EP2129244A2 - Tabac et matériau de conditionnement de tabac pour empêcher ou réduire une lésion associée au tabac dans les voies respiratoires et le tube digestif d'un sujet - Google Patents

Tabac et matériau de conditionnement de tabac pour empêcher ou réduire une lésion associée au tabac dans les voies respiratoires et le tube digestif d'un sujet

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Publication number
EP2129244A2
EP2129244A2 EP08702682A EP08702682A EP2129244A2 EP 2129244 A2 EP2129244 A2 EP 2129244A2 EP 08702682 A EP08702682 A EP 08702682A EP 08702682 A EP08702682 A EP 08702682A EP 2129244 A2 EP2129244 A2 EP 2129244A2
Authority
EP
European Patent Office
Prior art keywords
tobacco
saliva
activity
opo
manufacturing
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP08702682A
Other languages
German (de)
English (en)
Inventor
Abraham Z. Reznick
Rafael M. Nagler
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Technion Research and Development Foundation Ltd
Original Assignee
Technion Research and Development Foundation Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Technion Research and Development Foundation Ltd filed Critical Technion Research and Development Foundation Ltd
Publication of EP2129244A2 publication Critical patent/EP2129244A2/fr
Withdrawn legal-status Critical Current

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Classifications

    • A—HUMAN NECESSITIES
    • A24—TOBACCO; CIGARS; CIGARETTES; SIMULATED SMOKING DEVICES; SMOKERS' REQUISITES
    • A24B—MANUFACTURE OR PREPARATION OF TOBACCO FOR SMOKING OR CHEWING; TOBACCO; SNUFF
    • A24B15/00—Chemical features or treatment of tobacco; Tobacco substitutes, e.g. in liquid form
    • A24B15/18—Treatment of tobacco products or tobacco substitutes
    • A24B15/28—Treatment of tobacco products or tobacco substitutes by chemical substances
    • A24B15/30—Treatment of tobacco products or tobacco substitutes by chemical substances by organic substances
    • A—HUMAN NECESSITIES
    • A24—TOBACCO; CIGARS; CIGARETTES; SIMULATED SMOKING DEVICES; SMOKERS' REQUISITES
    • A24B—MANUFACTURE OR PREPARATION OF TOBACCO FOR SMOKING OR CHEWING; TOBACCO; SNUFF
    • A24B13/00—Tobacco for pipes, for cigars, e.g. cigar inserts, or for cigarettes; Chewing tobacco; Snuff
    • A—HUMAN NECESSITIES
    • A24—TOBACCO; CIGARS; CIGARETTES; SIMULATED SMOKING DEVICES; SMOKERS' REQUISITES
    • A24B—MANUFACTURE OR PREPARATION OF TOBACCO FOR SMOKING OR CHEWING; TOBACCO; SNUFF
    • A24B15/00—Chemical features or treatment of tobacco; Tobacco substitutes, e.g. in liquid form
    • A24B15/18—Treatment of tobacco products or tobacco substitutes
    • A24B15/28—Treatment of tobacco products or tobacco substitutes by chemical substances
    • A24B15/281—Treatment of tobacco products or tobacco substitutes by chemical substances the action of the chemical substances being delayed
    • A24B15/282—Treatment of tobacco products or tobacco substitutes by chemical substances the action of the chemical substances being delayed by indirect addition of the chemical substances, e.g. in the wrapper, in the case
    • A—HUMAN NECESSITIES
    • A24—TOBACCO; CIGARS; CIGARETTES; SIMULATED SMOKING DEVICES; SMOKERS' REQUISITES
    • A24D—CIGARS; CIGARETTES; TOBACCO SMOKE FILTERS; MOUTHPIECES FOR CIGARS OR CIGARETTES; MANUFACTURE OF TOBACCO SMOKE FILTERS OR MOUTHPIECES
    • A24D1/00—Cigars; Cigarettes
    • A24D1/02—Cigars; Cigarettes with special covers
    • A—HUMAN NECESSITIES
    • A24—TOBACCO; CIGARS; CIGARETTES; SIMULATED SMOKING DEVICES; SMOKERS' REQUISITES
    • A24D—CIGARS; CIGARETTES; TOBACCO SMOKE FILTERS; MOUTHPIECES FOR CIGARS OR CIGARETTES; MANUFACTURE OF TOBACCO SMOKE FILTERS OR MOUTHPIECES
    • A24D3/00—Tobacco smoke filters, e.g. filter-tips, filtering inserts; Filters specially adapted for simulated smoking devices; Mouthpieces for cigars or cigarettes
    • A24D3/06—Use of materials for tobacco smoke filters
    • A24D3/14—Use of materials for tobacco smoke filters of organic materials as additive

Definitions

  • the present invention relates to methods of preventing or reducing pathogenesis of oxidant stress-associated diseases of the aerodigestive tract. More particularly, the present invention relates to methods of employing hydroxocobalamin (vitamin B 12a, OH-CO), deferoxamine (DES) and reduced glutathione (GSH) to reduce or prevent tobacco-induced cellular or macromolecular damage in the aerodigestive tract.
  • hydroxocobalamin vitamin B 12a, OH-CO
  • DES deferoxamine
  • GSH reduced glutathione
  • tobacco consumption leads to development or enhancement of atherosclerosis, cardiovascular disease, chronic obstructive pulmonary disease and various forms of cancer, including carcinomas of the mouth, pharynx, esophagus and lung.
  • snuff is a fine-grain tobacco that often comes in teabag-like pouches, which users "pinch” or "dip” between their lower lip and gum.
  • Chewing tobacco comes in shredded, twisted, or "bricked” tobacco leaves that users put between their cheek and gum. Whether it's snuff, snus or chewing tobacco, the user consumes the tobacco letting it sit in the mouth and suck on the tobacco juices, spitting often to get rid of the saliva that builds up.
  • TS tobacco smoke
  • Oxygen free radicals include the superoxide free radical (/O2 ⁇ ) and the hydroxyl radical (OH-) which, together with hydrogen peroxide (H 2 O 2 ) and singlet oxygen (*O2), are jointly called reactive oxygen species (ROS). Due to their high reactivity they may lead to chemical modification and impairment of the components of living cells, such as proteins, lipids, carbohydrates and nucleotides.
  • ROS reactive oxygen species
  • Tobacco smoke mediated oxidant injury is similar to that induced by smog, thereby increasing such noxious stimuli to primary and secondary smokers in polluted atmospheric environments.
  • mainstream and side stream TS Most of constituents of TS have been identified in so-called mainstream and side stream TS.
  • the former is that volume of smoke drawn through the mouthpiece of the tobacco product during puffing while side stream smoke is that smoke emitted from the smoldering cigarette in between puffs.
  • mainstream smoke emission is also markedly reduced both in low and in ultra low tar yield cigarettes.
  • the emissions of toxic and carcinogenic components in side stream smoke are not significantly reduced in filter cigarettes when compared to non-filter counterparts.
  • side stream smoke is a major contributor to environmental smoke, affecting both the smoker and their non-smoking counterparts, so called secondary smokers.
  • Cigar smokers may spend one full hour smoking a single large cigar and commonly hold an unlit cigar in the mouth, allowing further exposure to toxins by local absorption. Thus, consumption of cigars may produce an equal or greater burden of toxic exposure to TS than cigarettes. Recently, sales of cigars have risen, partly due to their gaining popularity with women and the advent of the female friendly "cigar bar".
  • Tobacco whether smoked or chewed, causes common untoward effects in the oral cavity. Tobacco smoke has two chances to exert its deleterious effects in the mouth; when it is inhaled by the smoker and on its exit during exhalation.
  • Oral cancer kills 8,000 patients each year and only half of cases diagnosed annually have a five-year survival. The great majority of these patients are users of tobacco products.
  • Oral squamous cell carcinoma (SCC) is the most common malignancy of the head and neck with a worldwide incidence of over 300,000 new cases annually. The disease is characterized by a high rate of morbidity and mortality (approximately 50 %) and in this respect is similar to malignant melanoma (1-4).
  • the major inducer of oral SCC is exposure to tobacco which is considered to be responsible for 50-90 % of cases world-wide (5, 6).
  • leukoplakia is a benign oral lesion, it has a malignant potential.
  • tobacco contributes to other oral symptoms or pathologies of the mouth and teeth.
  • Tobacco may cause halitosis, may numb the taste buds, and interfere with the smell and the taste of food. It may stain teeth and contribute to dental caries.
  • Smokers have more dental tartar (calculus) than non-smokers.
  • Tobacco is associated also with destructive periodontal (gum) disease and tooth loss.
  • Acute necrotizing ulcerative gingivitis (“trench mouth") is a destructive, painful inflammatory condition occurring mainly in tobacco smokers. Swelling of the nasal and sinus membranes have also been associated, purportedly, in individuals who are "allergic" to TS.
  • Oral submucous fibrosis occurs mainly in India and is a chronic, progressive premalignant condition.
  • the etiology is chronic chewing of tobacco or areca nut or both.
  • the fibrosis results in restriction of mouth opening and involves the palates, tonsillar fossa, buccal mucosa and underlying muscle.
  • oropharyngeal carcinomas also with a high frequency in India and associated in 70% of cases with chewing tobacco.
  • Smokeless tobacco and areca nut usage is also common in Pakistan, Bangladesh and Java and in these and Indian immigrants to the United States and United Kingdom.
  • Tobacco smoke induces oxidative damage to lipids, DNA and proteins, particularly via protein-SH groups as a consequence of containing high levels of both free radicals as well as aldehydes, including acetaldehyde (ethanol), propanol and acrolein, as well as other deleterious molecules.
  • Tar contains high concentrations of free radicals.
  • Many tar extracts and oxidants are water-soluble and reduce oxygen to superoxide radical which can dismutate to form the potent oxidant H 2 O 2 .
  • Oxidants in gas-phase smoke are reactive carbon- and oxygen-centered radicals with extremely short half lives.
  • TS elicits protein carbonylation in plasma and that, in contrast, exposure of human plasma to gas-phase but not to whole TS produces oxidative damage to lipids.
  • Redox-active metals Redox-active metal ions, such as iron and copper, in the presence of H 2 O 2 and other low-reactive free radicals found in TS, such as superoxide radicals, participate in the deleterious Haber- Weiss and Fenton reactions, in which the highly reactive hydroxyl free radicals are produced.
  • Aldehydes Studies have shown that exposure of plasma to TS results in protein damage in the form of protein carbonylation (11) and in oxidation of plasma lipids and antioxidants (12). The source of the accumulation of protein carbonylation was found to be due to aldehydes present in TS (13, 14).
  • TS salivary enzymes
  • lactic dehydrogenase (LDH) lactic dehydrogenase
  • acid phosphatase were considerably affected by TS (14, 15), where both TS-based aldehydes, such as acrolein and crotonaldehyde, as well as oxygen free radicals were implicated as the causative agents affecting the above enzymes (14, 15).
  • Physiological antioxidants Glutathione: Glutathione, a sulfur-containing tripeptide (L- ⁇ -glutamyl-1- cysteine-glycine) is the most abundant non-protein thiol in mammalian cells and is recognized as the primordial antioxidant.
  • Glutathione in its reduced form, "GSH" acts as a substrate for glutathione-S-transferase and glutathione peroxidase, enzymes catalyzing reactions involved in detoxification of xenobiotic compounds and in antioxidation of ROS and other free radicals.
  • GSH Glutathione
  • This ubiquitous protein plays a vital function in maintaining the integrity of free radical sensitive cellular components. Under states of GSH depletion, including malnutrition and severe oxidative stress, cells may then become injured from excess free radical damage and die.
  • Oral peroxidation is the pivotal enzymatic activity of the salivary antioxidant system (16-19). Oral peroxidase activity is composed of the combined activity of two peroxidases, salivary peroxidase (SPO) and myeloperoxidase (MPO).
  • SPO salivary peroxidase
  • MPO myeloperoxidase
  • Salivary peroxidase which is secreted by the major salivary glands, mainly the parotid gland (18), contributes 80 % of the total oral peroxidase (OPO) activity, while MPO, produced by leukocytes (20), contributes the remaining 20 % of OPO activity.
  • Oral peroxidase performs two functions preventing oxidant injury; it reduces the level of H 2 O 2 excreted into the oral cavity from the salivary glands, by bacteria and by leukocytes, and it inhibits the metabolism and proliferation of various bacteria in the oral cavity.
  • SCN ⁇ thiocyanate ion
  • H 2 O 2 oxidizes SCN-, a detoxification product of cyanide secreted mainly by the parotid gland.
  • SCN ⁇ acts as the electron-donating component, similarly to GSH in other biological systems (20, 24, 25).
  • HOSCN hydrogen hypothiocyanite
  • OSCN its conjugated anion
  • HOSCN and OSCN stems from their ability to react with sulfhydryl groups of bacterial enzymes that are vital for glycolysis, such as hexokinase, aldolase and pyruvate kinase (20, 25-28).
  • cigarette filters are used to trap TS tar but do not the gas-phase compounds.
  • dipeptide compounds with pharmaceutical properties to increase glutathione levels were employed (34).
  • a further approach utilized a glycine carboxylic acid alkyl mono-ester of glutathione to increase cellular GSH levels (35).
  • the present inventors have previously described novel smoking filters and oral compositions for reducing tobacco associated damage in the aeurodigestive tract (see
  • compositions include active agents which are capable of reducing or preventing tobacco associated loss of peroxidase activity in the aerodigestive tract.
  • U.S. Patent Number 5,922,346 teaches a composition for reducing free radical damage induced by tobacco products and environmental pollutants comprising, as active ingredients, reduced glutathione and a source of selenium selected from the group consisting of elemental selenium, selenomethionine and selenocysteine, the active ingredients being combined with suitable carriers and flavorings for their intra-oral administration as gels, lozenges, tablets and gums in concentrations for reducing free radical damage induced by tobacco products and other environmental pollutants to the oral cavity, pharynx and upper respiratory tract of a user and secondary smokers.
  • Patent Number 5,906,811 teaches a method for reducing free radical damage induced by tobacco products and environmental pollutants comprising administering in a suitable carrier in concentrations for effectively reducing said free radical damage to the oro-pharynx and upper respiratory tract of a user a combination of from 0.01 and 10% (weight) glutathione, from 1.0 to 25% (weight) ascorbic acid, from
  • U.S. Patent No. 5,829,449 teach a composition for inclusion within a cigarette, cigar or pipe tobacco for reducing free radical damage to the oro-pharyngeal cavity, respiratory tract and lungs from tobacco smoke, said composition comprising L- glutathione and a source of selenium selected from the group consisting of L- selenomethionine and L-selenocyste ⁇ ne.
  • U.S. Patent No. 5,829,449 clearly states that the composition is supplied by smoke inhalation and not by direct contact with the aerodigestive tract (i.e., wet tissue).
  • U.S. Patent 6,138,683 teaches a composition for inclusion within a quantity of smokeless tobacco, selected from the group consisting of chewing tobacco and snuff, for reducing free radical induced damage to the oro-pharyngeal cavity of the user, said composition comprising L-glutathione and a source of selenium in combination with said smokeless tobacco.
  • a smoking product comprising an agent being capable of reducing or preventing tobacco associated death of cells in the aerodigestive tract of a subject, the smoking product being designed and configured so as to enable physico-chemical interaction between said agent and said tobacco smoke when in use by the subject.
  • an article of manufacturing comprising tobacco and a tobacco packaging material, at least a portion of said tobacco and/or tobacco packaging material being in contact with an aerodigestive tract of a subject in use thereof, and whereas said at least a portion of said tobacco and/or tobacco packaging material comprises at least one agent capable of reducing or preventing tobacco associated loss of peroxidase activity in said aerodigestive tract, said at least one agent being selected from the group consisting of a cyanide chelator and an iron chelator.
  • an article of manufacturing comprising a tobacco packaging material at least a portion of which being in contact with an aerodigestive tract of a subject in use thereof, wherein said at least a portion comprises an agent capable of reducing or preventing tobacco associated loss of peroxidase activity in said aerodigestive tract.
  • said tobacco packaging material comprises a filter and said at least one agent is impregnated in a paper of said filter being in contact with said aerodigestive tract of said subject in use thereof.
  • said tobacco is smokeless tobacco.
  • said tobacco is smoked tobacco.
  • said tobacco packaging material is selected from the group consisting of a rolling paper, a filter paper, a suns bag packaging, a cigarette, a pipe and a tin sheet packaging.
  • the article of manufacturing is selected from the group consisting of a snuff, a cigarette, a snus, a Gutka, a plug, a twist, a scrap and tobacco water.
  • said agent comprises a cyanide chelator.
  • said agent is hydroxocobalamin. According to still further features in the described preferred embodiments said agent is capable of reducing or preventing tobacco smoke-associated death of cells in the digestive tract of a subject.
  • said cells are lymphocytes and/or epithelial cells.
  • said agent comprises an iron chelator.
  • said agent is deferoxamine. According to still further features in the described preferred embodiments said agent comprises an antioxidant.
  • said agent is glutathione.
  • FIG. 1 is schematic diagram depicting the construction of a filter paper impregnated with agents of the present invention according to the present invention.
  • FIG. 2a is a diagram depicting the pathway of cyanate metabolism resulting from exposure to TS-derived HCN.
  • FIG. 2b is a data plot depicting reduced OPO activity following in vivo exposure (smoking) to TS in smoker and non-smoker subjects. The OPO activity of 17 smokers and 16 non-smokers was measured prior to and following smoking 1 cigarette.
  • FIG. 2c is a data plot depicting reduced OPO activity following in vitro exposure of saliva to TS in smoker and non-smoker subjects. The OPO activity of 7 smokers and 11 non-smokers was measured prior to and following exposure to 1 cigarette.
  • FIG. 3 is a histogram depicting KCSN-mediated resistance to TS exposure- induced decreases in OPO activity in saliva of non-smoker subjects exposed to the
  • TS from 1 cigarette in vitro.
  • Each value represents the average value obtained in experiments on 3 subjects ⁇ standard deviation (SD).
  • FIGs. 4a-b are photographs of Western immunoblotting analysis depicting increased levels of salivary protein carbonylation in representative non-smoker saliva following in vivo exposure of saliva (smoking) to the TS of 1 cigarette ( Figure 4a).
  • FIGs. 5a-b are photographs of Western immunoblotting analysis depicting increased levels of salivary protein carbonylation in representative non-smoker saliva following in vitro exposure of saliva to the TS of 1 cigarette ( Figure 5a). Proof that equal quantities of proteins were analyzed per sample is shown via Coomassie Blue staining ( Figure 5b) of the samples. Lane 1: prior to exposure to TS, Lane 2: 10 mins following exposure to TS, Lane 3: 30 mins following exposure to TS, Lane 4: 60 mins following exposure to TS.
  • FIG. 6 is a data plot depicting TS dose-dependent decrease in OPO activity in saliva exposed in vitro to 3 exposures of TS over a 1 h period. Each data point represents mean + standard error of the means (SEM) of results from experiments on saliva from 4 subjects.
  • FIG. 7 is a data plot depicting the effect of increasing concentrations of dapsone (4-aminophenylsulfone) on OPO activity in TS-treated saliva.
  • Abbreviations: NO.l, NO.2, NO.3, NO4 and NO.5 correspond to saliva from 5 different subjects and plasma corresponds to commercial MPO.
  • FIG. 8 is a data plot depicting the effect on OPO activity in saliva treated with TS in the presence or absence of 150 ⁇ M dapsone. Saliva samples were normalized with respect to initial OPO activity. OPO exposed to TS in the presence
  • FIG. 9 is a data plot depicting the effect of GSH, deferoxamine (DES) and ascorbate on OPO activity in TS-treated saliva.
  • FIG. 10 is a data plot depicting the effect of 100 ⁇ M FeCl 3 or ascorbate on OPO activity in TS-treated saliva. Data points represent data plotted as mean ⁇ SD using saliva samples obtained from 3 subjects.
  • FIG. 11 is a data plot depicting the effect of purified aldehydes present in TS on OPO activity in saliva from 4-5 subjects. Treatment: exposure to air for 3 h (0), 2 mM acetylaldehyde ( ⁇ ), 20 ⁇ M crotonaldehyde ⁇ ), 80 ⁇ M acrolein (X).
  • FIG. 12 is a histogram depicting the effect of 18 h of dialysis on OPO activity in TS-treated saliva having lost 68 % of initial OPO activity levels.
  • FIG. 13 is a histogram depicting recovery of OPO activity in saliva treated. with KCN following 18 the of dialysis.
  • FIG. 14 is a histogram depicting OH-CO mediated prevention of OPO activity loss in saliva resulting from TS-treatment. OPO activity in non-TS-treated saliva in the absence [1] or presence [2] of 1 mM OH-CO
  • Each value represents data calculated as average ⁇ SD of results from 3-4 experiments using saliva from 3-4 subjects.
  • FIG. 15 is a histogram depicting OH-CO mediated prevention of OPO activity loss in KCN-treated saliva. Each value represents data calculated as average + SD of results from 3 experiments using saliva from 3 subjects.
  • FIG. 16 is a data plot depicting death of lymphocytes incubated at 37 °C in the presence of TS and saliva.
  • FIG. 17 is a photograph of Western immunoblotting analysis depicting increased levels of protein carbonylation in lymphocytes treated with TS in the presence of saliva.
  • Lane 1 incubation in PBS alone
  • Lane 2 incubation in the presence of saliva
  • Lane 3 incubation in PBS + TS
  • Lane 4 incubation with TS + saliva. Incubations were performed for 80 min at 37 °C.
  • FIG. 18 is a photograph of Western immunoblotting analysis depicting the effects of saliva and uric acid on lymphocyte protein carbonylation. Lane 1: incubation in PBS alone, Lane 2: incubation in the presence of saliva, Lane 3: incubation in the presence of 10 ⁇ M uric acid, Lane 4: incubation in the presence of
  • FIG. 19 is a data plot depicting the effects of various antioxidants; 1 mM GSH, 1 mM NAC (N-acetylcysteine) and ImM deferoxamine (DES), on survival of lymphocytes incubated in the presence of TS and saliva at 37 0 C. Asc: ascorbate.
  • FIG. 20 is a photograph of Western immunoblotting analysis depicting the effects of several antioxidants on protein carbonylation levels in lymphocyte treated with TS and saliva.
  • Lane 1 incubation in the presence of TS and saliva
  • Lane 2 incubation in the presence of TS, saliva and 1 mM NAC
  • Lane 3 incubation in the presence of TS, saliva and 1 mM ascorbate
  • Lane 4 incubation in the presence of TS, saliva and 1 mM GSH
  • Lane 5 incubation in the presence of TS, saliva and 1 mM deferoxamine (DES). Incubations were performed at 37 0 C for 20 min.
  • DES deferoxamine
  • FIG. 21 is a photograph of Western immunoblotting analysis depicting the effects of several volatile aldehydes on lymphocyte carbonylation levels. Lane 1: incubation in the presence of PBS alone, Lane 2: incubation with 80 ⁇ M acrolein,
  • Lane 3 incubation with 20 ⁇ M crotonaldehyde
  • Lane 4 incubation with 2 ⁇ M acetaldehyde
  • Lane 5 incubation with 80 ⁇ M acrolein + 1 mM GSH
  • Lane 6 incubation with 20 ⁇ M crotonaldehyde + 1 mM GSH
  • Lane 7 incubation with 2 ⁇ M crotonaldehyde + 1 mM GSH. Incubations were performed at 37 °C for 20 min.
  • FIG. 22 is a photograph of Western immunoblotting analysis depicting the effects of saliva and acrolein on the lymphocyte protein carbonylation levels.
  • FIG. 23 is a histogram depicting survival of lymphocytes incubated at 37 °C for 20 min in the presence of saliva, various antioxidants and redox-active iron. Column 1: incubation in PBS alone, Column 2: incubation in the presence of saliva,
  • FIG. 24 is a histogram depicting survival of lymphocytes incubated at 37 °C for 80 min in the presence of TS, ascorbate and deferoxamine (DES).
  • Column 1 incubation in PBS alone
  • Column 2 incubation in the presence of TS
  • Column 3 incubation in the presence of 1 mM ascorbate
  • Column 4 incubation in the presence of 1 mM deferoxamine
  • Column 5 incubation in the presence of TS + 1 mM ascorbate
  • Column 6 incubation in the presence of TS + 1 mM ascorbate + 1 mM deferoxamine.
  • FIG. 25 is a photograph of Western immunoblotting analysis depicting the effects of GSH and deferoxamine (DES) on protein carbonylation levels in lymphocytes incubated for 20 or 80 minutes at 37 0 C in the presence of TS and saliva.
  • Lane 1 incubation in the presence of TS + saliva for 20 minutes
  • Lane 2 incubation in the presence of TS + saliva for 80 minutes
  • Lane 3 incubation in the presence of TS + saliva + 1 mM GSH for 20 minutes
  • Lane 4 incubation in the presence of TS + saliva + 1 mM GSH for 80 minutes
  • Lane 5 incubation in the presence of TS + saliva +
  • Lane 6 incubation in the presence of TS + saliva + 1 mM GSH + 1 mM deferoxamine for 80 minutes
  • Lane 7 incubation in the presence of TS + saliva + 1 mM GSH + 5 mM deferoxamine for 20 minutes
  • Lane 8 incubation in the presence of TS + saliva + 1 mM GSH + 5 mM deferoxamine for 80 minutes.
  • FIG. 26 is a diagram depicting mechanisms of cyanate metabolism following exposure to TS.
  • the present invention is of methods, pharmaceutical compositions, oral compositions, filters and tobacco products for preventing or reducing tobacco smoke- associated injury in the aerodigestive tract of a subject.
  • the present invention can be used to prevent or reduce loss of OPO activity or CN -, redox- active metal ion- or aldehyde-induced cell death resulting from TS-associated oxidative stress, all of which being capacities not provided by prior art methods.
  • the present inventors have previously described novel smoking filters and oral compositions for reducing tobacco associated damage in the aeurodigestive tract (see
  • compositions include active agents which are capable of reducing or preventing tobacco associated loss of peroxidase activity in the aerodigestive tract.
  • compositions and means for preventing or reducing any tobacco associated oxidant injury in the aerodigestive tract devoid of the above limitation. While reducing the present invention to practice, the present inventors uncovered that previously described agents (see U.S. Pat. No. 6,789,546) can be implemented in any tobacco and tobacco related products which come in direct contact with the aerodigestive tract (e.g., saliva) and as such can be ameliorate tobacco damage simultaneously with its use (e.g., chewing).
  • an article of manufacturing comprising tobacco and a tobacco packaging material, at least a portion of the tobacco and/or tobacco packaging material being in contact with an aerodigestive tract of a subject in use thereof, and whereas the at least a portion of the tobacco and/or tobacco packaging material comprises an agent capable of reducing or preventing tobacco associated loss of peroxidase activity in the aerodigestive tract.
  • an article of manufacturing comprising a tobacco packaging material at least a portion of which being in contact with an aerodigestive tract of a subject in use thereof, wherein the at least a portion comprises an agent capable of reducing or preventing tobacco associated loss of peroxidase activity in the aerodigestive tract.
  • tobacco refers to any tobacco species (e.g., crude or extract) which is compatible with human use.
  • the present invention also envisages the use of the agents of the present invention (in line with the above described aspects) with other smoked, dipped, chewed, snuff or snused herbs, compatible with human consumption and which causes damage to the aerodigestive tract due to the loss of peroxidase activity.
  • tobacco packaging material refers to any auxiliary means which packages the tobacco or facilitates its consumption (carrier). Examples include, but are not limited to, rolling paper, snus bags, filter paper, tin sheets and the like.
  • the agent may be impregnated in (attached to, absorbed in, coated with) a filter paper which comes in direct contact with the aerodigestive tract.
  • FIG 1 illustrates a cigarrete filter configuration of the tobacco smoke (TS) filter of the present invention which is referred to hereinunder as a cigarette filter 10.
  • Cigarette filter 10 is constructed of a paper lining 12 and a filter core 14 which is composed of glass fiber and is positioned adjacent to a tobacco filling 16.
  • the agent of the present invention can be impregnated in filter lining 12.
  • Such filters have been previously described in patent documents (39, 40), the teachings of which are herein incorporated by reference.
  • the rolling paper may be treated with the agent such that the agent is confined to that region of the paper which comes in contact with the aerodigestive tract (say about 1 cm margines).
  • saliva-lined tissues such as the lips, mouth, buccal cavity, tongue, oropharynx, throat, larynx, esophagus, upper digestive tract, saliva glands, saliva, as well as the similar mucous-lined tissues of the respiratory tract, such as the respiratory mucosa, alveoli, trachea, and lungs.
  • antioxidants such as CN ⁇ chelators can be used to treat TS-associated loss of OPO activity.
  • examples of CN- chelators, such as OH-CO and additional antioxidants which can be used by this aspect of the present invention are given hereinbelow.
  • the CN " chelator e.g., OH-CO
  • the CN " chelator is administered in a manner which enables establishment of a concentration of 0.5-2 mM, preferably 1 mM in body fluids, such as saliva.
  • CN ⁇ chelators can be effectively employed to prevent or reduce TS- associated injury in the aerodigestive tract since they act to sequester CN ⁇ which is injurious to OPO.
  • Such capacity of OH-CO, also known as the non-CN -bound form of cyanocobalamin, hydroxocobalamin or vitamin B 12a, to prevent TS-induced loss of OPO activity represents a marked improvement over prior art methods of preventing TS-mediated oxidant injury of the aerodigestive tract since such protection has never been demonstrated or suggested by prior art methods employing other antioxidants such as GSH, ascorbate or deferoxamine, as shown in Example 1 of the Examples section, below.
  • antioxidants functional as redox-active metal ion chelators can be used to treat TS- associated death of cells, such as lymphocytes.
  • Redox-active metal ion chelators such as deferoxamine
  • physiological antioxidants such as GSH
  • GSH physiological antioxidants
  • Redox-active metal ion chelators are used in a manner which enables establishment of about ImM concentration in body fluids (e.g.,saliva).
  • deferoxamine is administered in a manner which enables establishment of a concentration of about 1 mM, more preferably about 5 mM in body fluids. More preferably, a mixture of deferoxamine and GSH is used in a ratio of about 1:1, preferably 5:1, respectively.
  • deferoxamine and GSH body fluid concentrations of about 1 mM each are desirable although a deferoxamine concentration of 5 mM and a GSH concentration of 1 mM are also therapeutically effective.
  • antioxidants such as GSH and redox-active metal chelators, such as deferoxamine, preferably in combination, represents a significant improvement over prior art methods of preventing or reducing TS-mediated oxidant injury in the aerodigestive tract.
  • CN chelators suitable for use according to the present invention include, for example, epselen, vitamins A, C and E, selenium compounds, OH-CO, flavenoids, qui ⁇ ones (e.g., QlO, Q9), retinoids and carotenoids.
  • redox-active metal ion chelators suitable for use according to the present invention include, for example, epselen, desferoxamine, zinc- desferioxamine, polyamine chelating agents, ethylenediamine, diethylenetriamine, triethylenetetramine, triethylenediamine, tetraethylenepentamine, aminoethylethanolamine, aminoethylpiperazine, pentaethylenehexamine, triethylenetetramine-hydrochloride, tetraethylenepentamine-hydrochloride, pentaethylenehexamine-hydrochloride, tetraethylpentamine, captopril, penicilamine, N,N'-bis(3-aminopropyl)-l,3-propanediamine, N,N,Bis (2 aminoethyl) 1,3 propane diamine, l,7-dioxa-4,10-diazacyclododecane, 1,4,8, 11 -tetraaza
  • Articles of the present invention can further comprise at least one fiavorant such as wintergreen oil, oregano oil, bay leaf oil, peppermint oil, spearmint oil, clove oil, sage oil, sassafras oil, lemon oil, orange oil, anise oil, benzaldehyde, bitter almond oil, camphor, cedar leaf oil, marjoram oil, citronella oil, lavendar oil, mustard oil, pine oil, pine needle oil, rosemary oil, thyme oil, and cinnamon leaf oil.
  • fiavorant such as wintergreen oil, oregano oil, bay leaf oil, peppermint oil, spearmint oil, clove oil, sage oil, sassafras oil, lemon oil, orange oil, anise oil, benzaldehyde, bitter almond oil, camphor, cedar leaf oil, marjoram oil, citronella oil, lavendar oil, mustard oil, pine oil, pine needle oil, rosemary oil, thyme oil, and cinnamon leaf
  • the active ingredients may be introduced to the article of manufacturing as described above (e.g., snuff), such as in the forma dry powder, either as a mixture of antioxidants, or as a complex in protective liposomes, nanospheres or other acceptable delivery vehicles.
  • This powder may be added in the final process of manufacturing and may also contain suitable flavors or fragrances as not infrequently used in this industry.
  • Hydroxocobalamin provides protection from tobacco smoke-induced loss of aerodigestive tract antioxidant defenses
  • Many common and highly debilitating oral diseases such as cancer, periodontitis and gingivitis, result from, or are aggravated by consumption of tobacco products, such as tobacco smoking.
  • oral cancer a frequently lethal and highly debilitating disease, results from tobacco consumption in 50-90 % of cases world-wide.
  • One widely accepted mechanism whereby cancer progression is promoted is via oxidant injury, such as protein damage caused by exposure to free radicals.
  • the present inventors have therefore analyzed the effects of TS on aerodigestive tract antioxidant defenses and have invented means to prevent such effects, as described below. Exposure to tobacco smoke leads to a decrease in oral peroxidase and to an increase in protein damage:
  • Oral peroxidase is the critical salivary enzymatic defense against upper digestive tract oxidant injury resulting in, for example, macromolecular damage associated with progression of diseases such as malignancies of the upper digestive tract, periodontitis and gingivitis.
  • the mechanism of OPO mediated protection from antioxidant injury resulting from exposure to TS HCN is schematized in Figure 2a.
  • Saliva collection Whole saliva was collected under non-stimulatory conditions from healthy smokers having smoked at least 20 cigarettes a day for at least 10 years and from non-smoking subjects, as previously described (43).
  • Tobacco smoke was obtained from popular commercial cigarettes containing 14 mg of tar and 0.9 mg of nicotine ('Time' cigarettes, Dubek Ltd., Tel Aviv, Israel).
  • OPO activity was measured according to the 2- nitrobenzoic-thiocyanate (NBS-SCN) assay, as previously described (20). Briefly, DTNB is reduced to NBS by addition of ⁇ -mercaptoethanol and decreases in NBS concentration by reaction with OSCKT, a product of OPO, are monitored spectrophotometrically by measuring absorbance at 412 ran at pH 5.6 (18). One unit of enzyme activity was defined as the activity required to cleave 1 ⁇ mol of NBS/min at 22 °C, using a molar extinction coefficient of 12,800 (20).
  • Hydrocyanic acid mediates tobacco smoke-associated loss of oral peroxidase activity:
  • Tobacco smoke was obtained from popular commercial cigarettes containing 14 mg of tar and 0.9 mg of nicotine ('Time' cigarettes, Dubek Ltd., Tel Aviv, Israel).
  • In vitro exposure of saliva to TS The in vitro exposure of saliva samples to
  • TS was performed as described above with the modification that the same saliva samples were subjected to multiple exposures to the TS of 1 cigarette at 20 min intervals.
  • Analysis of OPO activity Analysis of OPO activity in TS-treated saliva samples was performed as described above.
  • Loss of OPO activity due to exposure to TS was found to be 60-85% in the presence of 150 ⁇ M of dapsone whereas control MPO activity was unaffected, ( Figure 7) and loss of OPO activity was similar in the presence or absence of 150 ⁇ M dapsone in saliva containing ⁇ 40 % MPO ( Figure 8) indicating that TS affects SPO and MPO activities to a similar extent.
  • Cyanide is a known inhibitor of heme peroxidase, and the gas-phase TS of various cigarette brands have been found to contain 2-233 ⁇ g of KCN (45).
  • saliva samples were treated with 150 ⁇ M KCN and analyzed for OPO activity. This concentration of KCN was observed to cause a loss of ⁇ 65 % of OPO activity after only 2 min of incubation, which loss was considerably reversed by dialysis ( Figure 13), thereby indicating that KCN is indeed capable of inhibiting OPO activity.
  • Hydroxocobalamin prevents tobacco smoke-associated loss of salivary peroxidase: Since CN ion was found to be involved in TS-associated loss of the OPO activity, the present inventors have attempted to provide a means of preventing such loss of OPO activity via the use of a CN ⁇ chelator.
  • OH- CO can be employed to effectively reduce or prevent the occurrence of diseases, such as malignancies of the aerodigestive tract, periodontitis and gingivitis associated with exposure to TS.
  • the antioxidants deferoxamine and glutathione prevent upper aerodigestive tract lymphocyte death associated with exposure to tobacco smoke
  • Saliva collection Saliva was collected from healthy subjects, 3 males and 3 females ranging from 21-47 years of age, under non-stimulatory conditions in a quiet room during the morning between 8 am and noon. Saliva collection was performed at least 1 h after eating by spitting into a recipient for 10 minutes, as previously described (43). Following collection, saliva was immediately centrifuged at 800 x g for 10 min at 4 °C to remove cells and cell debris and the resulting supernatant was used for biochemical analyses. Lymphocyte isolation: Blood from 10 consenting, healthy, non-smoking subjects, 5 males and 5 females ranging from 18-55 years of age, was drawn into EDTA-containing vacutainers.
  • Lymphocytes were isolated by Ficoll-Hypaque (Sigma) gradient centrifugation according to the manufacturer's instructions and lymphocytes were suspended at 10 7 cells/ml PBS (Beit-Ha'emek Industries, Israel), and used immediately in experiments.
  • Tobacco smoke was obtained from popular commercial cigarettes containing 14 mg of tar and 0.9 mg of nicotine ('Time' cigarettes, Dubek Ltd., Tel Aviv, Israel).
  • Exposure of lymphocytes to TS was carried out by attaching a Time cigarette (capable of removing particles > 0.1 mm in diameter) in which the filter tip was removed to a Cambridge filter which was combined with a vacuum system to aspirate gas-phase TS inside sealed 250 ml flasks containing lymphocytes suspended in 12-15 ml PBS, as previously described (14, 15). A reproducible vacuum was created in the flask and upon application of vacuum for 5 s, 80-100 ml of TS from the lit cigarette was drawn into the flask.
  • lymphocyte viability was assessed by Trypan Blue exclusion assay.
  • Lymphocytes were washed twice in PBS following treatment by centrifugation at 2,000 rpm for 2 minutes to remove saliva and other components of the incubation medium. Lymphocytes were then centrifuged at 14,000 rpm for 1 minute and lysed by sonication for 10 seconds in lysis buffer containing 20 mM Tris pH 7.4, 1 mM EGTA, 1 mM PMSF, 50 ⁇ M NaVO 4 and 50 mM NaF. Lysates were centrifuged at 14,000 rpm for 1 minute and supernatants were harvested for analysis. Carbonylation analysis was performed as described above.
  • Results for statistical analysis were obtained from the control subgroup (lymphocytes in PBS) and from the various treatment subgroups. Means, SDs and SEMs were computed and results between the subgroups were analyzed and compared via one-way analysis-of-variance (46) using the Bonferroni Multiple-Comparison Test Model (47) to determine significant differences between computed means. The means between each pair of means was analyzed via T-test
  • GSH and deferoxamine according to the present invention, can therefore be effectively employed to prevent or reduce diseases of the aerodigestive tract associated with cell death caused by tobacco consumption.
  • Epstein JB Scully C. Assessing the patient at risk for oral squamous cell carcinoma. SCD Special Care in Dentistry 1997;17:120-8.

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Abstract

L'invention concerne des articles de fabrication pour empêcher ou réduire une lésion associée à la fumée du tabac et associée au tabac sans fumée dans les voies respiratoires et le tube digestif d'un sujet.
EP08702682A 2007-01-22 2008-01-22 Tabac et matériau de conditionnement de tabac pour empêcher ou réduire une lésion associée au tabac dans les voies respiratoires et le tube digestif d'un sujet Withdrawn EP2129244A2 (fr)

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PCT/IL2008/000101 WO2008090552A2 (fr) 2007-01-22 2008-01-22 Tabac et matériau de conditionnement de tabac pour empêcher ou réduire une lésion associée au tabac dans les voies respiratoires et le tube digestif d'un sujet

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IT1396903B1 (it) * 2009-03-23 2012-12-20 River Pharma Srl Composizioni comprendenti antiossidanti e chelanti del ferro e loro uso per il trattamento del sovraccarico di ferro
US11766068B1 (en) * 2009-12-14 2023-09-26 Altria Client Services Llc Method of treating smokeless tobacco
EP2552245B1 (fr) 2010-03-26 2018-11-14 Philip Morris Products S.A. Inhibition de l'irritation sensorielle pendant la consommation de produits de tabac non fumables
CN102273732B (zh) * 2011-06-03 2013-04-10 湖南中烟工业有限责任公司 一种弱极性卷烟纸质滤材及其制备和应用方法
CN103815540A (zh) * 2014-03-14 2014-05-28 红塔烟草(集团)有限责任公司 一种槟榔风味的无烟烟草制品及其制备方法
CN104001478B (zh) * 2014-06-19 2016-02-03 云南中烟工业有限责任公司 一种醋酸铁交联蒙脱石滤嘴添加剂及其应用
CN106750811A (zh) * 2016-12-27 2017-05-31 广西中烟工业有限责任公司 一种抗菌、抗氧化、金属螯合性的多功能塑料及其制备方法
CN113749282A (zh) * 2020-06-01 2021-12-07 张家港外星人新材料科技有限公司 一种表面喷洒掺杂含硒有机酸的烟丝及含有该烟丝的制品
EP4408184A1 (fr) * 2021-09-30 2024-08-07 Nicoventures Trading Limited Composition de gomme orale

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5906811A (en) * 1997-06-27 1999-05-25 Thione International, Inc. Intra-oral antioxidant preparations
US6138683A (en) * 1997-09-19 2000-10-31 Thione International, Inc. Smokeless tobacco products containing antioxidants
US5829449A (en) * 1997-09-19 1998-11-03 Thione International, Inc. Smoking products containing antioxidants
US5922346A (en) * 1997-12-01 1999-07-13 Thione International, Inc. Antioxidant preparation
US6789546B2 (en) * 2001-06-26 2004-09-14 Technion Research & Development Foundation Ltd. Filters for preventing or reducing tobacco smoke-associated injury in the aerodigestive tract of a subject

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* Cited by examiner, † Cited by third party
Title
See references of WO2008090552A3 *

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US20100108086A1 (en) 2010-05-06

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