Early Prostate Carcinogenesis Research Articles

Progressive Spreading of DNA Methylation in the GSTP1 Promoter CpG Island across Transitions from Precursors to Invasive Prostate Cancer.

DNA hypermethylation at the GSTP1 promoter progressively spreads from being unmethylated in normal prostate to intermediate levels in precursor lesions to extensive methylation in cancer. This molecular progression of GSTP1 promoter methylation patterns in early prostate carcinogenesis could be useful for identification and interception of prostate cancer precursors.

Orchestration of miRNA Patterns by Testosterone and Dietary Tomato Carotenoids during Early Prostate Carcinogenesis in TRAMP Mice

BackgroundThe integrative effects of prostate cancer risk factors, such as diet and endocrine status, on cancer-associated miRNA expression are poorly defined. ObjectivesThis study aimed to define the influence of androgens and diet (tomato and lycopene) on prostatic miRNA expression during early carcinogenesis in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. MethodsWild type (WT) and TRAMP mice were fed control, tomato-containing, or lycopene-containing diets from 4 to 10 weeks of age. Mice underwent either sham (intact) or castration surgery at 8 wk, and half of the castrated mice received testosterone (2.5 mg/kg body weight/d) at 9 wk. Mice were killed at 10 wk, and dorsolateral prostate expression of 602 miRNAs was assessed. ResultsWe detected expression of 88 miRNAs (15% of 602), all of which were present in the TRAMP, in comparison with 49 miRNAs being detectable (8%) in WT. Expression of 61 miRNAs differed by TRAMP genotype, with the majority upregulated in TRAMP. Of the 61 miRNAs, 42 were responsive to androgen status. Diet affected 41% of the miRNAs, which differed by genotype (25/61) and 48% of the androgen-sensitive miRNAs (20/42), indicating overlapping genetic and dietary influences on prostate miRNAs. Tomato and lycopene feeding influenced miRNAs previously associated with the regulation of androgen (miR-145 and let-7), MAPK (miR-106a, 204, 145/143, and 200b/c), and p53 signaling (miR-125 and miR-98) pathways. ConclusionsExpression of miRNAs in early prostate carcinogenesis is sensitive to genetic, endocrine, and diet drivers, suggesting novel mechanisms by which tomato and lycopene feeding modulate early prostate carcinogenesis.

Study on Early Prostate Cancer Antigen (EPCA) and existent risk factors of prostate cancer, Sudan: A case-control study

Background: Early prostate cancer antigen (EPCA), a nuclear matrix protein, has recently been recommended as a hopeful biomarker for early prostate carcinogenesis. Objectives: To evaluate the validity of serum early prostate cancer antigen (EPCA) in the early detection of prostate cancer (PCa) and the characteristics of Sudanese patients diagnosed with prostate cancer. Method: In this study, seventy men were considered as a case subject, who were diagnosed as cancer prostate at Gezira Hospital for Renal Disease and Surgery (GHRDS), Sudan during the period February 2018 to July 2019. Randomly selected sixty patients of BPH patients and forty-five apparently healthy men as controls group. EPCA, and PSA estimations were performed from serum samples using the principle of Enzyme Linked Immunosorbent Assay (ELISA). Results: There was significant association between age, family history, residence, unhealthy habits, education, BMI, occupations and prostate cancer, results also revealed a significant elevation between the means of the serum levels of both early prostate cancer antigen and prostate specific antigen of the patients with prostate cancer when compared with the control groups. EPCA biomarker offered the best performance statically (P= 0.00) and highest specificity (81%) and sensitivity (94%) in prostate cancer detection in Sudanese males over PSA biomarker. Conclusions: Cases of studied prostate cancer linked to many risk factors. Serum levels of early prostate cancer antigen and prostate specific antigen were significantly increased in patients with prostate cancer, however, our data proposed that EPCA has higher power statistical value, so could be utilized as a prostate cancer specific and sensitive biomarker for early detection of prostate cancer.

Abstract 1: Transcriptional profiling of prostatic Skp2 knock-in mouse model and development of the associated prostate organoids for testing Skp2 targeting agents

Abstract S-phase kinase associated protein 2 (Skp2) is a promising drug target as studies have demonstrated that Skp2 is required for spontaneous tumor development that occurs in the retinoblastoma protein (pRb), Pten, or p53 deficient mice. We, therefore, aim to establish CRISPR human Skp2 (hSkp2) knock-in in the prostates of mice to study the molecular profiling features in prostate carcinogenesis. Prostate weights increased in transgenic mice and overexpression of hSkp2 induced prostatic lesions including hyperplasia, mouse prostate intraepithelial neoplasia (mPIN), and carcinoma, which suggested the initial role of hSkp2 in early prostate carcinogenesis. RNAseq results revealed Myl3 and Ctgf as the top down-expressed and up-expressed genes respectively in hSkp2 mice prostates. Gene set enrichment analysis (GSEA) demonstrated the significant upregulations of ribosome and proteasome pathways which had similar alterations in the human prostate cancer dataset with Skp2 overexpression, suggesting the potential role of ribosome biogenesis in prostate tumorigenesis and for drug development. In addition, Skp2 organoids derived from transgenic mice prostates were being developed for convenient and efficient screening of specific Skp2 inhibitors. Flavokawain A (FKA) and Skp2 inhibitor C1 both selectively decreased the viability and altered the morphologies of hSkp2 organoids, meanwhile FKA exhibited less toxicity on wild-type organoids. Citation Format: Liankun Song, Vyvyan Nguyen, Shan Xu, Jana Yamak, Kia Arabzadehkaffash, Ali Fazelpour, Merci Mino, Matthew Tippin, Shuang Meng, Xiaolin Zi. Transcriptional profiling of prostatic Skp2 knock-in mouse model and development of the associated prostate organoids for testing Skp2 targeting agents [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1.

Growth and differentiation factor 15 and NF‐κB expression in benign prostatic biopsies and risk of subsequent prostate cancer detection

ABSTRACTGrowth and differentiation factor 15 (GDF‐15), also known as macrophage inhibitory cytokine 1 (MIC‐1), may act as both a tumor suppressor and promotor and, by regulating NF‐κB and macrophage signaling, promote early prostate carcinogenesis. To determine whether expression of these two inflammation‐related proteins affect prostate cancer susceptibility, dual immunostaining of benign prostate biopsies for GDF‐15 and NF‐κB was done in a study of 503 case‐control pairs matched on date, age, and race, nested within a historical cohort of 10,478 men. GDF‐15 and NF‐κB expression levels were positively correlated (r = 0.39; p < 0.0001), and both were significantly lower in African American (AA) compared with White men. In adjusted models that included both markers, the odds ratio (OR) for NF‐κB expression was statistically significant, OR =0.87; p = 0.03; 95% confidence interval (CI) =0.77–0.99, while GDF‐15 expression was associated with a nominally increased risk, OR =1.06; p = 0.27; 95% CI =0.96–1.17. When modeling expression levels by quartiles, the highest quartile of NF‐κB expression was associated with almost a fifty percent reduction in prostate cancer risk (OR =0.51; p = 0.03; 95% CI =0.29–0.92). In stratified models, NF‐κB had the strongest negative association with prostate cancer in non‐aggressive cases (p = 0.03), older men (p = 0.03), and in case‐control pairs with longer follow‐up (p = 0.02). Risk associated with GDF‐15 expression was best fit using nonlinear regression modeling where both first (p = 0.02) and second (p = 0.03) order GDF‐15 risk terms were associated with significantly increased risk. This modeling approach also revealed significantly increased risk associated with GDF‐15 expression for subsamples defined by AA race, aggressive disease, younger age, and in case‐control pairs with longer follow‐up. Therefore, although positively correlated in benign prostatic biopsies, NF‐κB and GDF‐15 expression appear to exert opposite effects on risk of prostate tumor development.

Abstract 1146: Development of CRISPR human Skp2 knock-in in the prostate of mouse and the associated prostate organoids for testing Skp2 targeting agents

Abstract S-phase kinase associated protein 2 (Skp2) is a promising drug target as studies have demonstrated that Skp2 is required for spontaneous tumor development that occurs in the retinoblastoma protein (pRb), Pten or p53 deficient mice. We therefore have aimed to establish CRISPR human Skp2 (hSkp2) knock-in in the prostate of mouse to examine its role in prostate carcinogenesis. In addition, Skp2 overexpressing organoids are being developed for convenient and efficient screening of specific Skp2 inhibitors. cDNA of hSkp2 and porcine teschovirus-1 (P2A) was introduced into immediate upstream of mouse probasin coding sequencing using CRISPR knock-in targeting approach to establish transgenic mice specifically overexpressing hSkp2 in the prostate glands. Quantitative PCR, western blot and immunohistochemistry (IHC) were used to identify the expression levels of Skp2 and histopathological analyses were performed after H&amp;E staining. Prostate glands were dissected and digested for organoid culture, then morphological features and viability of organoids were evaluated following treatments of potential Skp2 inhibitors. Prostate glands and other tissues from mice of three transgenic lines which were genotyped positive were dissected and hSkp2 was detected in ventral, dorsal and lateral prostate lobes, but not in testis, kidney, spleen, and liver. Line 10092 was found to have the highest expression of hSkp2 among these three lines. Prostate weight slightly increased in transgenic mice compared to that of wild-type control mice. Hyperplasia, prostatic intraepithelial neoplasia (PIN), and carcinoma were noted, which suggested that overexpression of hSkp2 in the prostate of mice promoted proliferation and prostate tumorigenesis. Organoids that were derived from the prostate of hSkp2 transgenic mice recapitulated prostatic phenotypic features and maintained a high level of hSkp2. A novel Flavokawain A derivative was found to selectively alter the morphology of hSkp2 organoids and decrease the viability. Prostatic lesions found in the hSkp2 knock-in mice demonstrated a potential role of hSkp2 in early prostate carcinogenesis and organoids overexpressing hSkp2 have the utility for screening out compounds targeting Skp2 in prostate cancer. Citation Format: Liankun Song, Shan Xu, Kia Arabzadehkaffash, Ali Fazelpour, Dongjun Fu, Matthew Tippin, Xiaolin Zi. Development of CRISPR human Skp2 knock-in in the prostate of mouse and the associated prostate organoids for testing Skp2 targeting agents [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1146.

Dietary Tomato Varieties Similarly Inhibit Prostate Carcinogenesis in the TRAMP Model in Association with Distinct Transcriptomic and Metabolomic Profiles

Abstract Objectives Lycopene intake is associated with reduced risk of prostate cancer, especially lethal disease. Tangerine tomatoes contain highly bioavailable lycopene isomers, compared to red tomatoes, but impact of isomers on cancer preventive bioactivity are unknown. Our goals are: (1) to determine if feeding tomato containing diets with differing lycopene isomers inhibit transgenic adenocarcinoma of the mouse prostate (TRAMP) carcinogenesis, and (2) to determine key early metabolic and transcriptional pathways through which tomatoes act to prevent carcinogenesis. Methods We examine prostate carcinogenesis in the TRAMP model, comparing two tomatoes with distinct lycopene isomer profiles: all-trans-lycopene-rich red tomatoes and cis-lycopene-isomer-rich tangerine tomatoes. Weanling TRAMP and wild type (WT) mice were fed AIN-93 G control, 10% tangerine tomato + AIN-93 G (w/w), or 10% red tomato + AIN-93 G (w/w) diet until 10 or 18 wks. At 10 wks., prostate RNA-seq and plasma metabolomics were performed on TRAMP and WT samples. At 18 wks., plasma carotenoid analysis and histopathology was performed on TRAMP mice. Results At 18 wks., plasma lycopene concentrations were 2.84-fold greater in tangerine tomato-fed mice than red tomato-fed mice (p &amp;lt; 0.0001), yet both red and tangerine tomato diets similarly prevent carcinoma incidence compared to the control diet by 43% and 36% respectively (p &amp;lt; 0.001, both vs. control). Our investigation of early carcinogenesis (10 wks.) shows that both tomato diets similarly impact the plasma metabolome, and we define prostate transcriptomic profiles mediated by TRAMP genotype, such as inflammation- and immune-regulated pathways, that are inhibited by dietary tomato. We identify an inflammatory gene panel (Olr1, Il1a, Cscl9, and Ccl22) which is inhibited by tomato diets. In a human prostate cancer database, higher expression of this gene panel is associated with a more aggressive phenotype. Conclusions Together, our results implicate red and tangerine tomatoes as potentially beneficial for the inhibition of early prostate carcinogenesis through the modulation of transcriptional programs centered on inflammation and the immune response. Our findings support efforts to test novel tomato products for the inhibition of human prostate carcinogenesis. Funding Sources AICR, NIH P30, USDA NNF.

Tomato and Lycopene Feeding Impact Expression of Lipid and Cholesterol Metabolism Genes in Early TRAMP Mouse Model Prostate Carcinogenesis (OR05-05-19)

ObjectivesEpidemiologic evidence suggests consuming tomato and lycopene are associated with a reduced risk of advanced and lethal prostate cancer. Prior studies of prostate cancer prevention in the TRAMP mouse model indicate that lifelong tomato- or lycopene-feeding reduce incidence of early carcinoma by up to 70%, and this effect is dependent on the expression of Bco2 (beta-carotene oxygenase 2, a carotenoid metabolic enzyme). We hypothesize that gene expression patterns will reveal the mechanisms by which these diets act to disrupt early carcinogenesis. MethodsTo define the early transcriptional responses in the dorsolateral mouse prostate, TRAMP and wild-type littermate mice were crossed with Bco2+/+ or Bco2–/– mice (C57/Bl6 background). All 4 crosses were fed either control, lycopene, or 10% tomato powder diets from weaning at 3 weeks until 8 weeks. Expression of 84 genes was measured by a prostate cancer-focused PCR array (n = 5/group). Protein expression was measured by Western blotting (n = 4/group) and serum carotenoids by HPLC (n = 3–4/group). Statistical effects of TRAMP and Bco2 genotypes and diet treatment on final body mass (n = 6/group), serum lycopene, and gene expression were analyzed by ANOVA (alpha = 0.05). ResultsBody masses were not influenced by genotypes or diets. Serum lycopene concentrations were subject to a Bco2 genotype effect (P = 0.004), with greater lycopene being present in Bco2–/– mice than Bco2 +/+mice, but did not differ by TRAMP genotype nor between lycopene vs tomato diets. The TRAMP genotype influenced the expression of 49 genes, diet impacted expression of 11 genes, and Bco2 genotype influenced expression of 2 genes. Expression of 4 genes, Apc, Mto1, Nfkb1, and Rbm39, were subject to a significant Bco2 x diet interaction. Expression of 5 genes related to lipid metabolism, Fasn, Acaca, Srebf1, Hmgcr, and Ptgs1, were impacted by a diet effect. ConclusionsSemi-targeted analyses suggest tomato and lycopene may affect lipid metabolism in early carcinogenesis. Future studies may elaborate upon specific pathways modulated by tomato and lycopene in early prostate carcinogenesis. Funding SourcesNIH/National Cancer Institute, NIH/National Center for Complementary and Integrative Health, USDA/Agricultural Research Service.

Vitamin D Signaling Suppresses Early Prostate Carcinogenesis in TgAPT121 Mice.

We tested whether lifelong modification of vitamin D signaling can alter the progression of early prostate carcinogenesis in studies using mice that develop high-grade prostatic intraepithelial neoplasia that is similar to humans. Two tissue-limited models showed that prostate vitamin D receptor (VDR) loss increased prostate carcinogenesis. In another study, we fed diets with three vitamin D3 levels (inadequate = 25 IU/kg diet, adequate for bone health = 150 IU/kg, or high = 1,000 IU/kg) and two calcium levels (adequate for bone health = 0.5% and high = 1.5%). Dietary vitamin D caused a dose-dependent increase in serum 25-hydroxyvitamin D levels and a reduction in the percentage of mice with adenocarcinoma but did not improve bone mass. In contrast, high calcium suppressed serum 1,25-dihydroxyvitamin D levels and improved bone mass but increased the incidence of adenocarcinoma. Analysis of the VDR cistrome in RWPE1 prostate epithelial cells revealed vitamin D-mediated regulation of multiple cancer-relevant pathways. Our data support the hypothesis that the loss of vitamin D signaling accelerates the early stages of prostate carcinogenesis, and our results suggest that different dietary requirements may be needed to support prostate health or maximize bone mass. SIGNIFICANCE: This work shows that disrupting vitamin D signaling through diet or genetic deletion increases early prostate carcinogenesis through multiple pathways. Higher-diet vitamin D levels are needed for cancer than bone.

Dietary Tomato and Lycopene Impact Androgen Signaling- and Carcinogenesis-Related Gene Expression during Early TRAMP Prostate Carcinogenesis

Consumption of tomato products containing the carotenoid lycopene is associated with a reduced risk of prostate cancer. To identify gene expression patterns associated with early testosterone-driven prostate carcinogenesis, which are impacted by dietary tomato and lycopene, wild-type (WT) and transgenic adenocarcinoma of the mouse prostate (TRAMP) mice were fed control or tomato- or lycopene-containing diets from 4 to 10 weeks of age. Eight-week-old mice underwent sham surgery, castration, or castration followed by testosterone repletion (2.5 mg/kg/d initiated 1 week after castration). Ten-week-old intact TRAMP mice exhibit early multifocal prostatic intraepithelial neoplasia. Of the 200 prostate cancer-related genes measured by quantitative NanoString, 189 are detectable, 164 significantly differ by genotype, 179 by testosterone status, and 30 by diet type (P < 0.05). In TRAMP, expression of Birc5, Mki67, Aurkb, Ccnb2, Foxm1, and Ccne2 is greater compared with WT and is decreased by castration. In parallel, castration reduces Ki67-positive staining (P < 0.0001) compared with intact and testosterone-repleted TRAMP mice. Expression of genes involved in androgen metabolism/signaling pathways is reduced by lycopene feeding (Srd5a1) and by tomato feeding (Srd5a2, Pxn, and Srebf1). In addition, tomato feeding significantly reduced expression of genes associated with stem cell features, Aldh1a and Ly6a, whereas lycopene feeding significantly reduced expression of neuroendocrine differentiation-related genes, Ngfr and Syp. Collectively, these studies demonstrate a profile of testosterone-regulated genes associated with early prostate carcinogenesis that are potential mechanistic targets of dietary tomato components. Future studies on androgen signaling/metabolism, stem cell features, and neuroendocrine differentiation pathways may elucidate the mechanisms by which dietary tomato and lycopene impact prostate cancer risk.

MP52-04 COLOCALIZATION OF ANDROGEN RECEPTOR AND ESTROGEN RECEPTOR ALPHA IN THE STROMAL MICROENVIRONMENT SUPPORTS A ROLE FOR ESTROGENS IN EARLY PROSTATE CANCER PROGRESSION

You have accessJournal of UrologyProstate Cancer: Basic Research V1 Apr 2014MP52-04 COLOCALIZATION OF ANDROGEN RECEPTOR AND ESTROGEN RECEPTOR ALPHA IN THE STROMAL MICROENVIRONMENT SUPPORTS A ROLE FOR ESTROGENS IN EARLY PROSTATE CANCER PROGRESSION Tristan Nicholson, Priyanka Sehgal, Sally Drew, Wei Huang, and William Ricke Tristan NicholsonTristan Nicholson More articles by this author , Priyanka SehgalPriyanka Sehgal More articles by this author , Sally DrewSally Drew More articles by this author , Wei HuangWei Huang More articles by this author , and William RickeWilliam Ricke More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.1614AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail Introduction and Objectives Androgens, acting via androgen receptor (AR), are known to be important in prostate cancer (PRCA) progression; there is also an emerging role for estrogens, acting via estrogen receptors (ERs), in PRCA. We evaluated expression and localization of AR and ERα (considered the key estrogenic mediator of prostate carcinogenesis) to better understand the role of these sex steroid receptors in PRCA progression. We asked two questions: (1) what is the prevalence of cells expressing AR, ERα and both receptors during different stages of PRCA progression, and (2) what does the relative prevalence of double positive cells imply about the underlying biology of the stromal microenvironment during PRCA progression? Methods Multiplexed immunohistochemistry was performed to detect AR, ERα and smooth muscle alpha-actin (ACTA2) utilizing a tissue microarray consisting of benign prostate tissue (BPT), high-grade prostatic intraepithelial neoplasia (HGPIN), primary tumor from PRCA, and metastasis (MET). To assess the number of sex steroid receptor positive cells and marker staining intensity, we used an automated pathology platform. Results Expression and staining intensity of AR increased with PRCA progression in prostate epithelium, carcinoma cells and the stromal microenvironment. Epithelial and stromal ERα expression and staining intensity was highest in HGPIN and decreased in PRCA and MET, relative to BPT. However, among stromal cells negative for ACTA2, AR positive cells were less prevalent in PRCA compared to BPT, and there was no significant difference in the prevalence of ERα positive cells. In the epithelium and the stromal microenvironment, double positive (for AR & ERα) cells were most prevalent in HGPIN, relative to BPT. Conclusions Taken together, the present results underscore the importance of androgens and estrogens in the biology of PRCA progression. We found an increased prevalence of cells expressing both AR and ERα in early, but not late stages of PRCA progression, suggesting that while the androgens are important in all stages of progression, the role for estrogens and sex steroid synergy may be most prominent during early prostate carcinogenesis. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e581 Advertisement Copyright & Permissions© 2014MetricsAuthor Information Tristan Nicholson More articles by this author Priyanka Sehgal More articles by this author Sally Drew More articles by this author Wei Huang More articles by this author William Ricke More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Abstract 3701: Tomato carotenoids and testosterone modulate mRNA and miRNA profiles during prostate carcinogenesis.

Abstract There is considerable evidence supporting the hypothesis that tomato carotenoids may contribute to prostate cancer prevention however the mechanisms remain uncertain. We propose that bioactive tomato components, such as lycopene, may impact testosterone signaling to inhibit prostate carcinogenesis. Four week-old C57/BL6 WT and TRAMP male mice were fed either control (AIN93G), 10% tomato powder or 0.25% lycopene beadlets. At eight weeks of age, mice were randomized among treatments: intact, castration, or castration + 2.5mg/kg testosterone repletion and animals sacrificed at 10 weeks of age. Prostate gene expression was quantified by Nanostring nCounter was used to quantify prostate gene expression with a 200 gene murine prostate carcinogenesis custom codeset and miRNA with codeset v1.1. Carotenoid analysis is by HPLC and proliferation (Ki67) by immunohistochemical evaluation. Feeding dietary tomato or lycopene increased serum lycopene, for example, in WT mice concentrations reached 335 +/- 33 nmol/L and 337 +/- 34 nmol/L in tomato and lycopene fed animals, respectively. As expected, the TRAMP genotype and testosterone stimulate proliferation with diet as a modulating factor. In the WT intact prostate, the tomato powder and lycopene diet decreased proliferation (p&amp;lt;0.009). The TRAMP genotype and endocrine status impacted the expression of multiple genes and miRNAs in several distinct patterns. For example, Birc5, Ki67, PCNA, BRCA1, ATM, Aurora kinase A / B, Cdc25c, Cyclin B2, cyclin E2, Fox M1,and miRs 15b, 16, 25, 200a, 200b, and 429 are increased in the TRAMP prostate, only when testosterone is present. Alternatively, TMPRSS2, CD31, CD34, Cox 2, nCad, Cyclin D1, Keratin 5, Shh, and p63, and miRs 145, 205, 328, and 423-5p are decreased in the TRAMP prostate in the presence of testosterone. Nkx3.1, miR 148a, and miR 375 are decreased upon castration whereas beta catenin, FGFR1 and TNF alpha are increased. Tomato and / or Lycopene modestly decreased expression of eCad, HIF 1 alpha, Paxillin, RxR alpha, Sca 1 and Vimentin and increased expression of CD133 and miR 22. These highly controlled in vivo studies document the strong interactions between genetic defects in the prostate, the hormonal environment, and diet in modulating early prostate carcinogenesis. Citation Format: Jennifer M. Thomas-Ahner, Lei Wan, Hsueh-Li Tan, Nancy E. Moran, Amy C. Elsen, Dennis K. Pearl, John W. Erdman, Steven K. Clinton. Tomato carotenoids and testosterone modulate mRNA and miRNA profiles during prostate carcinogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3701. doi:10.1158/1538-7445.AM2013-3701

Bioactive tomato components inhibit cancer promoting activity of testosterone in the mouse prostate epithelium

We hypothesize that tomato phytochemicals, such as lycopene (LYC), may antagonize cancer promoting testosterone activity in the normal prostate. C57/BL6 male mice (4 wks; n=84) consumed a control AIN93G or an identical diet with 10% tomato powder (TP) or 0.04% LYC beadlets. Mice were randomized to intact, castration, or castration + testosterone at 8 wks. Serum LYC analysis was by HPLC and prostate proliferation (Ki67) and apoptosis (ApopTag) were evaluated. Prostate gene expression was by qRT‐PCR. Serum LYC concentrations were 366 nmol/L for TP and 371 nmol/L for LYC and were reduced by castration (−38% in TP, P=0.02; −42% in LYC, P&lt;0.001). Epithelial proliferation in TP and LYC fed intact mice was reduced by 50% (P&lt;0.001) with a 30% inhibition (TP, P=0.02; LYC, P&lt;0.001) observed in the castrate‐testosterone mice. Castration dramatically enhanced apoptosis which was further increased by TP (2.2%, P&lt;0.05). CMO1 expression is present in the liver and not detected in the prostate. The CMO2 gene, present in the liver and potentially involved in eccentric LYC cleavage, was detected in the prostate and reduced by castration (P=0.005) with no effect of diet. Tomato phytochemicals may inhibit early prostate carcinogenesis by impacting androgen signaling pathways that impact sensitivity to early steps in the carcinogenic process. Testosterone may impact CMO2 gene expression suggesting a diet and endocrine interaction.Grant Funding Source: NIH‐NCI/AICR

Implications for chemoprevention of prostate cancer with intake of cruciferous vegetables

Implications for chemoprevention of prostate cancer with intake of cruciferous vegetables

Abstract 2398: Gene expression profiling of TgAPT121 mouse prostate reveals biological processes associated with early prostate carcinogenesis

Abstract The TgAPT121 transgenic mice express a truncated SV40 T antigen (T121) in prostate epithelial cells and have been proposed as a model to study the early stages of prostate cancer (PCa) development. T121 expression inactivates the retinoblastoma family of proteins, causing epithelial hyperplasia, prostatic intraepithelial neoplasia (PIN) by 12-wk of age, and adenocarcinoma with microinvasion by 6-mo of age. Here we examined how the molecular features of the anterior prostate (AP) in this model relate to its histological presentation of PIN. TgAPT121 and non-transgenic control mice were raised to 12 wks, at which time the AP were removed and processed for histology (H&amp;E) and RNA isolation (SV Total RNA Isolation kit; Promega). Transcripts were analyzed using the Affymetrix Mouse Gene 1.0 ST array. Data were preprocessed using Bioconductor packages, differentially expressed genes were identified by Significance Analysis of Microarray, and functional analysis was conducted using the MetaCore analysis suite (GeneGo, Inc.). More than 80% of the prostate epithelium developed PIN lesions and 4310 transcripts were significantly altered (5% FDR, fold-change ≥ 1.5; 3775 unique genes, 2443 up, 1332 down). Consistent with the PIN phenotype, functional analysis showed enrichment of GO terms and pathways related to cell cycle/proliferation, transcriptional activation through E2F1, c-Myc, NF-κB, AP-1, ETS1, and EGR1, DNA damage response, and apoptosis. Signaling through the androgen receptor (AR) and estrogen receptor alpha (ERα) have been shown to contribute to human PCa initiation and progression. Transcripts for components of the AR signaling pathway were increased in TgAPT121 mice, as were the transcriptional targets of AR and ERα. Cell adhesion and gap-junction components were downregulated, suggesting the loss of normal cell attachment and cell-cell communication controlling proliferation and differentiation. We next analyzed a published dataset of human PIN samples collected by laser-capture microdissection (GEO Database, GDS3289). Of the 4247 differentially expressed transcripts (5% FDR; 3868 unique genes, 2016 up, 1852 down), 489 overlapped with the differentially regulated genes from TgAPT121 mice (328 up, 161 down). Functional analysis of this subset of genes showed enrichment of pathways involved in cell cycle progression, apoptosis, DNA damage repair, as well as the disruption of gap junctions. This demonstrates that the early prostatic lesions in TgAPT121 mice share several essential molecular features with human PIN. Future research is needed to determine how these early features contribute to, and are altered along with, the progression of PCa in these mice. Supported by NIH Award CA101113 (JCF), NCI Cancer Prevention Interdisciplinary Education Program (YL), and Purdue Center for Cancer Research (JCF). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2398. doi:10.1158/1538-7445.AM2011-2398

Research Resource: Estrogen-Driven Prolactin-Mediated Gene-Expression Networks in Hormone-Induced Prostatic Intraepithelial Neoplasia

Cotreatment with testosterone (T) and 17β-estradiol (E2) is an established regimen for inducing of prostatic intraepithelial neoplasia (PIN) and prostate cancer in rodent models. We previously used the pure antiestrogen ICI 182,780 (ICI) and bromocriptine, a dopamine receptor agonist, to inhibit PIN induction and systemic hyperprolactinemia in Noble rats and found that the carcinogenic action of T+E2 is mediated directly by the effects of E2 on the prostate and/or indirectly via E2-induced hyperprolactinemia. In this study, we delineate the specific action(s) of E2 and prolactin (PRL) in early prostate carcinogenesis by an integrated approach combining global transcription profiling, gene ontology, and gene-network mapping. We identified 2504 differentially expressed genes in the T+E2-treated lateral prostate. The changes in expression of a subset of 1990 genes (∼80%) were blocked upon cotreatment with ICI and bromocriptine, respectively, whereas those of 262 genes (∼10%) were blocked only by treatment with ICI, suggesting that E2-induced pituitary PRL is the primary mediator of the prostatic transcriptional response to the altered hormone milieu. Bioinformatics analyses identified hormone-responsive gene networks involved in immune responses, stromal tissue remodeling, and the ERK pathway. In particular, our data suggest that IL-1β may mediate, at least in part, hormone-induced changes in gene expression during PIN formation. Together, these data highlight the importance of pituitary PRL in estrogen-induced prostate tumorigenesis. The identification of both E2- and pituitary PRL-responsive genes provides a comprehensive resource for future investigations of the complex mechanisms by which changes in the endocrine milieu contribute to prostate carcinogenesis in vivo.

Androgen-induced programs for prostate epithelial growth and invasion arise in embryogenesis and are reactivated in cancer

Cancer cells differentiate along specific lineages that largely determine their clinical and biologic behavior. Distinct cancer phenotypes from different cells and organs likely result from unique gene expression repertoires established in the embryo and maintained after malignant transformation. We used comprehensive gene expression analysis to examine this concept in the prostate, an organ with a tractable developmental program and a high propensity for cancer. We focused on gene expression in the murine prostate rudiment at three time points during the first 48 h of exposure to androgen, which initiates proliferation and invasion of prostate epithelial buds into surrounding urogenital sinus mesenchyme. Here, we show that androgen exposure regulates genes previously implicated in prostate carcinogenesis comprising pathways for the phosphatase and tensin homolog (PTEN), fibroblast growth factor (FGF)/mitogen-activated protein kinase (MAPK), and Wnt signaling along with cellular programs regulating such 'hallmarks' of cancer as angiogenesis, apoptosis, migration and proliferation. We found statistically significant evidence for novel androgen-induced gene regulation events that establish and/or maintain prostate cell fate. These include modulation of gene expression through microRNAs, expression of specific transcription factors, and regulation of their predicted targets. By querying public gene expression databases from other tissues, we found that rather than generally characterizing androgen exposure or epithelial budding, the early prostate development program more closely resembles the program for human prostate cancer. Most importantly, early androgen-regulated genes and functional themes associated with prostate development were highly enriched in contrasts between increasingly lethal forms of prostate cancer, confirming a 'reactivation' of embryonic pathways for proliferation and invasion in prostate cancer progression. Among the genes with the most significant links to the development and cancer, we highlight coordinate induction of the transcription factor Sox9 and suppression of the proapoptotic phospholipid-binding protein Annexin A1 that link early prostate development to early prostate carcinogenesis. These results credential early prostate development as a reliable and valid model system for the investigation of genes and pathways that drive prostate cancer.

Mitochondrial redox signaling by p66Shc is involved in regulating androgenic growth stimulation of human prostate cancer cells.

p66Shc is shown to negatively regulate the life span in mice through reactive oxygen species (ROS) production. Recent reports, however, revealed that p66Shc protein level is significantly elevated in several human cancer tissues and growth-stimulated carcinoma cells, suggesting a mitogenic and carcinogenic role for p66Shc. In this communication, we demonstrate for the first time that p66Shc mediates androgenic growth signals in androgen-sensitive human prostate cancer cells through mitochondrial ROS production. Growth stimulation of prostate cancer cells with 5alpha-dihydrotestosterone (DHT) is accompanied by increased p66Shc level and ROS production, which is abolished by antioxidant treatments. However, antioxidant treatments do not affect the transcriptional activity of androgen receptor (AR) as observed by its inability to block DHT-induced prostate-specific antigen expression, an AR-dependent correlate of prostate cancer progression. Elevated expression of p66Shc by cDNA transfection increases the basal cell proliferation and, thus, reduces additional DHT-induced cell proliferation. Furthermore, DHT increases the translocation of p66Shc into mitochondria and its interaction with cytochrome c. Conversely, both redox-negative p66Shc mutant (W134F), which is deficient in cytochrome c interaction, and p66Shc small interfering RNA decrease DHT-induced cell proliferation. These results collectively reveal a novel role for p66Shc-ROS pathway in androgen-induced prostate cancer cell proliferation and, thus, may play a role in early prostate carcinogenesis.

ERG upregulation and related ETS transcription factors in prostate cancer

The aim of this study was to identify and validate differentially expressed genes in matched pairs of benign and malignant prostate tissue. Samples included 29 histologically verified primary tumors and 23 benign controls. Microarray analysis was initially performed using a sequence verified set of 40,000 human cDNA clones. Among the genes most consistently and highly upregulated in prostate cancer was the ETS family transcription factor ERG (ETS related gene). This finding was validated in an expanded patient series (37 tumors and 38 benign samples) using DNA oligonucleotide microarray and real-time quantitative PCR assays. ERG was 20- to more than 100-fold overexpressed in prostate cancer compared with benign prostate tissue in more than 50% of patients according to quantitative PCR. Surprisingly, ERG mRNA levels were found to be significantly higher in the endothelial cell line, HUVEC, than in the prostate cell lines PC3, DU145 and LNCaP. In situ hybridization of prostate cancer tissue revealed that ERG was abundantly expressed in both prostate cancer cells and associated endothelial cells. The consistency and magnitude of ERG overexpression in prostate cancer appeared unique, but several related ETS transcription factors were also overexpressed in matched pairs of tumor and benign samples, whereas ETS2 was significantly underexpressed. Our findings support the hypothesis that ERG overexpression and related ETS transcription factors are important for early prostate carcinogenesis.

Protein profiling of microdissected prostate tissue links growth differentiation factor 15 to prostate carcinogenesis.

Identification of proteomic alterations associated with early stages in the development of prostate cancer may facilitate understanding of progression of this highly variable disease. Matched normal, high-grade prostatic intraepithelial neoplasia (hPIN) and prostate cancer cells of predominantly Gleason grade 3 were procured by laser capture microdissection from serial sections obtained from snap-frozen samples dissected from 22 radical prostatectomy specimens. From these cells, protein profiles were generated by surface-enhanced laser desorption/ionization-time of flight mass spectrometry. A 24-kDa peak was observed at low or high intensity in profiles of prostate cancer cells in 19 of 27 lesions and at low intensity in 3 of 8 hPIN lesions but was not detectable in matched normal cells. SDS-PAGE analysis of prostate cancer and matched normal epithelium confirmed expression of a prostate cancer-specific 24-kDa protein. Mass spectrometry and protein data-based analysis identified the protein as the dimeric form of mature growth differentiation factor 15 (GDF15). The increased expression of mature GDF15 protein in prostate cancer cells cannot be explained on the basis of up-regulation of GDF15 mRNA because reverse transcription-PCR analysis showed similar amounts of transcript in normal, hPIN, and prostate cancer cells that were obtained by laser capture microdissection in the same set of serial sections from which the protein profiles were obtained. Our findings suggest that early prostate carcinogenesis is associated with expression of mature GDF15 protein.