Interrelationships between dietary restriction, the IGF-I axis, and expression of vascular endothelial growth factor by prostate adenocarcinoma in rats.

Human studies suggest that excessive energy intake and obesity may influence prostate cancer progression. Rodent experiments demonstrate that diet restriction attenuates tumor growth in parallel with reduced vascular density. The present study examines changes in the insulin-like growth factor I (IGF-I) axis caused by dietary restriction and their association with the expression of vascular endothelial growth factor (VEGF) in prostate cancer. Weanling male Copenhagen rats were randomized into control or 40% dietary restricted groups (n = 5). After 8 wk, rats were implanted with rat AT6.3 prostate adenocarcinoma cells. Two weeks later, the animals were sacrificed and serum, normal prostate, liver, and prostate tumor samples were collected for analyses. Dietary restriction reduced serum concentrations of IGF-I by 35% (P < 0.05) and increased IGF-binding protein-3 (IGFBP3) by sevenfold (P < 0.0001). Lower circulating IGF-I concentrations were correlated with reduced IGF-I mRNA expression in the liver, the primary source of circulating IGF-I. Dietary restriction also lowered mRNA expression of IGF-I (45%, P = 0.0242) and its receptor IGFIR (40%, P = 0.0083) in prostate tumors. Similarly, reduced VEGF mRNA (30%, P = 0.0176) and secreted VEGF protein (33%, P = 0.0003) were observed in prostate cancer of restricted rats. An in vitro study employing AT6.3 prostate cancer cells demonstrated dose- and time-dependent stimulation of VEGF expression by IGF-I. These results suggest that dietary restriction reduces endocrine and prostate tumor autocrine/paracrine IGF-I expression, which contributes to reduced VEGF expression and signaling, to inhibit tumor angiogenesis associated with prostate tumorigenesis.

The effects of L-748706, a selective cyclooxygenase-2 inhibitor, on N-nitrosomethylbenzylamine-induced rat esophageal tumorigenesis.

Epidemiological studies suggest that the frequent intake of non-steroidal anti-inflammatory drugs (NSAIDs) is associated with a decreased risk of developing esophageal squamous cell carcinoma (SCC). This decrease is thought to correlate with the inhibition of cyclooxygenase (COX) activity. The production of prostaglandin E2 (PGE2), a major metabolite of COX, is increased in numerous human cancers including esophageal SCC, therefore, inhibition of COX activity and subsequent suppression of the formation of PGE2 may be chemopreventive in the esophagus. The objective of the present study was to determine whether L-748706 (L-706), a novel selective COX-2 inhibitor, would prevent N-nitrosomethylbenzylamine (NMBA)-induced esophageal tumor progression in the Fischer 344 (F344) rat. In rats pretreated with a low-dose of NMBA (0.25 mg/kg body weight), L-706 at 100 p.p.m. in the diet significantly reduced tumor multiplicity but not tumor incidence. At 150 p.p.m. in the diet, L-706 alone and in combination with 200 p.p.m. piroxicam produced significant reductions in both tumor incidence and multiplicity. Inhibition of tumor development in low-dose NMBA-treated rats was associated with reductions in esophageal cell proliferation rates and PGE2 levels in preneoplastic tissues. In contrast, in rats treated with a higher dose of NMBA (0.5 mg/kg body weight), neither L-706 alone nor in combination with piroxicam reduced esophageal tumor incidence or multiplicity in spite of the fact that they reduced esophageal PGE2 levels in preneoplastic tissues and in papillomas. Cell proliferation rates were reduced only in animals treated with L-706 + piroxicam. Our data suggest that the chemopreventive treatments were effective in inhibiting tumor development in NMBA-treated animals only when they reduced PGE2 levels in preneoplastic esophageal tissues approximately to those levels found in normal esophagus.

Perillyl alcohol as a chemopreventive agent in N-nitrosomethylbenzylamine-induced rat esophageal tumorigenesis.

Perillyl alcohol (POH) is a monoterpene found in lavender, spearmint, and cherries. Phase I clinical trials with this agent have shown a favorable toxicity profile and preliminary data indicate some chemotherapeutic efficacy in advanced cancers. Animal studies have demonstrated the ability of POH to inhibit tumorigenesis in the mammary gland, liver, and pancreas. Although the precise mechanism of action is unclear, POH has been shown to inhibit the farnesylation of small G-proteins, including Ras, up-regulate the mannose-6-phosphate receptor, and induce apoptosis. Previous studies in our laboratory using the rat model of squamous cell carcinoma of the esophagus have shown that a specific Ha-ras codon 12 mutation is important for tumor promotion and progression. Given the limited toxicity of POH in humans, its proven efficacy in several animal models and its potential to inhibit Ha-ras farnesylation, we conducted an animal study to evaluate the efficacy of POH as a chemopreventive agent for squamous cell carcinoma of the esophagus. Male Fischer-344 rats were treated s.c. with 0.25 mg/kg b.w. of N-nitrosomethylbenzylamine three times a week for 5 weeks. Three days after the final carcinogen dose, they were started either on control diet or diets containing 0.5 or 1.0% POH. At 25 weeks, the animals were sacrificed, and esophageal tumors were counted. Animals fed either dose of POH showed a significant increase in dysplasia when compared with controls (P < 0.05) and a nonsignificant trend toward increased tumor multiplicity. Additionally, 1.0% POH did not affect Ras membrane localization. These data indicate that POH has a weakly promoting effect early in nitrosamine-induced esophageal tumorigenesis and suggest that POH may not be an effective chemopreventive agent for esophageal cancer in humans.

Piroxicam is an ineffective inhibitor of N-nitrosomethylbenzylamine-induced tumorigenesis in the rat esophagus.

Epidemiological studies indicate an association between the frequent use of nonsteroidal anti-inflammatory drugs and decreased risk for esophageal cancer. These studies suggest that limiting excess prostaglandin production, via inhibition of cyclooxygenase (COX)-mediated arachidonic acid metabolism, may be an important strategy for the prevention of this type of malignancy. N-Nitrosomethylbenzylamine (NMBA)-induced tumorigenesis in the rat esophagus is a model of human esophageal squamous cell carcinoma used for investigations of chemical carcinogenesis and for the evaluation of putative chemopreventive agents. In this study, we characterized COX-mediated arachidonic acid metabolism in NMBA-induced rat esophageal tumorigenesis by measuring COX-1 and COX-2 expression and prostaglandin E(2) production. In addition, we evaluated the ability of piroxicam, a potent COX inhibitor, to prevent postinitiation events of NMBA-induced tumorigenesis in the rat esophagus. After a 2-week acclimatization period, groups of 30 male F344 rats received s.c. injections of NMBA (0.5 mg/kg b.w.) three times/week for 5 weeks. Seventy-two h after the final NMBA treatment and for the remainder of the study, piroxicam was administered in the diet at 200 and 400 ppm. Twenty-five weeks after the initiation of NMBA treatment, we observed an elevation in COX mRNA and protein expression and prostaglandin E(2) production in NMBA-treated esophageal tissues compared with normal epithelium. However, these changes were associated with data indicating that a COX inhibitor is not preventive in NMBA-induced rat esophageal tumorigenesis. Administration of piroxicam in the diet produced no significant reductions in esophageal tumor incidence, multiplicity, or size. The reasons for the lack of effect are largely unknown but may be related to the inability of piroxicam to modulate other biochemical pathways involved in NMBA-induced tumorigenesis.

Functional role of cytochrome p-450 2a3 in N-nitrosomethylbenzylamine metabolism in rat esophagus.

Previous in vitro studies demonstrated that the rat esophageal carcinogen N-nitrosomethylbenzylamine (NMBA) is metabolically activated by cytochrome P-450s (CYP) 2A3 and 2E1. However, the in vivo role of these P-450s in the metabolism of NMBA has not been fully evaluated. In this study, the effects of single and multiple doses of NMBA were investigated on CYP2A3 and CYP2E1 mRNA expression in the rat esophagus and lung. Seven- to 8-wk old male Fischer 344 rats were administered a single subcutaneous dose of NMBA at either 0.5 mg/kg or 2 mg/kg body weight, after which the rats were sacrificed at 1, 3, 6, 12, 24, 48, and 72 h. In the multiple-dose experiment, 2 groups of rats were dosed with 0.5 mg/kg body weight NMBA 3 times per week for 1 wk or 3 wk. The animals were sacrificed 24 h following the last treatment. Semiquantitative reverse-transcription polymerase chain reaction (RT-PCR) analysis demonstrated a reduction of CYP2A3 mRNA expression in lung and esophagus from NMBA-treated animals compared to dimethyl sulfoxide (DMSO)-treated vehicle controls. This reduction in CYP2A3 mRNA was significant at 48 h in the esophagus and at 24 and 48 h in the lung following a single dose of 2 mg/kg body weight NMBA. In contrast, CYP2E1 mRNA expression remained unchanged in rat lung following NMBA treatment and no consistent pattern of expression could be observed in the esophagus. In the multiple-dose study, a 32% and 25% reduction in esophageal CYP2A3 mRNA expression was observed at 1 and 3 wk, respectively. Similar reductions in CYP2A3 mRNA expression were also observed in the lung. Further, esophageal explants derived from animals pretreated with NMBA in vivo demonstrated a reduced ability to metabolize the carcinogen in vitro as compared to explants from vehicle control animals. Taken together, these data provide further support for a potential role of CYP2A3 in NMBA metabolism in the rat esophagus. Data suggest that CYP2A3 levels in the rat esophagus can be a determinant of its ability to metabolize this carcinogen in vivo.