Chemoprevention of tobacco-smoke lung carcinogenesis in mice after cessation of smoke exposure

Hanspeter Witschi 0 Dale Uyeminami 0 Dexter Moran 0 Imelda Espiritu 0 0 Institute of Toxicology and Environmental Health and Department of Molecular Biosciences, School of Veterinary Medicine, University of California , One Shields Avenue, Davis, CA 95616, USA 1To whom correspondence should be addressed Email: Male strain A/J mice were exposed for 6 h per day, 5 days per week to a mixture of 89% cigarette sidestream smoke and 11% mainstream smoke. Total suspended particulate concentrations were 137 mg/m3. In experiment 1, animals were exposed for 5 months to tobacco smoke and given a 4 month recovery period in air. Lung tumor multiplicity was 2.4 and incidence 89%. Animals exposed to filtered air had 1.0 tumor per lung (65% incidence). In animals kept for 5 months in smoke, removed into air and then fed a diet containing a mixture of myoinositol and dexamethasone, tumor multiplicity was 1.0 and incidence was 62%. These values were significantly (P < 0.01) lower than in animals exposed to smoke and identical to values seen in controls. In animals fed a diet containing 250 mg/kg each of phenethyl isothiocyanate and benzyl isothiocyanate during the entire 9 months, lung tumor multiplicity was 2.1 and incidence 96%, not significantly different from animals exposed to smoke and fed control diet. In experiment 2, animals were exposed for 5 months to smoke, followed by a 4 month recovery period in air and were fed during the entire period a diet containing either D-limonene or 1,4-phenylenebis(methylene)selenoisocyanate (p-XSC). In animals exposed to tobacco smoke and fed control diet, lung tumor multiplicity was 2.8, whereas in the animals fed D-limonene it was 2.6 and in the animals fed p-XSC it was 2.4. The differences to the controls were statistically not significant. It was concluded that myoinositol-dexamethasone successfully prevents the development of tobacco smoke-induced lung tumors even if administered when the animals have 'quit' smoking. On the other hand, agents otherwise shown to prevent lung tumor formation following administration of 4-(methylnitrosamino)-1-(3-pyridyl)-1butanone or benzo[a]pyrene were ineffective against tobacco smoke. Abbreviations: BITC, benzyl isothiocyanate; NNK, 4-(methylnitrosamino)1-(3-pyridyl)-1-butanone; p-XSC, 1,4-phenylenebis(methylene)selenoisocyanate; PEITC, phenethyl isothiocyanate. - The incidence and death rates of lung cancer in the USA have decreased in men. In females the annual increase appears to have slowed down. There is some concern, however, that changing smoking habits of teens and the substitution of cigarettes by cigars may possibly reverse this trend (1). In other countries of the world, particularly in Asia, the lung cancer epidemic continues to spread and tobacco eventually will kill millions of people (24). Complete cessation of smoking would be the most effective way to prevent this burden on the public health. It is recognized that a substantial number of smokers is unable to quit. Chemoprevention might be a possible way to mitigate the impact of smoking (510). Chemopreventive agents, such as drugs or naturally occurring constituents of the diet, might provide some protection to active smokers. Perhaps more importantly, they might help to reduce further the risk of developing lung cancer in individuals who have quit smoking or in individuals exposed to tobacco smoke in their environment. During the last few years, a substantial number of agents has been examined for their possible effects on lung cancer chemoprevention. Practically all experiments used animals treated with lung-tumor-producing chemicals, most often tobacco-specific nitrosamines or polycyclic aromatic hydrocarbons (PAHs). Lung tumors in strain A/J mice have become the preferred test system. (11). These tumors resemble in many ways human lung adenocarcinoma (12), a tumor that during the past decades has been found with increasing frequency in man (13,14). The experimental model is attractive as a screening tool because a substantial overlap exists between man and mouse in the genetic alterations thought to be responsible for lung tumorigenesis (15). Already it has been firmly established that lung tumorigenesis in strain A/J mice can effectively be prevented by a large variety of chemicals. The carcinogenic action of tobacco-specific nitrosamines or PAHs can be counteracted by isothiocyanates (16), non-steroidal anti-inflammatory agents (17), green and black tea (18), organoselenium compounds (19), glucocorticoid hormones (20,21), perillyl alcohol (22) and others. On occasion, some doubts have been raised whether studies with model compounds may substitute exposure to the full and complex mixture of tobacco smoke (15,23). We have addressed this problem in a series of experiments in which we examined the effects of chemopreventive agents on lung tumors induced in strain A/J mice by a mixture of 89% sidestream smoke and 11% mainstream smoke (24,25). So far we found that green tea, acetylsalicylic acid and N-acetyl cysteine were ineffective. Phenethyl isothiocyanate (PEITC) had a possible marginal, but statistically not significant effect. The only agent that was found to be highly effective against tobacco smoke was a combination of dietary myoinositol and dexamethasone (25). In the present investigation we examined three additional chemopreventive agents, D-limonene, 1,4-phenylenebis (methylene) selenoisocyanate (p-XSC) and a mixture of PEITC and benzyl isothiocyanate (BITC). We also examined whether it would be possible to achieve chemoprevention with myoinositoldexamethasone in ex-smokers, i.e. in mice that were only fed the chemopreventive diet once they were removed from the smoke atmosphere. Materials and methods Animals Male strain A/J mice, 68 weeks old, were purchased from Jackson Laboratories, Bar Harbor, ME. Randomly chosen animals were sent to the Comparative Pathology Laboratory, UC Davis, for a standard rodent health surveillance screen. No evidence for infectious disease (pathogenic agents) or presence of parasites or ova in pelage and cecum were reported. Histopathology was not processed since no significant lesions were noted. Serology was negative for mouse hepatitis virus, Sendai virus, Reovirus type 3, pneumonia virus, parvo, ectromelia and mycoplasma pulmonis. The animals were housed, four to a cage, in polypropylene cages with tightly fitting wire screen lids on conventional bedding material. At all times during the experiment, including during smoke exposure, water and the test diets were provided ad libitum. The animals were monitored daily and weighed weekly. Kentucky 1R4F reference cigarettes were purchased from the Tobacco Research Institute, University of Kentucky, Lexington, KY. PEITC, BITC, myoinositol, dexamethasone, D-limonene and corn oil were obtained from Sigma Chemical Co., St Louis, MO. Anhydrous acetone and ,-debromo-p-xylene were obtained from Aldrich Chemicals, Milwaukee, WI, and KSeCN from Acros (...truncated)