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We evaluated the chemopreventive efficacy of the organoselenium compound 1,4-phenylenebis(methylene)selenocyanate (p-XSC) against the development of tumors of the lung and forestomach induced by a mixture of benzo(a)pyrene (B(a)P) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), two of the major lung carcinogens present in tobacco smoke. A/J mice (20 mice/group) were given intragastric doses of a mixture of B(a)P (3 micromol/mouse) and NNK (3 micromol/mouse) in cottonseed oil (0.1 ml) once a week for eight consecutive weeks. Mice were fed either AIN-76A control diet or control diet containing p-XSC (10 ppm selenium), either during or after carcinogen administration. Dietary p-XSC significantly reduced lung tumor multiplicity, regardless of whether it was given during or after carcinogen administration. p-XSC was also an effective inhibitor of tumor development in the forestomach. To provide some biochemical insights into the protective role of p-XSC, its effect on selected phase I and II enzyme activities involved in the metabolism of NNK and B(a)P was also examined in vivo in this animal model. Dietary p-XSC significantly inhibited the activities of the phase I enzymes, methoxyresorufin O-dealkylase (MROD) and N-nitrosodimethylamine N-demethylase (NDMAD), in mouse liver, but it had no effect on ethoxyresorufin O-dealkylase (EROD), pentoxyresorufin O-dealkylase (PROD), and erythromycin N-demethylase (ERYTD). Total glutathione S-transferase (GST) enzyme activity, as well as GST-pi and GST-mu enzyme activities, were significantly induced by dietary p-XSC in both the lung and liver. Glutathione peroxidase (GPX) activity was also induced by p-XSC in mouse lung, but not in the liver. Dietary p-XSC had no effect on selenium-dependent glutathione peroxidase (GPX(Se)), GST-alpha, and UDP-glucuronosyl transferase (UDPGT) enzyme activities in either the lung or the liver. These studies suggest that the chemopreventive efficacy of p-XSC, when fed during carcinogen administration, may be, in part, due to the inhibition of certain phase I enzymes involved in the metabolic activation of these carcinogens, and the induction of specific phase II enzymes involved in their detoxification. The mechanisms that account for the effect of p-XSC when fed after carcinogen administration remain to be determined.

Humans are exposed to 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 1-nitropyrene (1-NP) via several environmental sources and both are known mammary carcinogens in rodents, with the former being more potent (K. El-Bayoumy, Y.-H. Chae, P. Upadhyaya, A. Rivenson, K. Kurtzke, B. Reddy, S.S. Hecht, Comparative tumorigenicity of benzo[a]pyrene, 1-nitropyrene, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine administered by gavage to female CD rats, Carcinogenesis 16 (1995) 431-434). Following their metabolic activation, both carcinogens are known to bind covalently to DNA. However, it remains to be determined whether these carcinogens can also induce DNA-base oxidation. Our goal was to determine the effects of PhIP and 1-NP on the levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG; a marker of oxidative DNA damage) in rat mammary glands and to evaluate the effect of the chemopreventive agent 1,4-phenylenebis(-methylene)selenocyanate (p-XSC) as an inhibitor of such damage. As an established potent mammary carcinogen, the synthetic 7,12-dimethylbenz[a]anthracene (DMBA) was included in this study. Female CD rats were fed a high-fat AIN-76A diet (23.5% corn oil) supplemented with p-XSC (10 ppm as selenium) or unsupplemented control diet for 1 week. At 50 days of age, each rat (12 rats/group) was gavaged with either PhIP (22 mg (100 micromol) per rat) or I-NP (20 mg (80 micromol) per rat) in trioctanoin (0.5 ml), DMBA (5 mg (20 micromol) per rat] in olive oil (0.2 ml), or the corresponding vehicle. Rats were sacrificed 6 and 24 h after carcinogen treatment (six rats per time point). Mammary fat pads were excised and DNA was isolated and enzymatically hydrolyzed. The hydrolysates were analyzed for 8-OHdG using HPLC with EC detection. PhIP significantly increased the levels of 8-OHdG by 83% after 6 h (P < 0.05), but the increase (47%) at the 24 h point was not significant. p-XSC alone had no effect on the levels of 8-OHdG. However, the elevation of 8-OHdG caused by PhIP at 6 h was significantly inhibited by p-XSC to levels similar to those measured in rats treated with the vehicle only (P < 0.05). p-XSC had no effect on PhIP-induced 8-OHdG at 24 h. I -NP had no effect on the levels of 8-OHdG at either time point. Levels of 8-OHdG were increased by 22% 6 h after DMBA administration and, significantly, rose to 84% at 24 h (P < 0.01); at either time point, this elevation was not inhibited by p-XSC. Although the mechanisms remain to be determined, to our knowledge, this is the first report demonstrating that PhIP and DMBA are capable of enhancing 8-OHdG levels in the rat mammary tissue in vivo.

1,4-Phenylenebis(methylene)selenocyanate (p-XSC) is an effective chemopreventive agent against 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung adenoma in female A/J mice. While p-XSC can effectively inhibit NNK-induced DNA methylation in female A/J mice and in male F344 rats, its effect on NNK-induced oxidative DNA damage had not been determined. Thus, the effect of p-XSC on the levels of 8-hydroxy-2'-deoxyguanosine (8-OH-dG) in lung DNA from A/J mice and F344 rats treated with NNK was examined. Mice were given NNK by gavage (0.5 mg/mouse in 0.2 ml corn oil, three times per week for 3 weeks) or by a single i.p. injection (2 mg/mouse in 0.1 ml saline) while maintained on a control diet (AIN-76A) or control diet containing p-XSC at 10 or 15 p.p.m. (as Se) starting 1 week before NNK administration and continuing until termination. Mice were killed 2 h after the last NNK gavage in the multiple administration protocol or 2 h after the single i.p. injection. Treatment with NNK by gavage significantly elevated the levels of 8-OH-dG in lung DNA of A/J mice from 0.7 +/- 0.1 to 1.6 +/- 0.2 adducts/10(5) 2'-deoxyguanosine (dG) (P < 0.001), while dietary p-XSC (at 10 p.p.m. Se) prevented significant elevation of the levels of this lesion caused by NNK, keeping them at 0.9 +/- 0.1 adducts/10(5) dG (P < 0.003). Injection of NNK in saline also significantly increased the levels of 8-OH-dG in lung DNA of A/J mice from 1.2 +/- 0.6 to 3.6 +/- 0.8/10(5) dG adducts (P < 0.01), while dietary p-XSC (at 15 p.p.m. Se) kept these levels at 1.9 +/- 0.5 adducts/10(5) dG (P < 0.03). Rats were given a single i.p. injection of NNK (100 mg/kg body wt) in saline while being maintained on control diet (AIN-76A) or control diet containing p-XSC (15 p.p.m. as Se) starting 1 week before NNK administration and continuing until termination. The rats were killed 2 h after injection. Treatment with NNK using this protocol significantly elevated the levels of 8-OH-dG in lung DNA of F344 rats from 2.6 +/- 0.5 to 3.5 +/- 0.5 adducts/10(5) dG (P < 0.03), while dietary p-XSC (at 15 p.p.m. Se) kept the levels of this lesion at 2.2 +/- 0.6 adducts/10(5) dG (P < 0.01). Our findings suggest that the chemopreventive efficacy of p-XSC against NNK-induced lung tumorigenesis in A/J mice and F344 rats may be due in part to inhibition of oxidative DNA damage.

1,4-Phenylenebis(methylene)selenocyanate (p-XSC) inhibits chemically induced tumors in several laboratory animal models. To understand its mode of action, we synthesized p-[14C]XSC, examined its excretion pattern in female CD rats and also the nature of its metabolites. p-[14C]XSC was synthesized from alpha,alpha-dibromo-p-[ring-14C]xylene in 80% yield. The excretion profile of p-[14C]XSC (15.8 mg/kg body wt, 200 microCi/rat, oral administration, in 1 ml corn oil) in vivo was monitored by measuring radioactivity and selenium content. On the basis of radioactivity, approximately 20% of the dose was excreted in the urine and 68% in the feces over 3 days. The cumulative percentages of the dose excreted over 7 days were 24% in urine and 75% in feces, similar to excretion rates of selenium. According to selenium measurement, <1% of the dose was detected in exhaled air; radioactivity was not detected. Only 15% of the dose was extractable from the feces with EtOAc and was identified as tetraselenocyclophane (TSC). Most of the radioactivity remained tightly bound to the feces. Approximately 10% of this bound material converted to TSC on reduction with NaBH4. Organic soluble metabolites in urine did not exceed 2% of the dose; sulfate (9 % of urinary metabolites) and glucuronic acid (19.5% of urinary metabolites) conjugates were observed but their structural identification is still underway. Co-chromatography with a synthetic standard led to the detection of terephthalic acid (1,4-benzenedicarboxylic acid) as a minor metabolite. The major urinary conjugates contained selenium. Despite the low levels of selenium in the exhaled air, the reductive metabolism of p-XSC to H2Se cannot be ruled out. Identification of TSC in vivo indicates that a selenol may be a key intermediate responsible for the chemopreventive action of p-XSC.