Dorsolateral Prostate Research Articles

Alpha-santalol, a derivative of sandalwood oil prevents development of prostate cancer in TRAMP mice

BackgroundAlpha-santalol, a major component of sandalwood oil inhibits growth of cultured prostate cancer cells in vitro by causing apoptosis. However, its in vivo efficacy against prostate cancer development is not established. Hypothesis/purposeProstate cancer is one of the most commonly diagnosed cancers in men. The present study was undertaken to determine the in vivo efficacy of alpha-santalol using TRAMP (Transgenic Adenocarcinoma of Mouse Prostate) mice as a model. MethodsTRAMP mice were used for the in vivo studies. Immunohistochemical analysis was used to assess prostate/tumor sections and Ki-67 staining. Western blot analysis was used to determine expression of various proteins. TUNEL assay was used to detect apoptotic bodies. ResultsAdministration of alpha-santalol (100 mg/kg body weight) decreased the incidence of prostate tumors, average wet weights of urogenital organs and prostate weight compared to control mice. In addition, the area occupied by normal prostate was significantly higher, along with poorly differentiated carcinoma that was significantly lower in the dorsolateral prostate sections of alpha-santalol-treated mice compared with the controls. Furthermore, the dorsolateral sections of prostate from alpha-santalol-treated mice exhibited decreased cell proliferation (Ki-67 staining) in association with induction of apoptosis (TUNEL-positive cells). In agreement with immunohistochemical analysis, Western blotting analysis of prostate/tumor samples from alpha-santalol-treated group revealed a decrease in survivin, XIAP, PCNA, cyclin D and CDC2 levels compared to control samples. ConclusionThe present study shows for the first time, that alpha-santalol administration inhibits the development of prostate cancer in TRAMP mice by decreasing cell proliferation and inducing apoptosis, and warrants future studies for its clinical development.

Differential Effect of Non-Steroidal Anti-Inflammatory Drugs Aspirin and Naproxen against TMPRSS2-ERG (Fusion)-Driven and Non-Fusion-Driven Prostate Cancer.

The consumption of the non-steroidal anti-inflammatory drug (NSAID) aspirin is associated with a significant reduction in the risk of developing TMPRSS2-ERG (fusion)-positive prostate cancer (PCa) compared to fusion-negative PCa in population-based case-control studies; however, no extensive preclinical studies have been conducted to investigate and confirm these protective benefits. Thus, the focus of this study was to determine the potential usefulness of aspirin and another NSAID, naproxen, in PCa prevention, employing preclinical models of both TMPRSS2-ERG (fusion)-driven (with conditional deletion of Pten) and non-TMPRSS2-ERG-driven (Hi-Myc+/- mice) PCa. Male mice (n = 25 mice/group) were fed aspirin- (700 and 1400 ppm) and naproxen- (200 and 400 ppm) supplemented diets from (a) 6 weeks until 32 weeks of Hi-Myc+/- mice age; and (b) 1 week until 20 weeks post-Cre induction in the fusion model. In all NSAID-fed groups, compared to no-drug controls, there was a significant decrease in higher-grade adenocarcinoma incidence in the TMPRSS2-ERG (fusion)-driven PCa model. Notably, there were no moderately differentiated (MD) adenocarcinomas in the dorsolateral prostate of naproxen groups, and its incidence also decreased by ~79-91% in the aspirin cohorts. In contrast, NSAIDs showed little protective effect against prostate tumorigenesis in Hi-Myc+/- mice, suggesting that NSAIDs exert a specific protective effect against TMPRSS2-ERG (fusion)-driven PCa.

Lobe-specific responses of TRAMP mice dorsolateral prostate following celecoxib and nintedanib therapy.

Delayed cancer progression in the ventral prostate of the Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) model has been previously reported upon celecoxib and nintedanib co-administration. Herein, we sought to further investigate the effects of these drugs association in some of their direct molecular targets (COX-2, VEGF and VEGFR-2) and in reactive stroma markers (TGF-β, αSMA, vimentin and pro-collagen 1) in the dorsolateral prostate, looking for lobe-specific responses. Male TRAMP mice were treated with celecoxib (10mg/Kg, i.o.) and/or nintedanib (15mg/Kg, i.o.) for 6weeks and prostate was harvested for morphological and protein expression analyses. Results showed that combined therapy resulted in unique antitumor effects in dorsolateral prostate, especially due to the respective stromal or epithelial antiproliferative actions of these drugs, which altogether led to a complete inversion in high-grade (HGPIN) versus low-grade (LGPIN) premalignant lesion incidences in relation to controls. At the molecular level, this duality in drug action was paralleled by the differential down/upregulation of TGF-β signaling by celecoxib/nintedanib, thus leading to associated changes in stroma composition towards regression or quiescence, respectively. Additionally, combined therapy was able to promote decreased expression of inflammatory (COX-2) and angiogenesis (VEGF/VEGFR-2) mediators. Overall, celecoxib and nintedanib association provided enhanced antitumor effects in TRAMP dorsolateral as compared to former registers in ventral prostate, thus demonstrating lobe-specific responses of this combined chemoprevention approach. Among these responses, we highlight the ability in promoting TGF-β signaling and its associated stromal maturation/stabilization, thus yielding a more quiescent stromal milieu and resulting in greater epithelial proliferation impairment.

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.

Alterations in the balance of sex hormones may affect rat prostatic inflammation and fibrosis, and osteopontin might be involved in this process.

This study aimed to investigate the effects of sex hormone imbalance on rat prostatic inflammation and fibrosis and identify the key molecules involved. Castrated Sprague-Dawley (SD) rats were treated with a constant dose of oestradiol (E2) and different doses of dihydrotestosterone (DHT) to achieve different oestrogen/androgen ratios. After 8weeks, serum E2 and DHT concentrations, relative seminal vesicle weights, histopathological changes and inflammation were measured, collagen fiber content and oestrogen receptor (ER) and androgen receptor (AR) expression were detected, mRNA sequencing and bioinformatics analysis were performed to identify differentially expressed genes (DEGs). The severity of inflammation in the rat dorsolateral prostate (DLP) was higher, collagen fibre content and ER expression in the rat DLP and prostatic urethra were increased and AR expression in the rat DLP was decreased in the 1:1 E2/DHT-treated group than that in the 1:10 E2/DHT-treated group. RNA-seq analysis identified 487 DEGs, and striking increases in the expression of mRNAs encoding collagen, collagen synthesis and degradation enzymes, growth factors and binding proteins, cytokines and chemokines, and cell-surface molecules were confirmed in the 1:1 E2/DHT-treated group compared to the 1:10 E2/DHT-treated group. mRNA expression of secreted phosphoprotein 1 (Spp1) and protein expression of osteopontin (OPN, encoded by Spp1) were increased in the 1:1 E2/DHT-treated group compared to the 1:10 E2/DHT-treated group, and Spp1 expression correlated positively with Mmp7, Cxcl6 and Igfn1 expression. The imbalance in the oestrogen/androgen ratio may affect rat prostatic inflammation and fibrosis, and OPN might be involved in this process.

Effect of Dietary Methylseleninic Acid and Se-Methylselenocysteine on Carcinogen-Induced, Androgen-Promoted Prostate Carcinogenesis in Rats

Selenomethionine (SeMet) did not prevent prostate cancer in the SELECT trial and in two hormone-driven rat models. However, we have shown that daily oral bolus administration of next-generation selenium forms, methylseleninic acid (MSeA) and Se-methylselenocysteine (MSeC) at 3 mg Se/kg body weight, inhibits prostate carcinogenesis in the TRAMP and pten-deficient mouse models and In Vivo growth of human prostate cancer cells. Here, we determined whether these Se forms prevent prostate cancer in a chemically induced-androgen promoted carcinogenesis rat model in which SeMet was not preventive. WU rats were treated with methylnitrosourea, and one week later, slow-release testosterone implants when they were randomized to groups fed AIN-93M diet supplemented with 3 ppm selenium as MSeA or MSeC or control diet. Mean survival, tumor incidence in all accessory sex glands combined (dorsolateral and anterior prostate plus seminal vesicle) and the incidence of tumors confined to dorsolateral and/or anterior prostate were not statistically significantly different among the groups. Thus, MSeA and MSeC feeding was not preventive in this model. The contrast with the inhibitory effects of MSeA and MSeC in mouse models may be due to differences in carcinogenic mechanisms, selenium dosage, delivery mode, and pharmacokinetics or fundamental rat-mouse differences in selenium metabolism.

The MNU Plus Testosterone Rat Model of Prostate Carcinogenesis.

Animal models of prostate cancer are essential to identify chemopreventive treatments against this major male malignancy. The N-methyl-N-nitrosourea (MNU) plus testosterone rat model of prostate carcinogenesis is a reliable animal model that recapitulates human prostate cancer in many respects and has been used extensively in chemoprevention studies with good predictive value for the results of human clinical trials. The objective of this article is to describe the induction protocol of this model, demonstrate its robustness and reproducibility over time and across rat strains, provide diagnostic criteria for the identification of prostate lesions, and present the current tumor induction protocol so that others can use this model in a reliable manner. The majority of accessory sex gland tumors in this model are adenocarcinomas originating in the anterior and dorsolateral prostate that metastasize to lungs and abdominal structures. The rat strain used is of critical importance, with the commercially available Wistar WU and Fischer F344 strains yielding the highest tumor incidences. Low dose, long-term testosterone treatment is essential for a high tumor incidence, but in advanced stage, large adenocarcinomas do not appear to be androgen dependent. This rat model is a robust and reproducible prostate cancer animal model of human prostate cancer.

Characterization of stage-specific tumor progression in TMPRSS2-ERG (fusion)-driven and non-fusion-driven prostate cancer in GEM models.

In the present study, we performed a comparative stage-specific pathological and molecular marker evaluation of TMPRSS2-ERG fusion and PTEN loss-driven (TMPRSS2-ERG. Ptenflox/flox ) versus non-fusion-driven prostate tumorigenesis (Hi-Myc) in mice. Anterior, ventral, and dorsolateral prostates were collected from mice at different ages (or time points post-Cre induction). Results indicated that growth and progression of prostatic intraepithelial lesions to adenocarcinoma stages occurred in both mice models albeit at different rates. In the TMPRSS2-ERG. Ptenflox/flox mice, the initiation of tumorigenesis was slow, but subsequent progression through different stages became increasingly faster. Adenocarcinoma stage was reached early on; however, no high-grade undifferentiated tumors were observed. Conversely, in the Hi-Myc+/- mice, tumorigenesis initiation was rapid; however, progression through different stages was relatively slower and it took a while to reach the more aggressive phenotype stage. Nevertheless, at the advanced stages in the Hi-Myc+/- mice, high-grade undifferentiated tumors were observed compared to the later stage tumors observed in the fusion-driven TMPRSS2-ERG. Ptenflox/flox mice. These results were corroborated by the stage specific-pattern in the molecular expression of proliferation markers (PCNA and c-Myc); androgen receptor (AR); fusion-resultant overexpression of ERG; Prostein (SLC45-A3); and angiogenesis marker (CD-31). Importantly, there was a significant increase in immune cell infiltrations, which increased with the stage of tumorigenesis, in the TMPRSS2-ERG fusion-positive tumors relative to fusion negative tumors. Together, these findings are both novel and highly significant in establishing a working preclinical model for evaluating the efficacy of interventions during different stages of tumorigenesis in TMPRSS2-ERG fusion-driven PCa.

Validation of the Rat Model of Prostate Cancer: Correlating Seminal Vesicle Lesions With Dorsolateral Prostate Lesions.

Lesions in the seminal vesicle are described in the most used protocols for prostate cancer (PCa) induction. This study aimed to characterize the lesions of seminal vesicles associated with a protocol of PCa induction in rats to contribute to better characterization of this model. Forty-five male Wistar Unilever rats were randomly divided into two control groups: CONT1 (n=10) and CONT2 (n=10); and two PCa-induced groups: IND1 (n=10) and IND2 (n=15), sacrificed at 35 and 61 weeks, respectively. Animals from the induced groups were exposed to a multistep protocol for PCa induction. Animals, seminal vesicles and dorsolateral prostate were weighed. Seminal vesicles and dorsolateral prostate were submitted to histopathological and immunohistochemical analysis. Animals in which PCa was induced had a lower mean body weight when compared with the control animals (p<0.05). The relative mean seminal vesicle weight was higher in groups with PCa when compared with control groups (p<0.05). Although the differences were not statistically significant, animals from the IND2 group developed more lesions than animals from the IND1 and CONT2 groups. It is worth noting that the animals from group IND2 developed papillary adenomas and carcinomas in situ, which were not observed in any other group. Similar to observations in seminal vesicles, animals from group IND2 developed more dorsolateral prostate lesions than animals from the IND1 group (p<0.05). We observed that the longer the exposure to testosterone was, the greater was the incidence of preneoplastic and neoplastic lesions in both the seminal vesicle and the prostate, suggesting that testosterone exposure affects the spectrum of developed lesions.

Effects of obesity in rats on prostate histology and expression of leptin receptor, prolactin receptor, IL-6, and NF-κB

Hypercaloric intake can lead to obesity, which is a major risk factor associated with chronic subclinical inflammation and many types of cancer. It can increase the serum levels of leptin, prolactin, nuclear factor kappa B (NF-кB) and interleukin (IL)-6, implicated in cell proliferation, differentiation and survival. To explore the effects of obesity induced by chronic hypercaloric diet in rats on the long-term expression of leptin receptor (OB-R), prolactin receptor, NF-кB, and IL-6, and the changes of histology in rat prostate. From postnatal day 21, experimental males were fed with normal chow or chow plus enriched hypercaloric liquid diet. On the postnatal day 90 (13week old), the animals were euthanized for prostate histology (hematoxylin and eosin staining) and hormone receptors analysis by Western blot. Hypercaloric diet resulted in obesity (32% higher body weight). The prostates of the obese males showed epithelium anisocytosis and compressed interstice. There was also greater volume of lipidic content, anisokaryosis, alterations of the nucleus-cytoplasm ratio, and apparent proplasia. Measures in the ventral prostate (VP) showed that alveoli area increased, but epithelium height and nucleus area were reduced. In the dorsolateral prostate, there was only reduction of nucleus area and presence of mononuclear cells in the lumen. Hypercaloric males also expressed a trend for more OB-R 130kD in the VP, but no changes were observed with regard to prolactin receptor, NF-кB and IL-6. The obesity due to chronic consumption of hypercaloric diet affects both prostatic regions, but VP is possibly more sensitive via OB-R. We suggest that longer periods of obesity are needed to alter other receptors or the molecular markers of inflammation.

Protective effect of the association of curcumin with piperine on prostatic lesions: New perspectives on BPA-induced carcinogenesis

Bisphenol A (BPA) is a chemical agent which can exert detrimental effects on the male reproductive system, especially the prostate gland. In this study we described the efficacy of the dietary agent curcumin, alone or combined with piperine, to suppress the impact of BPA on the prostate. Adult gerbils were divided into nine experimental groups (n = 7 each group), regarding control (water and oil), exposed to BPA (50 μg/kg/day in water) or curcumin (100 mg/kg) and/or piperine (20 mg/kg). To evaluate the effects of the phytotherapic agents, the other groups received oral doses every two days, BPA plus curcumin (BCm), piperine (BP), and curcumin + piperine (BCmP). BPA promoted prostatic inflammation and morphological lesions in ventral and dorsolateral prostate lobes, associated with an increase in androgen receptor-positive cells and nuclear atypia, mainly in the ventral lobe. Curcumin and piperine helped to minimize these effects. BPA plus piperine or curcumin showed a reduction in nuclear atypical phenotype, indicating a beneficial effect of phytochemicals. Thus, these phytochemicals minimize the deleterious action of BPA in prostatic lobes, especially when administered in association. The protective action of curcumin and piperine consumption is associated with weight loss, anti-inflammatory potential, and control of prostate epithelial cell homeostasis.

A 10% Tomato Diet Selectively Reduces Radiation-Induced Damage in TRAMP Mice

BackgroundTomatoes contain carotenoids that have the potential to alter the effects of external beam radiation therapy (EBRT). ObjectivesWe hypothesized that dietary lyophilized tomato paste (TP) would reduce apoptosis within carotenoid-containing nonneoplastic tissues in EBRT-treated TRansgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice. MethodsMale TRAMP mice (n = 73) were provided an AIN-93G diet or a modified AIN-93G diet containing 10% TP (wt:wt) at 4 wk of age. Prostate tumor growth was monitored by ultrasound. The caudal half of the mouse was irradiated with 7.5 Gy (Rad) or 0 Gy (sham) at 24 wk of age or after the tumor volume exceeded 1000 mm3 with a Cobalt-60 source. Mice were euthanized 24 h postradiation. Carotenoids and α-tocopherol were measured by HPLC and compared by a t test. Tissues were assessed for radiation-induced changes (hematoxylin and eosin) and apoptosis [cleaved caspase-3 (CC3)] and compared by Kruskal–Wallis test or Freedman–Lane's permutation test. ResultsSerum concentrations of lycopene (52% lower), phytoene (26% lower), and α-tocopherol (22% lower) were decreased in TP-fed irradiated mice (TP-Rad) compared with TP-fed sham mice (P < 0.05). CC3 scores increased within the prostate tumor with radiation treatments (P < 0.05), but were not affected by tomato consumption. In nonneoplastic tissues, TP-Rad had a lower percentage of CC3-positive cells within the cranial (67% lower) and caudal (75% lower) duodenum than irradiated mice on the control diet (Rad) (P < 0.005). Likewise, CC3 scores within the dorsolateral prostate of TP-Rad trended toward lower scores than for Rad (P = 0.07). ConclusionsTP selectively reduces radiation-induced apoptosis in extratumoral tissues without decreasing radiation-induced apoptosis within the prostate tumor in TRAMP mice. Additional studies are needed to confirm and expand upon these findings.

Abstract 2497: Inositol Polyphosphate 4-Phosphatase Type-II B protects mice from benign prostatic hyperplasia

Abstract BPH is the most common tumor-like condition that afflicts more than 50% of men between the ages of 51-60 and over 90% in men older than 80. While connection between obesity and BPH is well established, the driving signaling pathways are not clear. BPH is a nonmalignant enlargement of prostate epithelium that requires androgen receptor (AR) signaling. INPP4B and EZH2 are highly expressed in prostate epithelium where they regulate AR signaling and play an important role in prostate cancer etiology. EZH2 is an epigenetic silencer that methylates histone H3 on K9 and K27 residues and INPP4B is a dual specificity phosphatase that dephosphorylates PI(3,4)P2 and PI(4,5)P2, membrane lipids that activate Akt and PKC signaling pathways respectively. We discovered that Inpp4b-/- male mice develop BPH at 3 months old of age and subsequently prostate intraepithelial neoplasia (PIN) when fed high fat diet. While expression of AR in the prostates of these mice is similar, AR activity is reduced. The loss of INPP4B caused reduction in EZH2 protein levels in prostate cancer cell lines and in mouse prostate. Inhibition of Akt and PKC signaling reduced EZH2 levels in ventral and dorsolateral prostate lobes of Inpp4b-/- males, suggesting that INPP4B upregulates EZH2 levels by suppressing Akt and PKC signaling pathways. Consistent with decline in EZH2 levels, Inpp4b-/- males have reduced levels of H3K27me3 and H3K9me2 in the ventral prostate, and high fat diet exacerbated this phenotype. Our data suggests that INPP4B modulates epigenetic profile of prostate epithelium by stabilizing EZH2 levels. Both loss of INPP4B and high fat diet treatment lead to decline in EZH2 levels and activity and contribute to development of BPH in mouse model. Citation Format: Yasemin Ceyhan, Manqi Zhang, Irina U. Agoulnik. Inositol Polyphosphate 4-Phosphatase Type-II B protects mice from benign prostatic hyperplasia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2497.

Evaluation of the chemopreventive potential of polyprenols from the needles of Picea abies L. and Pinus sylvestris L. using the model of prostate carcinogenesis in male Wistar rats

For decades, one of the most popular ideas in reducing the burden of cancer worldwide is the idea of cancer chemoprevention with low-toxic agents. There is an evidence that polyprenols (polyisoprenoid alcohols) are perspective agents for cancer chemoprevention. The aim of the study is to investigate anticancer activity of the polyprenol complex (the main polyprenols are prenol-14, prenol-15, prenol-16 and prenol-17) against prostate cancer (PCa) and its precursor (prostatic intraepithelial neoplasia, PIN). Material and methods. For induction of prostate carcinogenesis we used modified protocol of two-stage model. Mature male Wistar rats were administrated with single intravenous injection of N-methyl-N-nitrosourea and chronic intraperitoneal injections of four testosterone esters following surgical castration. Animals with an initial body weight of 328±39 g were randomized into 3 groups: experimental group (n=32) - animals received polyprenols complex 12.5 mg/kg per os diluted in vegetable oil 5 days a week; control group (n=38) - animals received vehicle (vegetable oil) 0.5 ml per os 5 days a week; intact group (n=12) - animals without intervention. Duration of the experiment was 56 weeks. At the end of the study and in euthanized animals in terminal state as well, the prostate and seminal vesicles were processed by histological technique and serial step sections of all lobes were assessed blindly by two independent pathologists. Incidence and multiplicity of precancerous lesion (PIN) and prostate tumors were calculated. Results. Compared with the control group, long-term consumption of polyprenols significantly reduced the overall incidence of PIN from 76.5 to 44.8% (р=0.0183; relative risk, RR=0.59; 95% CI 0.38-0.91), multiplicity of PIN per rat per group by 48.4% (р=0.0081), the incidence of PIN in 2 or 3 lobes of the prostate from 55.9 to 27.6% (р=0.0402), the incidence of PIN in the dorsolateral prostate from 70.6 to 41.4% (р=0.0240), the incidence of PIN in the ventral lobe from 47.1 to 20.7% (р=0.0362). The polyprenol complex exhibited a pronounced anticancer effect against induced prostate tumors. In the group of animals receiving the tested agent, there was a significant decrease in the total incidence of PCa from 64.7 to 34.5% (р=0.0234; RR=0.53; 95% CI 0.30-0.93), as well as the multiplicity of PCa per rat per group and per PIN carrier by 63.4% (р=0.0024) and by 30.6% (р=0.0240) respectively. In the polyprenols group we observed significant decrease in the incidence of prostate cancer in 2 or 3 lobes from 32.4 to 6.9% (р=0.0147) and the incidence of PCa in the dorsolateral prostate from 58.8 to 24.1% (р=0.0101). Oral administration of polyprenols provided a nonsignificant trend towards a decrease in the incidence of metastatic prostate cancer from 32.4 to 20.7% (р=0.3962). Conclusion. Chronic oral consumption of polyprenols from the needles of Picea abies L. and Pinus sylvestris L. (12.5 mg/kg per day) significantly suppresses the development of PIN and PCa induced by MNM and a mixture of testosterone esters in male Wistar rats. Despite the exploratory nature of the study, our results can serve the rationale for further investigation of polyprenols in prostate cancer chemoprevention trials.

Distinct lipid signatures are identified in the plasma of rats with chronic inflammation induced by estradiol benzoate and sex hormones.

Prostatitis is likely to occur in younger or middle-aged men, while prostate cancer is likely to occur in older men. Although amino acids and lipids as biomarkers of prostate cancer have been examined using prostate cancer cell lines/tissues, no previous studies have evaluated amino acids or lipids as potential chronic prostatitis biomarkers. The study's aim was to identify amino acids and lipids that could serve as potential biomarkers of chronic prostatitis. We profiled the amino acids and lipids found in plasma from rats collected in a previous study. In brief, a total of 148 Sprague-Dawley rats (offspring) were dosed with estradiol benzoate (EB) on postnatal days (PNDs) 1, 3 and 5, and subsequently dosed with testosterone (T)/estradiol (E) tubes via subcutaneous implants from PND 90 to 200. Plasma was collected on PNDs 30, 90, 100, 145 and 200. Analysis was conducted with a Xevo TQ-S triple-quadrupole mass spectrometer using a Biocrates AbsoluteIDQ p180 kit. Plasma acylcarnitines [(C2, C16:1, C18, C18:1, C18:1-OH, and C18:2)], glycerophospholipids (lysophosphatidylcholine-acyl, -di-acyl, and -di-acyl acyl-alkyl) and sphingomyelins [SM (OH) C16:1, SM C18:0, SM C18:1, and SM C20:2] significantly increased on PND 145, when chronic inflammation was observed in the dorsolateral prostate of rats dosed with EB, T, and E. No statistical significances of amino acid levels were observed in the EB + T + E group on PND 145. Exposure to EB, T, and E altered lipid levels in rat plasma with chronic prostate inflammation. These findings suggest that the identified lipids may be predictive chronic prostatitis biomarkers. The results require confirmation through additional nonclinical and human studies.

The prostaglandin synthases, COX-2 and L-PGDS, mediate prostate hyperplasia induced by low-dose bisphenol A

This study aimed to identify prostaglandin synthases (PGS) that mediate bisphenol A (BPA)-induced prostatic hyperplasia and explore their underlying mechanisms. In an in vivo study, male adult Sprague–Dawley rats were treated with different concentrations of BPA (10, 30, 90, or 270 μg/kg, i.g., daily), or with vehicle for 4 weeks. Results revealed that low-dose BPA induced prostatic hyperplasia with increased PCNA/TUNEL ratio. It significantly upregulated the expression of cyclooxygenase-2 (COX-2) and NF-κB in the dorsolateral prostate (P < 0.05) and the expression of lipocalin-type prostaglandin D synthase (L-PGDS) in ventral prostate (P < 0.05). The level of estradiol (E2)/testosterone (T) and expression of androgen receptor (AR) and estrogen receptor α (ERα) were also altered. In vitro studies showed that low-dose BPA (0.1–10 nM) promoted the proliferation of human prostate fibroblasts and epithelial cells, and significantly upregulated the expression of COX-2 and L-PGDS in the cells. The two types of cell proliferation induced by BPA were inhibited by COX-2 inhibitor (NS398) and L-PGDS inhibitor (AT56), with increased apoptosis level. These findings suggested that COX-2 and L-PGDS could mediate low-dose BPA-induced prostatic hyperplasia through pathways involved in cell proliferation and apoptosis, which might be related to the functions of ERα and AR. The role of COX-2/NF-κB pathway in dorsolateral prostate requires further research.

Abstract IA12: Role of endocrine disruptors in cancer development

Abstract Multiple lines of evidence show that brief, early-life exposures to bisphenol A (BPA) can increase the risk of prostate cancer (PCa) with aging. Initial work from our laboratory determined that low-dose BPA exposure during rat development reprograms the prostate and enhances incidence and severity of neoplastic lesions (PIN) in response to elevated adult estrogen levels, as occurs in aging men (Prins et al., Repro Tox 2007). Subsequent BPA dose-response studies revealed that elevated carcinogenic risk is found at doses that yield undetectable serum free-BPA with documented PIN progression to PCa (Prins et al., EHP 2017). Epigenetic modifications (altered DNA and histone methylation) provide a mechanistic framework that may connect early-life BPA to later-life predisposition to prostate carcinogenesis. Our studies within the CLARITY-BPA Consortium, a collaboration with the NIEHS, NTP, and FDA, found that continuous low-dose BPA exposure from fetal life to 6 months altered adult dorsal prostate stem cell homeostasis, increased stem cell numbers and progenitor cell proliferation, and shifted lineage commitment to favor basal progenitor cells (Prins et al., EHP 2018). At 1 year, dorsolateral prostate ducts had heightened multiplicity of estrogen-driven adenocarcinoma as a function of chronic BPA exposure (2.5 μg/kg/day). We propose that the BPA-induced stem cell alterations may underpin increased carcinogenic risk with aging. To provide translational evidence for human exposures, two approaches were used. First, we generated a pioneer in vitro model of directed differentiation of hESC into prostatic organoids. Exposure to 1-10 nM BPA resulted in focal clusters of stem cells within the organoids, a phenotype not observed in controls, which suggests aberrant stem-cell self-renewal (Calderon and Prins, PLOS One 2015). Next, we developed in vitro and in vivo models utilizing primary prostate cells from young, disease-free organ donors. BPA exposure to 3-D human prostasphere cultures increased stem-progenitor cell self-renewal and stem-related gene expression through membrane-initiated ER signaling pathways. Tissue-cell recombination of human prostasphere cells and rat UGM with renal grafting into nude mice was used as an in vivo model to assess carcinogenicity (Prins et al., Endocrinol 2014). Developmental BPA exposure combined with adult estradiol led to a heightened PIN/PCa incidence (33-46%) in the human prostate epithelium of the chimeric prostate tissue compared to oil controls given adult estrogen (13%; P&amp;lt;0.01). Together, these findings indicate that stem and progenitor cells may be direct BPA targets and suggest that humans may be susceptible to endocrine-disrupting chemical (EDC)-imprinted prostate disease in a manner similar to the rodent models. Supported by NIH grants ES015584, ES018748, ES020886, ES027792, CA172220. Citation Format: Gail S. Prins. Role of endocrine disruptors in cancer development [abstract]. In: Proceedings of the AACR Special Conference on Environmental Carcinogenesis: Potential Pathway to Cancer Prevention; 2019 Jun 22-24; Charlotte, NC. Philadelphia (PA): AACR; Can Prev Res 2020;13(7 Suppl): Abstract nr IA12.

Brazilian Berry Extract, High‐Fat Diet and Prostate Cancer: In Vitro and In Vivo Evaluation of Inflammation and Extracellular Matrix

The high incidence of prostatic cancer (PCa) has encouraged studies in prostatic cellular biology all over the world. In addition, Brazilian berries, such as Myrciaria jaboticaba (Vell.) Berg, show a high polyphenol concentration in the peel, characterizing antioxidative and anti‐inflammatory properties. The aim herein was to analyze the anti‐inflammatory and remodeling tissue effects of the Jaboticaba peel extract (JPE), a patented product, in early prostatic adenocarcinoma development associated with high‐fat diet (HFD) intake, as well as to investigate the JPE role in human PCa and LNCaP cell lines. TRAMP mice from control groups received standard diet or HFD and the treated groups followed the same diet protocol plus JPE (5.8g/Kg/body weight during 5 days per week for 2 months). The dorsolateral prostate was collected for immunohistochemistry and Western Blotting analyses. PC‐3 and LNCaP cells were treated with different doses of JPE (125, 250, 500 ug/mL) and used for cellular viability assay and Western Blotting analyses. The results showed that the HFD intake led to an MMP‐9 increase and a Laminin protein level decrease, worsening the PCa profile in this transgenic model. Conversely, JPE led to decreased MMP‐9 and also TGF‐beta after HFD intake. Laminin level increased in both groups treated with JPE. Dose and time dependent JPE exposure reduced cellular viability, especially, in androgen‐responsive LNCaP cells. In addition, JPE induced NfkB and iNOS reduction not only in LNCaP but also in PC‐3 cell lines, suggesting that the growth inhibitory effect of JPE in PCa may be associated with its anti‐inflammatory role. To conclude, the tissue damage caused by HFD led to the deregulation of the extracellular matrix elements, which intensified cancer progression, compromising the epithelial‐stromal interaction in the dorsolateral prostate. However, JPE was beneficial for the dynamic interaction between the epithelium and stroma in vivo, and altered cancer cell growth by means of anti‐inflammatory pathway, pointing towards a possible adjuvant therapy for PCa delay, highlighting the natural compounds as an important strategy in prostatic disorders.Support or Funding InformationSao Paulo Research Foundation/FAPESP (2018 04579‐7). And Fund for Support to Teaching, Research and Outreach Activities FAEPEX/UNICAMP (2080/20)

Increased estrogen levels altered microRNA expression in prostate and plasma of rats dosed with sex hormones

Elevated estrogen (E) levels caused by aging or exposure to endocrine disrupting chemicals are related to prostate disease development. Sixty to seventy percent of prostate cancer or benign prostatic hyperplasia patients are over the age of 65, while prostatitis is likely to occur in men under 45years. MicroRNAs currently represent a class of distinctive biological indicators to be used for clinical disease diagnosis and treatment monitoring. This study aims to identify microRNAs that could serve as potential biomarkers for prostate disorders induced by elevated E levels according to their altered expression in prostate or plasma. Groups of Sprague Dawley rats (offspring) were dosed with estradiol benzoate (EB) on postnatal days 1, 3, and 5, and subcutaneously implanted with tubes containing testosterone (T)/E on postnatal day 90. Expression levels of prostate and plasma microRNAs were evaluated using microRNA microarray and validated via qRT-PCR. The expression levels of the potential targeted genes of a set of identified microRNAs were also examined by qRT-PCR. Postnatal administration of EB, T, and E elevated serum E levels with decreased serum T levels in rats. Chronic inflammation was observed in the dorsolateral prostate. Significant changes in expression levels of several microRNAs (rno-miR-146-5p, rno-miR-329-3p, and rno-miR-126a-3p) in the dorsolateral prostate and of a microRNA (rno-miR-329-3p) in the plasma were found in the dosed rats. The target gene expression levels of the altered microRNAs also changed accordingly. Chronic inflammation in the dorsolateral prostate of rats dosed with EB, T, and E resulted in deregulated expression in a set of microRNAs whose target genes were related to tumor growth or abnormal proliferation. Our findings suggest the identified microRNAs and their target genes the potential use as biomarkers to predict prostate cancer development. Validation using human samples is warranted.

Gene expression profiling in dorsolateral prostates of prepubertal and adult Sprague-Dawley rats dosed with estradiol benzoate, estradiol, and testosterone.

The imbalance of testosterone to estradiol ratio has been related to the development of prostate diseases. Although rat models of prostate diseases induced by endocrine-disrupting chemicals (EDCs) and/or hormone exposure are commonly used to analyze gene expression profiles in the prostate, most studies utilize a single endpoint. In this study, microarray analysis was used for gene expression profiling in rat prostate tissue after exposure to EDCs and sex hormones over multiple time points (prepubertal through adulthood). We used dorsolateral prostate tissues from Sprague-Dawley rats (male offspring) and postnatally administered estradiol benzoate (EB) on postnatal days (PNDs) 1, 3, and 5, followed by treatment with additional hormones [estradiol (E) and testosterone (T)] on PNDs 90-200, as described by Ho et al. Microarray analysis was performed for gene expression profiling in the dorsolateral prostate, and the results were validated via qRT-PCR. The genes in cytokine-cytokine receptor interaction, cell adhesion molecules, and chemokines were upregulated in the EB+T+E group on PNDs 145 and 200. Moreover, early-stage downregulation of anti-inflammatory gene: bone morphogenetic protein 7 gene was observed. These findings suggest that exposure to EB, T, and E activates multiple pathways and simultaneously downregulates anti-inflammatory genes. Interestingly, these genes are reportedly expressed in prostate cancer tissues/cell lines. Further studies are required to elucidate the mechanism, including analyses using human prostate tissues.