Mechanistic Approaches To The Prevention Of Mutation And Cancer

Cancer and other mutation-related diseases can be prevented at three levels: primary prevention, which is addressed to healthy individuals in order to prevent occurrence of the disease; secondary prevention, which is addressed to early stage patients in order to prevent progression of the disease; and tertiary prevention, which is addressed to patients after therapy in order to prevent relapses of the disease. Although the most obvious approach to prevention is to minimize exposures to recognized risk factors, a complementary strategy is represented by chemoprevention, using dietary and pharmacological agents that reinforce the host defence machinery. Since 1988, I proposed detailed classifications of mechanisms of inhibitors of mutagenesis and carcinogenesis. They may apply not only to cancer but also to other degenerative diseases that have replaced infectious diseases as the leading causes of death in the population. In fact, certain mechanisms, such as damage to nuclear DNA and mtDNA, oxidative stress, chronic inflammation, signal transduction alterations and epigenetic changes may be involved in the pathogenesis of different diseases. Studies performed in our laboratory have shown that certain genomic alterations that are usually investigated in cancer research may also be detected in other chronic diseases, such as atherosclerosis, degenerative heart diseases, chronic obstructive pulmonary diseases, neurological disorders, eye diseases, skin ageing, and alopecia. Similar alterations were investigated in critical periods of life, such as birth and ageing. The nucleotide alterations occurring at birth in the lung render the newborn particularly vulnerable to the action of environmental agents. In fact, we demonstrated that cigarette smoke becomes a potent carcinogen in mice when exposure starts at birth and continues early in life. We investigated a number of chemopreventive agents by evaluating modulation of intermediate biomarkers and carcinogenicity. An optimal agent should not excessively alter the physiological patterns of gene expression, microRNA and proteome profiles, but at the same time it should be effective in inhibiting alterations induced by mutagens and carcinogens. It should be noted that most chemopreventive agents possess pleiotropic properties. The knowledge of mechanisms can be exploited to combine different agents working with complementary mechanisms. Like the therapy of important diseases, such as cancer, cardiovascular diseases, AIDS, etc., uses combinations of drugs, combined chemoprevention is a quite promising strategy. Although a large number of agents are potentially able to prevent cancer, we are in search of tools to predict, hopefully in the single individual, their efficacy and safety in humans

The generation of DNA single-strand breaks during the reduction of chromate by ascorbic acid and/or glutathione in vitro.

The potential role of iron and copper and the involvement of hydroxyl radicals in the DNA cleavage caused by chromate and glutathione (GSH) has been investigated. We have also studied the ability of chromate, on reaction with ascorbate as well as in mixed solutions of ascorbate and GSH, to cause DNA strand breaks. In both fully demetalated and conventional (i.e., metal contaminated) systems, chromate and GSH induced similar numbers of DNA strand breaks. This observation suggests that traces of iron or copper contaminating the reaction mixtures do not play a major role in the DNA cleavage caused by chromate and GSH. A series of hydroxyl radical scavengers exhibited a protective influence on the induction of DNA strand breaks. However, glucose and sucrose, both strong hydroxyl radical scavengers, showed no concentration-dependent inhibition of DNA cleavage. Competition kinetics studies yielded apparent rate constants that were not consistent with hydroxyl radicals being the species responsible for DNA strand breaks. Ascorbate in combination with chromate was also found to induce strand breaks in DNA; this damage could be attributed to reactive intermediates generated during the reduction. When mixed systems of ascorbate and GSH in the presence of chromate were investigated, there were clearly interactions between the two reductants

Attenuation of influenza-like symptomatology and improvement of cell-mediated immunity with longterm N-acetylcysteine treatment

ABSTRACT: N-acetylcysteine (NAC), an analogue and precursor of reduced glutathione, has been in clinical use for more than 30 yrs as a mucolytic drug. It has also been proposed for and/or used in the therapy and/or prevention of several respiratory diseases and of diseases involving an oxidative stress, in general. The objective of the present study was to evaluate the effect of long-term treatment with NAC on influenza and influenza-like episodes. A total of 262 subjects of both sexes (78% ≥65 yrs, and 62% suffering from nonrespiratory chronic degenerative diseases) were enrolled in a randomized, doubleblind trial involving 20 Italian Centres. They were randomized to receive either placebo or NAC tablets (600 mg) twice daily for 6 months. Patients suffering from chronic respiratory diseases were not eligible, to avoid possible confounding by an effect of NAC on respiratory symptoms. NAC treatment was well tolerated and resulted in a significant decrease in the frequency of influenza-like episodes, severity, and length of time confined to bed. Both local and systemic symptoms were sharply and significantly reduced in the NAC group. Frequency of seroconversion towards A/H 1 N 1 Singapore 6/86 influenza virus was similar in the two groups, but only 25% of virus-infected subjects under NAC treatment developed a symptomatic form, versus 79% in the placebo group. Evaluation of cell-mediated immunity showed a progressive, significant shift from anergy to normoergy following NAC treatment. Administration of N-acetylcysteine during the winter, thus, appears to provide a significant attenuation of influenza and influenza-like episodes, especially in elderly high-risk individuals. N-acetylcysteine did not prevent A/H 1 N 1 virus influenza infection but significantly reduced the incidence of clinically apparent disease. Eur Respir J 1997; 10: 1535-154

Extracellular Vesicles in Biological Fluids. A Biomarker of Exposure to Cigarette Smoke and Treatment with Chemopreventive drugs

Extracellular vesicles (EVs) are released from cells and enter into body fluids thereby providing a toxicological mechanism of cell-cell communication. The present study aimed at assessing (a) the presence of EVs in mouse body fluids under physiological conditions, (b) the effect of exposure of mice to cigarette smoke for 8 weeks, and (c) modulation of smoke-related alterations by the nonsteroidal anti-inflammatory drug celecoxib, a selective cyclooxygenase-2 inhibitor. To this purpose, ICR (CD-1) mice were either unexposed or exposed to cigarette smoke, either treated or untreated with oral celecoxib. EVs, isolated from bronchoalveolar lavage fluid (BALF), blood serum, and urines, were analyzed by nanoparticle tracking analysis and flow cytometry. EVs baseline concentrations in BALF were remarkably high. Larger EVs were detected in urines. Smoking increased EVs concentrations but only in BALF. Celecoxib remarkably increased EVs concentrations in the blood serum of both male and female smoking mice. The concentration of EVs positive for EpCAM, a mediator of cell-cell adhesion in epithelia playing a role in tumorigenesis, was much higher in urines than in BALF, and celecoxib significantly decreased their concentration. Thus, the effects of smoke on EVs concentrations were well detectable in the extracellular environment of the respiratory tract, where they could behave as delivery carriers to target cells. Celecoxib exerted both protective mechanisms in the urinary tract and adverse systemic effects of likely hepatotoxic origin in smoke-exposed mice. Detection of EVs in body fluids may provide an early diagnostic tool and an end-point exploitable for preventive medicine strategies.

Carcinogen metabolism in human lung tissues and the effect of tobacco smoking: results from a case--control multicenter study on lung cancer patients.

Cigarette smoking is the strongest risk factor for lung cancer, but genetically determined variations in the activities of pulmonary enzyme that metabolize tobacco-derived carcinogens may affect individual risk. To investigate whether these enzymes (e.g., CYP1A-related) can serve as markers for carcinogen-DNA damage, lung tissue specimens were taken during surgery from middle-aged men with either lung cancer or non-neoplastic lung disease. Phase I [aryl hydrocarbon hydroxylase (AHH), ethoxycoumarin O-deethylase (ECOD)] and phase II (epoxide hydrolase, UDP-glucuronosyltransferase, glutathione S-transferase) enzyme activities, glutathione and malondialdehyde contents were determined in lung parenchyma and/or bronchial tissues; some samples were also analyzed for DNA adducts, using 32P-postlabeling. The data were then analyzed for the following: a) differences in metabolic profiles between bronchial and parenchymal lung tissue; b) the effect of recent exposure to tobacco smoke on enzyme inducibility and benzo[a]pyrene metabolism; c) differences in enzyme inducibility between lung cancer and non-lung cancer patients; d) the effect of smoking on metabolism of mutagens in vitro; e) pulmonary DNA adduct levels and AHH activity in lung parenchyma of smokers and ex-smokers; f) lipid peroxidation products in lung tissue from lung cancer and non-lung cancer patients, as related to smoking habits and degree of airway obstruction; and g) prognostic value of AHH pulmonary activity in lung cancer patients. The results demonstrate a pronounced effect of tobacco smoke on pulmonary metabolism of xenobiotics and prooxidant state and suggest the existence of a metabolic phenotype at higher risk for tobacco-associated lung cancer

Chromium(VI) reduction by ascorbate: role of reactive intermediates in DNA damage in vitro.

Reaction of chromium(VI) with one equivalent of ascorbate was studied by electron paramagnetic resonance spectroscopy in the presence of 0.10 M 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) at room temperature in 0.10 M (N-[2-hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid]) (HEPES) and 0.05 M tris(hydroxymethyl)aminomethane hydrochloride (Tris-HCl) buffers (pH 7.0 room temperature). Chromium(V), ascorbyl radical, and carbon-based DMPO-radical adducts were observed. A higher level of Cr(V) was observed in HEPES buffer and a higher level of the DMPO-radical adducts was observed in Tris-HCl buffer. Chromium-DNA binding studies were carried out in vitro for calf thymus DNA incubated with Cr(VI) and ascorbate in both buffers at 37 degrees C. Higher Cr-DNA binding was observed in HEPES buffer. DNA strand-break studies were carried out in vitro on pBR322 DNA incubated with Cr(VI) and ascorbate in both buffers at 37 degrees C. Higher percent nicking was observed in Tris-HCl buffer. Addition of DMPO decreased nicking levels in Tris-HCl buffer. These results suggest that free radicals are more reactive than Cr(V) in producing DNA strand breaks and that Cr(V) will react with DNA to produce Cr-DNA adducts. The fact that buffer affects the nature of the reactive intermediates produced upon reduction of Cr(VI) may be related to differences in intracellular metabolism of Cr(VI) and resulting DNA damage observed in various cell culture systems and animal tissues in vivo