Control charts 101: a guide to health care applications.

A control chart can be an effective way to display process data over time. It can differentiate common cause variation from special cause variation through the application of several probability-based interpretation rules. In addition, a control chart can be helpful in evaluating the effectiveness of a change. There are numerous types of control charts, and this tutorial was developed to guide the user through the selection process.

Elevated 8-hydroxy-2'-deoxyguanosine levels in lung DNA of A/J mice and F344 rats treated with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and inhibition by dietary 1,4-phenylenebis(methylene)selenocyanate.

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.

Inhibition of lung carcinogenesis by black tea in Fischer rats treated with a tobacco-specific carcinogen: caffeine as an important constituent.

Here, we examined the effect of black tea and caffeine on lung tumorigenesis in F344 rats induced by the nicotine-derived carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in a 2-year bioassay. NNK was administered s.c. at a dose of 1.5 mg/kg body weight three times weekly for 20 weeks. Animals were given either black tea as drinking water at concentrations of 2%, 1%, or 0.5%, or caffeine in drinking water at concentrations identical to those in 2% and 0.5% tea infusions for 22 weeks. The treatment period began 1 week before and ended 1 week after the NNK administration. The animals were sacrificed on week 101 for the examination of tumors in target organs, including lung, liver, nasal cavity, and other major organs. The NNK-treated group, given 2% black tea, showed a significant reduction of the total lung tumor (adenomas, adenocarcinomas, and adenosquamous carcinomas) incidence from 47% to 19%, whereas the group given 1% and 0.5% black tea showed no change. The 2% tea also reduced liver tumor incidence induced by NNK from 34% in the group given only deionized water to 12%. The tumor incidence in the nasal cavity, however, was not affected by either black tea or caffeine at any of the concentrations tested. The most unexpected finding was the remarkable reduction of the lung tumor incidence, from 47% to 10%, in the group treated with 680 ppm caffeine, a concentration equivalent to that found in the 2% tea. This incidence is comparable to background levels seen in the control group. This study demonstrated for the first time in a 2-year lifetime bioassay that black tea protects against lung tumorigenesis in F344 rats, and this effect appears to be attributed, to a significant extent, to caffeine as an active ingredient of tea.

Inhibition of N'-nitrosonornicotine-induced esophageal tumorigenesis by 3-phenylpropyl isothiocyanate.

The ability of dietary isothiocyanates to inhibit the esophageal metabolism of N'-nitrosonornicotine (NNN) was examined in F344 rats. Following feeding of benzyl isothiocyanate (BITC), phenethyl isothiocyanate (PEITC), 3-phenylpropyl isothiocyanate (PPITC), 4-phenylbutyl isothiocyanate (PBITC) or 6-phenylhexyl isothiocyanate for 2 weeks, rats were killed and the esophagi were incubated in vitro with [5-3H]NNN. While dietary BITC, PEITC and PBITC all decreased NNN metabolism, dietary PPITC had the greatest effect, yielding inhibition ranging from 55 to 91% of the control production of various NNN metabolites. To determine the chemopreventive efficacy of PPITC on NNN-induced esophageal tumorigenesis, rats were fed AIN-76A diets containing 0, 1.0 or 2.5 micromol/g PPITC and were given untreated drinking water or drinking water containing 5 p.p.m. NNN. After 87 weeks, the experiment was terminated and the esophageal tumors were counted. Rats that were given untreated drinking water developed no tumors. Rats that were given 5 p.p.m. NNN and unadulterated AIN-76A diet had an esophageal tumor incidence of 71% and a multiplicity of 1.57 tumors/animal. The two dietary concentrations of PPITC reduced the incidence and multiplicity of NNN-induced esophageal tumors by >95%. These results demonstrate the remarkable chemopreventive efficacy of PPITC in the NNN-induced esophageal tumor model.

Wound infection following trauma and burn injuries.

Wounds that result from trauma and burn injuries may lead to the development of infection from the mechanism of injury, bacterial contamination, exogenous and endogenous sources, and impaired host defenses. Knowledge of these factors, combined with thorough wound assessment, differentiation between contaminated and infected wounds, the appropriate use of wound cultures and antibiotics, and the appropriate selection of wound-management techniques, is important in the optimization of wound healing.

Enhancement of esophageal carcinogenesis in male F344 rats by dietary phenylhexyl isothiocyanate.

The purpose of this study was to evaluate the potential effects of dietary 6-phenylhexyl isothiocyanate (PHITC) on N-nitrosomethylbenzylamine (NMBA)-induced esophageal carcinogenesis in rats. Groups of 15 male F344 rats received weekly s.c. injections of NMBA in 20% dimethylsulfoxide or the vehicle alone for 15 consecutive weeks. Two weeks prior to initiation of carcinogen or vehicle injections rats were provided with modified AIN-76A diet or modified AIN-76A diet containing PHITC at levels of 0.4, 1.0 or 2.5 mumol/g diet. Experimental controls consisted of groups that received only the vehicle (vehicle controls), NMBA (carcinogen controls) or PHITC at the high dose level of 2.5 mumol/g diet. No esophageal tumors or preneoplastic lesions were detected in rats that received the vehicle or PHITC alone. In contrast, all rats treated with NMBA alone or PHITC + NMBA exhibited esophageal tumors and preneoplastic esophageal lesions. In groups that received PHITC + NMBA tumor multiplicity was increased by 21-69% when compared with rats treated with NMBA alone, indicating that PHITC enhanced esophageal tumorigenesis in this model system. These results, in conjunction with our previous work, demonstrate that arylalkyl isothiocyanates may inhibit or enhance esophageal tumorigenesis in the NMBA-treated rat. The ability of isothiocyanates to inhibit or enhance experimental tumorigenesis may depend on alkyl chain length of the isothiocyanate, the animal species examined and the specific carcinogen employed.

Structure-activity relationships of isothiocyanates as mechanism-based inhibitors of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced lung tumorigenesis in A/J mice.

A structure-activity relationship study was carried out to identify structural features in arylalkyl and alkyl isothiocyanates that are associated with the inhibitory potency of these compounds against lung tumorigenesis induced in A/J mice by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). These features include the alkyl chain length, phenyl substitution, and secondary isothiocyanates. The naturally occurring allyl isothiocyanate, phenethyl isothiocyanate, and the synthetic analogues such as 6-phenylhexyl isothiocyanate, 8-phenyloctyl isothiocyanate, 10-phenyldecyl isothiocyanate, 1,2-diphenylethyl isothiocyanate, 2,2-diphenylethyl isothiocyanate, and alkyl isothiocyanates (with 1-hexyl, 2-hexyl, and 1-dodecyl as alkyl moieties) were assayed in mice for their tumor inhibitory potential. The isothiocyanates were given in corn oil by gavage at doses of either 0.04, 0.1, and 0.2 mumol or 1 and 5 mumol 2 h prior to a single i.p. injection of 10 mumol NNK. Mice were sacrificed 16 weeks later and lung adenomas were counted. At 0.2 mumol, 8-phenyloctyl isothiocyanate and 10-phenyldecyl isothiocyanate were stronger inhibitors than the previously tested 6-phenylhexyl isothiocyanate, but the difference in potency was not obvious at the lower doses. At both 1 and 5 mumol, allyl isothiocyanate was inactive, while the other five synthetic isothiocyanates were considerably more potent than phenethyl isothiocyanate. In the alkyl isothiocyanate series, 2-hexyl isothiocyanate was more potent than 1-hexyl isothiocyanate, while 1-dodecyl isothiocyanate was the most potent at 1 mumol, reducing tumor multiplicity in the group treated with NNK alone from 11.1 to the background level. Also, 1,2-diphenylethyl isothiocyanate appeared to be a stronger inhibitor than 2,2-diphenylethyl isothiocyanate. In this study we have shown that the phenyl moiety is not essential for the inhibitory activity since alkyl isothiocyanates exhibit strong inhibitory effects against lung tumorigenesis. We have also shown that secondary isothiocyanates possess a higher potency than their structural isomers bearing a primary isothiocyanate. From results of this study and of seven previously studied isothiocyanates, we conclude that the observed inhibitory potency of isothiocyanates in the A/J mouse lung tumor model is correlated with their partition coefficients (log P) and the pseudo first order rate constants for the reaction of isothiocyanates toward glutathione (kobs). These results reveal that both high lipophilicity and low reactivity of isothiocyanates are important for inhibitory activity toward NNK-induced lung tumorigenesis. These observations provide a structural basis for the discovery of more effective chemopreventive agents.

Inhibition of tobacco-specific nitrosamine-induced lung tumorigenesis in A/J mice by green tea and its major polyphenol as antioxidants.

In this study we examined the effects of green tea and its major components, (-)-epigallocatechin gallate (EGCG) and caffeine, on the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung tumorigenesis in A/J mice. We also studied the effects of green tea and EGCG on O6-methylguanine and 8-hydroxydeoxyguanosine (8-OH-dGuo) formation in lung tissues caused by NNK treatment. Mice were given 2% tea, 560 ppm EGCG, or 1120 ppm caffeine in drinking water for 13 weeks. During this time, NNK (11.65 mg/kg body weight) was administered by gavage three times weekly for 10 weeks from weeks 3 to 12. The bioassay was terminated 6 weeks after the last NNK treatment. Mice treated with NNK developed 22.5 lung adenomas per mouse, whereas NNK-treated mice that drank green tea or EGCG as drinking water developed only 12.2 (P less than 0.01) and 16.1 (P less than 0.05) tumors per mouse, respectively. Mice that drank green tea or caffeine solution showed lower body weight gains, although little difference in water and diet consumption was noted in these groups. While green tea and EGCG exerted little effect on the formation of O6-methylguanine, a critical DNA lesion in NNK lung tumorigenesis, both treatments suppressed the increase of 8-OH-dGuo levels in mouse lung DNA. The inhibition of 8-OH-dGuo formation in lung DNA by green tea and EGCG is consistent with their ability to inhibit lung tumorigenesis by NNK. Because 8-OH-dGuo is a DNA lesion caused by oxidative damage, these results suggest that the mechanism of inhibition by green tea and EGCG in NNK-induced lung tumorigenesis is due at least partly to their antioxidant properties.

Effect of frequency of isothiocyanate administration on inhibition of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced pulmonary adenoma formation in A/J mice.

The abilities of phenethyl isothiocyanate (PEITC) and 6-phenylhexyl isothiocyanate (PHITC) to inhibit 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung tumorigenicity, when administered by a standard four-dose protocol or by a single-dose protocol, were determined. Corn oil or isothiocyanates were administered once or for four consecutive days by gavage, with the final (or single) administration of corn oil or inhibitor occurring 2 h prior to a single i.p. injection of NNK (10 mumol/mouse). Sixteen weeks following NNK administration, the experiment was terminated and pulmonary adenomas were quantitated. Pretreatment with PEITC at a dose of 5 mumol or PHITC at a dose of 0.2 mumol resulted in significant reductions of tumor multiplicity compared to control, regardless of whether each isothiocyanate was administered once or four times. For both isothiocyanates, there were no statistically significant differences between dosing frequencies in inhibitory effects on tumor multiplicities and tumor incidences. Furthermore, the results achieved following a single pretreatment with either isothiocyanate were in good agreement with previous results obtained utilizing the four-dose protocol. Thus, it appears that most or all of the inhibitory potential of the four-dose protocol is due to the final dose of isothiocyanate.

Structure-activity relationships for inhibition of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone lung tumorigenesis by arylalkyl isothiocyanates in A/J mice.

Phenethyl isothiocyanate (PEITC), 3-phenylpropyl isothiocyanate (PPITC), 4-phenylbutyl isothiocyanate (PBITC), and the newly synthesized 5-phenylpentyl isothiocyanate (PPeITC), 6-phenylhexyl isothiocyanate (PHITC), and 4-(3-pyridyl)butyl isothiocyanate (PyBITC) were tested for their abilities to inhibit tumorigenicity and DNA methylation induced by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in the lungs of A/J mice. Mice were administered isothiocyanates by gavage for 4 consecutive days at doses of 5, 1, or 0.2 mumol/day prior to administration of 10 mumol of NNK by i.p. injection. Mice were sacrificed 16 weeks after NNK administration and pulmonary adenomas were quantitated, PEITC effectively inhibited NNK-induced lung tumors at a dose of 5 mumol/day but was not inhibitory at doses of 1 or 0.2 mumol/day. PPITC, PBITC, PPeITC, and PHITC were all considerably more potent inhibitors of NNK lung tumorigenesis than PEITC. While virtually no differences in inhibitory activity could be ascertained for PPITC, PBITC, and PPeITC, PHITC appeared to be the most potent tumor inhibitor of all of the compounds. At a dose of 0.2 mumol/day, PHITC pretreatment reduced tumor multiplicity by 85%. PyBITC, an analogue of both NNK and PBITC, was ineffective as an inhibitor. Using the same protocol, the compounds were found to have qualitatively similar inhibitory effects on NNK-induced DNA methylation when administered at 1 mumol/day. These results extend our previous findings that increased alkyl chain length enhances the inhibitory activity of an arylalkyl isothiocyanate toward NNK lung tumorigenesis and demonstrate the exceptional chemopreventive potentials of two new isothiocyanates, PPeITC and PHITC.

Characterization of a glucuronide metabolite of 4-(methyl-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and its dose-dependent excretion in the urine of mice and rats.

Following analysis by reversed-phase HPLC, a previously uncharacterized metabolite of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) was found in the urine of A/J mice treated with NNK. Treatment with beta-glucuronidase converted the metabolite to a peak that co-eluted with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL). Treatment with sulfatase or beta-glucuronidase plus saccharic acid 1,4-lactone did not change the retention time of the metabolite. These data suggested that the unknown metabolite was a glucuronic acid conjugate of NNAL. Upon isolation and purification of larger quantities of the metabolite from the urine of A/J mice, CD-1 mice and F344 rats, 1H and 13C NMR and MS confirmed that the unknown metabolite was 4-(methylnitrosamino)-1-(3-pyridyl)-1-butyl beta-D-glucopyranosiduronic acid (NNAL Glu). To determine the quantitative relationship between NNK dose and NNAL Glu production and to compare the importance of glucuronidation relative to other metabolic pathways, [5-3H]NNK was administered to F344 rats and A/J mice at doses of 500-0.005 mumol/kg. At 500 mumol/kg, NNAL Glu accounted for 22% of the total urinary excretion of NNK in A/J mice, and for 8% in F344 rats 48 h after dosing. The proportions of excreted glucuronide and NNAL decreased with diminishing doses of NNK, yielding undetectable levels of each metabolite in both mice and rats at a dose of 0.005 mumol/kg NNK. Since substantial amounts of metabolites formed via alpha-hydroxylation and N-oxidation pathways were observed at the lower doses of NNK, these data demonstrate that NNAL glucuronidation is a quantitatively unimportant metabolic pathway at low doses of NNK.

Effects of indole-3-carbinol on lung tumorigenesis and DNA methylation induced by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and on the metabolism and disposition of NNK in A/J mice.

The effects of indole-3-carbinol (I3C) on lung neoplasia induced by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) were assessed in an A/J mouse pulmonary adenoma bioassay. Mice were administered corn oil or I3C (25 or 125 mumol/mouse/day) by gavage for 4 consecutive days. Two h after the final pretreatment, mice were administered a single dose of NNK (10 mumol/mouse) i.p. Pulmonary adenomas were quantitated 16 wk after NNK dosing. Mice pretreated with corn oil developed 10.7 tumors/mouse; I3C pretreatment at either dose level inhibited tumor multiplicity by approximately 40%. The effects of I3C on NNK-induced DNA methylation in the lungs and livers of A/J mice were assessed using the same dosing regimen as in the bioassay. Both dose levels of I3C inhibited pulmonary O6-methylguanine formation by at least 50%, but enhanced hepatic DNA methylation at 2 or at 6 h after NNK administration. The effects of I3C pretreatment on NNK metabolism were also investigated. Hepatic microsomes of I3C-pretreated mice showed increased formation of alpha-hydroxylation products, while no significant effect of I3C pretreatment was observed in pulmonary microsomes. The effects of I3C on [5-3H]NNK disposition were also evaluated. I3C pretreatment produced lower levels of total radioactivity in the lung when compared with controls. Additionally, lower proportions of NNK and its carcinogenic metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol were found in the lungs of I3C-pretreated mice. These results demonstrate that I3C inhibits NNK-induced lung neoplasia in A/J mice and suggest that the basis of this inhibition is the decrease in O6-methylguanine formation in A/J lung caused by I3C pretreatment. This decrease in lung DNA methylation appears to be due to the decreased bioavailability of NNK and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in the lungs of I3C-treated mice which, in turn, may be a result of increased metabolic alpha-hydroxylation of NNK by the liver.

Effect of dietary aromatic isothiocyanates fed subsequent to the administration of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone on lung tumorigenicity in mice.

Naturally-occurring aromatic isothiocyanates, benzyl isothiocyanate (BITC) and phenethyl isothiocyanate (PEITC), were tested for their post-treatment effects on lung tumorigenicity by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in A/J mice. Mice at 7 weeks of age were administered a single i.p. dose of NNK (10 mumol/mouse). One week after NNK dosing, mice were placed on AIN-76A diet containing 1 or 3 mumol/g diet of BITC or PEITC. The control group was maintained on AIN-76A diet after NNK administration. Mice were killed 16 weeks after NNK treatment and lung adenomas were counted. The results showed mice fed control diet developed 7.8 tumors/mouse. Mice fed PEITC at concentrations of 1 or 3 mumol/g diet had 8.2 or 6.1 tumors/mouse, respectively. Feeding BITC at 1 mumol/g diet resulted in a tumor yield of 8.0 tumors/mouse, whereas BITC diet at 3 mumol/g diet gave 5.2 tumors/mouse, a small but significant inhibition. However, in the high BITC dose group, a loss in weight gain due to reduced food intake was noted. The results of this study showed that post-treatment of aromatic isothiocyanates had little, if any, effect on NNK lung tumorigenicity in A/J mice. This is in contrast to our previous findings in which pretreatment with PEITC greatly inhibited lung tumor induction by NNK in A/J mice and suggests that tumor inhibition by PEITC is due to inhibition of NNK metabolic activation.

Effects of alkyl chain length on the inhibition of NNK-induced lung neoplasia in A/J mice by arylalkyl isothiocyanates.

Six homologous arylakyl isothiocyanates were evaluated for their abilities to inhibit pulmonary adenomas induced by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in A/J mice. Four consecutive daily doses (5 mumol/mouse) of phenyl isothiocyanate (PITC), benzyl isothiocyanate (BITC), phenethyl isothiocyanate (PEITC), 3-phenylpropyl isothiocyanate (PPITC), 4-phenylbutyl isothiocyanate (PBITC), 4-oxo-4-(3-pyridyl)-butyl isothiocyanate (OPBITC) and corn oil were administered to mice by gavage. Two hours following the final dosing, mice were administered saline or 10 mumol of NNK in saline i.p. Pulmonary adenomas were counted at 16 weeks after NNK administration. The mice administered only corn oil prior to NNK developed an average multiplicity of 9.2 tumors/mouse. Pretreatment with PITC, BITC and OPBITC had no significant effects on NNK-induced lung neoplasia. However, PEITC pretreatment resulted in a 64% reduction of lung tumor multiplicity, but did not affect the percentage of mice that developed tumors. Both PPITC and PBITC decreased tumor multiplicity by 96% and the percentage of tumor-bearing animals by greater than 60%. These results, in conjunction with our previous work, demonstrate a general trend of increasing inhibition of NNK-induced lung neoplasia by arylalkyl isothiocyanates with increasing alkyl chain length. This study also demonstrates the remarkable inhibitory activities of PPITC and PBITC, two isothiocyanates that had not previously been tested as chemopreventive agents.

Effects of aromatic isothiocyanates on tumorigenicity, O6-methylguanine formation, and metabolism of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in A/J mouse lung.

Phenethyl isothiocyanate (PEITC), benzyl isothiocyanate (BITC), and phenyl isothiocyanate (PITC) were tested for their abilities to inhibit lung tumorigenesis and O6-methylguanine formation in lung DNA induced by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in A/J mice. Pretreatment with PEITC for 4 consecutive days at daily doses of 5 or 25 mumol inhibited tumor multiplicity induced by a single 10-mumol dose of NNK by approximately 70% or 97%, respectively. The 25-mumol daily dose of PEITC also reduced the percentage of animals that developed tumors by 70%. In contrast, both BITC and PITC failed to significantly reduce tumor multiplicity or the percentages of mice that developed tumors. Using an identical dosing regimen, parallel results were observed in the effects of these isothiocyanates on O6-methylguanine formation in the lung, in which PEITC at either dose resulted in considerable inhibition at 2 or 6 h after NNK administration, while BITC or PITC had little effect. PEITC was further tested for its ability to inhibit lung microsomal metabolism of NNK. A single administration of PEITC (5 or 25 mumol) resulted in 90% inhibition of NNK metabolism. These results in conjunction with recent results obtained using F344 rats firmly establish PEITC as an effective inhibitor of NNK lung tumorigenesis and suggest that the basis of this inhibition is the reduction of DNA adduct formation caused by the inhibition of enzymes responsible for NNK activation.

Inhibition of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced DNA adduct formation and tumorigenicity in the lung of F344 rats by dietary phenethyl isothiocyanate.

F344 rats fed diets containing phenethyl isothiocyanate (PEITC, 3 mumol/g diet), a cruciferous vegetable component, before and during treatment with the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), developed about 50% fewer lung tumors than NNK-treated rats fed control diets. NNK-induced liver and nasal cavity tumors in rats were, however, not affected by this dietary treatment. The effects of PEITC diets on the formation of DNA adducts by NNK were also investigated in these target tissues. DNA methylation and pyridyloxobutylation by NNK were both decreased by 50% in lung of rats fed PEITC diets compared to that of rats fed control diets, but the levels of DNA methylation were not affected in liver and nasal mucosa. These results correlated with those from the carcinogenicity bioassay, suggesting that DNA alkylations could be used as indicators for screening inhibitors of NNK tumorigenesis. A slight increase in the number of tumors of the exocrine pancreas was observed in PEITC-fed rats with or without NNK treatments. However, these incidences were not statistically significant when compared to the control groups. The potential toxicity of PEITC at concentrations ranging from 0.75 mumol to 6 mumol/g diet was evaluated in a 13-week study. The only toxicity caused by this treatment was minimal fatty metamorphosis in the liver. Considering the widespread human exposure to NNK through tobacco use, it is of practical importance to demonstrate inhibition of lung tumors induced by this carcinogen. These results provide a basis for studies designed to discover agents of better efficacy for the prevention of NNK-induced tumorigenesis.

Effects of dietary sinigrin or indole-3-carbinol on O6-methylguanine-DNA-transmethylase activity and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced DNA methylation and tumorigenicity in F344 rats.

The effects of dietary sinigrin and indole-3-carbinol (I3C) on DNA methylation and O6-methylguanine--DNA-transmethylase activity, factors which may be of importance in the induction of tumorigenicity by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), were investigated. Additionally, the effects of dietary sinigrin on NNK tumorigenicity were assessed in a two-year bioassay in F344 rats. DNA methylation in target tissues of NNK tumorigenesis was examined in F344 rats administered [3H-CH3]NNK (0.6 mg/kg, four doses) s.c. and fed control or experimental diets for two weeks. Dietary sinigrin at a concentration of 3 mumol/g diet decreased 7-methylguanine formation in hepatic DNA, but had no effect on 7-methylguanine levels of lung or nasal mucosa DNA. Dietary I3C at a concentration of 30 mumol/g diet increased 7-methylguanine levels in hepatic DNA, but decreased DNA methylation in lung and nasal mucosa. No effects on O6-methylguanine--DNA-transmethylase activity were observed in tissue extracts derived from the livers, lungs and nasal mucosae of rats fed diets containing sinigrin or I3C. These results suggested that dietary sinigrin might reduce the incidence of NNK-induced hepatic tumors with no effect on NNK tumorigenesis of the lung and nasal cavity, whereas I3C might increase hepatic tumor incidence and reduce NNK tumorigenesis of the lung and nasal cavity. The bioassay results showed that dietary sinigrin had no effect on NNK tumorigenesis in these target tissues. However, dietary sinigrin plus NNK resulted in a significant incidence of pancreatic tumors, a rare occurrence in F344 rats. While the results from DNA methylation studies are in agreement with the bioassay data for lung and nasal cavity, the absence of any inhibitory effect of dietary sinigrin on NNK hepatic tumorigenesis indicates that factors other than DNA methylation and O6-methylguanine repair should be considered in assessing the effects of dietary compounds on NNK hepatic tumorigenesis. The contrary effects on NNK-induced hepatic DNA methylation by sinigrin and I3C, two major components of cruciferous vegetables, demonstrate the complexities of dietary modulation of carcinogenesis.