (AR) and the emergence of androgen-independent prostate cancer. Here we report the identification, isolation, and characterization of distinct classes of AR variants from spontaneous prostate tumors in the TRAMP model. Using cDNA cloning, single stranded conformation polymorphism and sequencing strategies, 15 unique somatic mutations in the AR were identified in prostate tumors obtained from eight TRAMP mice between 24 and 29 weeks of age. At least one mutation was isolated from each mouse. All mutations were single base substitutions, 10 were missense and 5 were silent. Nine mutations in the AR were identified in tumors of four mice that were castrated at 12 weeks of age. Interestingly, the majority of mutations (seven out of nine, 78%) identified in the androgen-independent tumors colocalized in the AR transactivation domain. The remaining mutations colocalized in the AR ligand binding domain. In general, the AR variants demonstrated promoter-, cell-, and cofactor-specific activities in response to various hormones. All AR variants isolated in this study maintained strong sensitivity for androgens, and four AR variants isolated from castrated mice demonstrated increased activities in the absence of ligand. The K638M and F677S variants demonstrated increased activities in response to androgen, and K638M also demonstrated increased response to estradiol. In the presence of AR coactivator ARA70 the E231G variant demonstrated increased activity in response to both androgen and estradiol. However, in the presence of AR coactivator ARA160 the E231G variant was selectively responsive to androgen. Collectively these analyses not only indicate that somatic mutations in the AR gene occur spontaneously in TRAMP tumors but also how changes in the hormonal environment may drive the selection of spontaneous somatic mutations that provide a growth advantage.

Centrosome-centriole abnormalities are markers for abnormal cell divisions and cancer in the transgenic adenocarcinoma mouse prostate (TRAMP) model.

We utilized the transgenic adenocarcinoma mouse prostate (TRAMP) model to study the formation of abnormal mitosis in malignant tumors of the prostate. The results presented here are focused on centrosome and centriole abnormalities and the implications for abnormal cell divisions, genomic instability, and apoptosis. Centrosomes are microtubule organizing organelles which assemble bipolar spindles in normal cells but can organize mono-, tri-, and multipolar mitoses in tumor cells, as shown here with histology and electron microscopy in TRAMP neoplastic tissue. These abnormalities will cause unequal distribution of chromosomes and can initiate imbalanced cell cycles in which checkpoints for cell cycle control are lost. Neoplastic tissue of the TRAMP model is also characterized by numerous apoptotic cells. This may be the result of multipolar mitoses related to aberrant centrosome formations. Our results also reveal that centrosomes at the poles in mitotic cancer cells contain more than the regular perpendicular pair of centrioles which indicates abnormal distribution of centrioles during separation to the mitotic poles. Abnormalities in the centriole-centrosome complex are also seen during interphase where the complex is either closely associated with the nucleus or loosely dispersed in the cytoplasm. An increase in centriole numbers is observed during interphase, which may be the result of increased centriole duplication. Alternatively, these centrioles may be derived from basal bodies that have accumulated in the cell's cytoplasm, after the loss of cell borders. The supernumerary centrioles may participate in the formation of abnormal mitoses during cell division. These results demonstrate multiple abnormalities in the centrosome-centriole complex during prostate cancer that result in abnormal mitoses and may lead to increases in genomic instability and/or apoptosis.

Chemoprevention of prostate carcinogenesis by alpha-difluoromethylornithine in TRAMP mice.

Development of effective chemopreventive agents for human consumption requires conclusive evidence of their efficacy in animal models that have relevance to human diseases. Transgenic adenocarcinoma mouse prostate (TRAMP) is an excellent model of prostate cancer that mimics progressive forms of human disease inasmuch as 100% of males develop histological PIN by 8-12 weeks of age that progress to adenocarcinoma with distant site metastases by 24-28 weeks of age. In these animals, ornithine decarboxylase (ODC) activity (>3-fold) as well as protein expression (>4-fold) was found to be markedly higher in the dorsolateral prostate as compared with the nontransgenic littermates, suggesting their suitability to determine the chemopreventive effect of alpha-difluoromethylornithine (DFMO), an enzyme-activated irreversible inhibitor of ODC, against prostate cancer. Using male TRAMP mice, we studied the effect of oral consumption of DFMO on development of prostate carcinogenesis and surrogate end point biomarkers related to prostate cancer progression. In two independent experiments, each consisting of 8 animals on test, the cumulative incidence of prostatic cancer development at 28 weeks of age in 16 untreated TRAMP mice was 100% (16 of 16), whereas 94% (15 of 16) and 69% (11 of 16) of the animals exhibited distant site metastases to lymph nodes and lungs, respectively. Oral consumption of 1% DFMO (w/v) in the drinking water to TRAMP mice from 8 to 28 weeks of age resulted in a significant decrease in (a) weight (59%) and volume (66%) of prostate, (b) genitourinary weight (63%), and (c) ODC enzyme activity (52%) in the dorsolateral prostate. Importantly, in none of the DFMO-fed TRAMP mice were any distant metastases to lymph node and lungs observed. Furthermore, DFMO treatment resulted in the marked reduction in the protein expression of proliferation cell nuclear antigen, ODC, and probasin in the dorsolateral prostate. The protein expression of antimetastases markers, i.e., E-cadherin and alpha- and beta-catenin, was found to be restored in DFMO-fed animals as compared with the non-DFMO-fed mice. These chemopreventive effects of DFMO were further confirmed by immunohistochemical analysis of the dorsolateral prostate. Histological analysis of the dorsolateral prostate of DFMO-fed animals displayed marginal epithelial stratification, a small number of cribriform structures, elongated hyperchromatic epithelial nuclei, and a significant increase in apoptotic index. Non-DFMO-fed animals, on the other hand, displayed extensive epithelial stratification with profound cribriform structures accompanied with marked thickening, remodeling, and hypercellularity of the fibromuscular stroma. In nontransgenic littermates fed with DFMO, no significant alterations in the above parameters were evident. These data demonstrate that ODC represents a promising and rational target for chemoprevention of human prostate cancer and that TRAMP mice are excellent models for screening of novel drugs and chemopreventive regimens for potential human use.

Genomic instability-based transgenic models of prostate cancer.

To develop animal models that represent the broad spectrum of human prostate cancer, we created transgenic mice with targeted prostate-specific expression of two genes (ECO:RI and c-fos) implicated in the induction of genomic instability. Expression of the transgenes was restricted to prostate epithelial cells by coupling them to the tissue-specific, hormonally regulated probasin promoter (PB). The effects of transgene expression were examined histologically in prostate sections at time points taken from 4 to 24 months of age. The progressive presence of regions of mild-to-severe hyperplasia, low- and high-grade prostatic intra-epithelial neoplasia, and well-differentiated adenocarcinoma was observed in both PBECO:RI lines but no significant pathology was detected in the PBfos line. Prostate tissue of PBECO:RI mice was examined for expression of p53, proliferating cell nuclear antigen (PCNA) and Ki67 at multiple time points. Although p53 does not appear to be mutated, levels of PCNA and Ki67 are elevated and correlate with the severity of the prostatic lesions. Overall, pre-neoplastic and neoplastic stages represented in the PBECO:RI model showed similarity to corresponding early stages of the human disease. This genomic instability-based model will be used to study the mechanisms involved in the early stages of prostate carcinogenesis and to investigate the nature of subsequent events necessary for the progression to advanced disease.

Combination immunotherapy of primary prostate cancer in a transgenic mouse model using CTLA-4 blockade.

We have previously shown that antibodies to CTLA-4, an inhibitory receptor on T cells, can be effective at inducing regression of transplantable murine tumors. In this study, we demonstrate that an effective immune response against primary prostate tumors in transgenic (TRAMP) mice can be elicited using a strategy that combines CTLA-4 blockade and an irradiated tumor cell vaccine. Treatment of TRAMP mice at 14 weeks of age resulted in a significant reduction in tumor incidence (15% versus control, 75%), as assessed 2 months after treatment. Histopathological analysis revealed that treated mice had a lower tumor grade with significant accumulation of inflammatory cells in interductal spaces when treated with anti-CTLA-4 and a granulocyte-macrophage colony-stimulating factor-expressing vaccine. Vaccination of nontransgenic mice with this regimen resulted in marked prostatitis accompanied by destruction of epithelium, indicating that the immune response was, at least in part, directed against normal prostate antigens. These findings demonstrate that this combinatorial treatment can elicit a potent antiprostate response and suggest potential of this approach for treatment of prostate cancer.

E-7869 (R-flurbiprofen) inhibits progression of prostate cancer in the TRAMP mouse.

E-7869 (R-flurbiprofen) is a single enantiomer of a racemic nonsteroidal anti-inflammatory drug. E-7869 does not inhibit either cyclooxygenase-1 or cyclooxygenase-2. We used the transgenic adenocarcinoma mouse prostate (TRAMP) mouse, a prostate cancer model, to evaluate the effect of this drug on prostate cancer progression. Sixty 12-week-old male TRAMP mice were placed randomly into five groups. The animals were treated by daily oral gavage with vehicle (1% carboxymethylcellulose) or E-7869 for 18-weeks. During the course of the study, two diets were used. Three groups (vehicle, 15-mg/kg, and 20-mg/kg drug treatments) received a Teklad diet containing 2.4% saturated fat [a high saturated fat (HSF) diet], and two groups (vehicle and 20 mg/kg drug treatment) received an AIN-93G diet containing 1.05% saturated fat [a low saturated fat (LSF) diet]. At necropsy, the urogenital system and periaortic lymph nodes were removed and weighed. The prostate lobes, seminal vesicles, lungs, and periaortic lymph nodes were preserved and sectioned for histological evaluation. The lung and periaortic lymph nodes were graded as to the presence (+) or absence (-) of metastasis; the urogenital tissues were graded on a 1-6 scale for degree of neoplasia/carcinoma. For both diets, the urogenital wet weights and lymph node wet weights in the 20-mg/kg treatment groups were significantly lower as compared to vehicle control groups. In addition, treatment with 20 mg/kg E-7869 in the LSF diet group resulted in a significantly lower primary tumor incidence (P < 0.05) and reduced incidence of metastasis. In this treatment group, the reduced incidence of metastasis was not statistically significant because the LSF diet itself resulted in a remarkably lower incidence of metastasis in the vehicle control group (10% LSF versus 40% HSF). Treatment with 20 mg/kg E-7869 on the HSF diet resulted in a significantly lower incidence of metastasis (P < 0.05) and a reduction in the primary tumor incidence. These results suggest that E-7869 is a promising chemopreventive and treatment for human prostate cancer.

Androgens and growth factors in prostate cancer: a transgenic perspective.

With the advent of transgenic technology, novel animal models are being developed to facilitate prostate cancer research. The ability to perform genetic perturbation studies in a temporally and spatially restricted manner has established a new paradigm for investigations of the molecular mechanisms involved in the initiation, progression and metastogenesis of prostate cancer. Using the transgenic adenocarcinoma of the mouse prostate (TRAMP) model we have begun to identify specific molecular events related to the emergence of the androgen-insensitive phenotype. In addition, a number of new models have recently been generated to characterize how the deregulation of stromal to epithelial interactions mediated by polypeptide growth factor signaling could facilitate transformation of the prostate. It is anticipated that these models and their successors will ultimately provide new molecular pathway-based strategies for the prevention, detection and treatment of prostate cancer. Prostate Cancer and Prostatic Diseases (2000) 3, 224-228

Early castration reduces prostatic carcinogenesis in transgenic mice.

OBJECTIVES: To test the hypothesis that transgenic mouse models of prostate cancer could be useful for testing chemoprevention strategies by evaluating the effects of early castration on prostate carcinogenesis in TRAMP mice. Human prostate cancer, unlike other cancers, requires androgens for oncogenesis yet acquires partial androgen independence in the castrated milieu. This paradigm is the basis for an ongoing clinical trial using selective androgen deprivation for prostate cancer chemoprevention. However, preclinical correlates for hormonal prevention or other chemoprevention strategies of prostate cancer have not previously been demonstrated in autochthonous models of prostate carcinogenesis. METHODS: Magnetic resonance imaging was used to longitudinally measure prostate growth in castrated and noncastrated TRAMP mice, and mice were prospectively examined for the onset of advanced, palpable prostate cancer. Modulation of androgen-responsive oncogene expression, as well as oncogene expression in refractory cancers, was evaluated by Western blot. RESULTS: Early castration significantly reduced prostate tumor growth as measured by magnetic resonance imaging and improved cancer-free survival. Prevention of prostate cancer development in these mice was associated with durable suppression of androgen-responsive oncogene expression (T-antigen expression not detectable by Western blot); prostate cancers refractory to the hormonal prevention strategy demonstrated androgen-independent oncogene expression. CONCLUSIONS: These findings suggest that carcinogenesis related to androgen-responsive oncogene expression can be prevented in some cases by hormonal manipulation and that transgenic TRAMP mice are useful for the preclinical evaluation of hormonal and possibly other strategies of prostate cancer chemoprevention.

Elimination of residual metastatic prostate cancer after surgery and adjunctive cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade immunotherapy.

Cancer relapse after surgery is a common occurrence, most frequently resulting from the outgrowth of minimal residual disease in the form of metastases. We examined the effectiveness of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade as an adjunctive immunotherapy to reduce metastatic relapse after primary prostate tumor resection. For these studies, we developed a murine model in which overt metastatic outgrowth of TRAMP-C2 (C2) prostate cancer ensues after complete primary tumor resection. Metastatic relapse in this model occurs reliably and principally within the draining lymph nodes in close proximity to the primary tumor, arising from established metastases present at the time of surgery. Using this model, we demonstrate that adjunctive CTLA-4 blockade administered immediately after primary tumor resection reduces metastatic relapse from 97.4 to 44%. Consistent with this, lymph nodes obtained 2 weeks after treatment reveal marked destruction or complete elimination of C2 metastases in 60% of mice receiving adjunctive anti-CTLA-4 whereas 100% of control antibody-treated mice demonstrate progressive C2 lymph node replacement. Our study demonstrates the potential of adjunctive CTLA-4 blockade immunotherapy to reduce cancer relapse emanating from minimal residual metastatic disease and may have broader implications for improving the capability of immunotherapy by combining such forms of therapy with other cytoreductive measures including surgery.