Abstract LB-422: A novel mechanism for the anti-cancer effects of licofelone against testosterone and estradiol-17ß-induced prostate carcinogenesis: Selective modulation of the cross-talk between estrogen receptor and NF-κB

Abstract

Abstract The chemopreventive and therapeutic potential of licofelone, an anti-inflammatory drug, has been reported in preclinical models against the cancers of the colon, breast, pancreas, and bladder, including prostate cancer (PCa). In contrast to nonsteroidal anti-inflammatory drugs (NSAIDs) and selective cyclooxygenase-2 (COX-2) inhibitors, licofelone exerts significantly less adverse effects under chronic use, and is currently undergoing phase III trials for osteoarthritis. However, the anticancer mechanism of licofelone is unclear. To elucidate the molecular mechanisms of licofelone, we used a short-term Noble (NBL) rat PCa model. Male NBL rats at 10 weeks of age were randomly assigned to control and three experimental groups (n=18 per group), and they were fed a semi-purified AIN-93M diet containing 0, 100, 200, and 300ppm of licofelone. After 2 weeks on control and licofelone diets, hormonal implants, the silastic tubing packed with testosterone (T) and estradiol-17 (E2), were placed subcutaneously in rats under anesthesia. Empty silastic implants (n=5 rats) served as vehicle control, and were fed semi-purified AIN-93M diet only. Dietary regimen was resumed immediately after the hormonal implantation, and all rats were sacrificed at the end of 16 weeks. To examine the anti-cancer activities of licofelone, histological, cell proliferation (ki67 staining), and apoptosis (TUNEL assay) analyses were performed using rat dorsolateral prostate (DLP) tissues. Serum T and E2 levels were measured using radioimmunoassay (RIA) kits. The levels of TNF-α, PGE2, and LTB4 in DLP tissues were analyzed using enzyme immunoassay EIA) kits. Real-time PCR, Western blot and immunohistochemical methods were used to determine the molecular targets at the mRNA and protein levels. Dietary administration of licofelone in NBL rats did not induce any toxic effect or weight loss. A 16-week hormonal treatment contributed chronic inflammation, and developed prostate intraepithelial neoplasia (PIN) in 88% of the rats (n=18). Histological analysis revealed that licofelone treatment suppressed PIN lesions significantly, in a dose-dependent manner; suppressed 21% at 100ppm, 47% at 200ppm, and 68% at 300ppm compared to the untreated control (p<0.001). Further, licofelone at 300ppm significantly decreased the rate of tumor cell proliferation by 68%, and enhanced apoptosis by 8-fold in contrast to the untreated controls. T + E2 treatment-induced production of critical inflammatory mediators were significantly decreased by licofelone treatment at 300ppm (p<0.001); prostaglandin E2 (PGE2) was reduced from 674 to 128 pg/mg protein, leukotriene B4 (LTB4) from 169 to 17 pg/mg protein, and tumor necrosis factor alpha (TNF-α) from 540 to 84 pg/mg protein. However, licofelone treatment had a minimal effect on COX-2 expression at the mRNA and protein level, and COX-1 level was not altered. These findings provide evidence that licofelone effectively suppressed the hormone-induced prostate carcinogenesis in NBL rats, independent of COX-2. Since estrogen receptors (ERα and ERß) are the main mediators of estrogen action in the prostate, it is plausible that a positive interaction between ERα and NF-κB (NF-κB) may trigger the transcriptional activation of pro-inflammatory NF-κB signal transduction pathway and may lead to cancer progression. To test this hypothesis, we further examined the expression of ERα, ERß, and NF- Bp65 expression in PIN lesions of the DLP. Our findings revealed that licofelone treatment at 300ppm significantly downregulated ERα and NF-κBp65 expression, whereas ERß expression was upregulated compared to untreated control (p<0.001). In summary, our findings suggest that licofelone modulated several pro-inflammatory targets, and effectively suppressed the hormone-induced PIN development in Noble rats, without significantly affecting COX-2. Most interestingly, our findings are the first to report evidence that licofelone may selectively bind to ERα, and inhibit ERα-mediated activation of NF-κB signaling in prostate carcinogenesis. Further studies are in progress to confirm this unique molecular mechanism of licofelone binding to ER in PCa chemoprevention (Supported by NIH/NCI R03 CA133929 grant to N. Narayanan). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-422.