Tobacco Carcinogenesis

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Tobacco carcinogenesis is the process by which tobacco products and their constituents interact with cells to cause cancer. Cigarettes are the main tobacco product worldwide. Manufactured cigarettes are available in all countries, but in some areas of the world, roll-your-own cigarettes are still popular. Other smoked products include kreteks, clove-flavored cigarettes popular in Indonesia, and "sticks" which are smoked in Papua, New Guinea. Bidis, a small amount of tobacco wrapped in temburni leaf and tied with a string, are very popular in India and neighboring areas and have recently taken hold in the United States.       
Cigars are presently increasing in popularity, and pipes are still used. A substantial amount of tobacco is consumed worldwide in the form of smokeless tobacco products. These include chewing tobacco, dry snuff used for nasal inhalation, moist snuff, which is placed between the cheek and gum, a popular practice in Scandinavia and North America, and pan or betel quid, a product used extensively in India. All products are complex mixtures of defined compounds, many of which are capable of inducing tumors. The compounds that induce tumors are called carcinogens. The study of cancer induction by tobacco products and their constituents results in the elucidation of mechanisms by which many of the common human cancers develop.


Due to the addictive power of nicotine, worldwide tobacco use is staggering. According to estimates by the World Health Organization, there are about 1100 million smokers in the world, representing approximately one-third of the global population aged 15 years or higher. China alone has approximately 300 million smokers, about the same number as in all developed countries combined. Globally, about 47% of men and 12% of women smoke. Smoking prevalence varies widely by country. For example, in Korea, 68% of men smoke daily, while the corresponding figure for Sweden is 22%. Among women, the highest smoking prevalence is in Denmark, where 37% of women smoke, while in many Asian and developing countries, prevalence is reported to be less than 10%. Although smoking prevalence is lower in the less developed countries in general, it is expected that this will increase markedly as smoking takes hold and larger numbers of young smokers grow older.      
Tobacco in all forms was consumed to the extent of 6.5 billion kg annually in the period 1990-1992, and there were six trillion cigarettes sold. It does not appear that tobacco use will disappear in the near future. Worldwide, smoking is estimated to have caused about 1.05 million cancer deaths in 1990. About 30% of all cancer death in developed countries is caused by smoking. The corresponding figure for developing countries is 13%. Lung cancer is the dominant malignancy caused by smoking, with 514,000 lung cancer deaths attributed to smoking in developed countries in 1995.      
Smoking is also an important cause of bladder cancer, cancer of the renal pelvis, oral cancer, oropharyngeal cancer, hypopharyngeal cancer, laryngeal cancer, esophageal cancer, and pancreatic cancer. Other cancers that may be caused by smoking include renal adenocarcinoma, cancer of the cervix, myeloid leukemia, colon cancer, and stomach cancer. In the United States, about 30% of all cancer death is caused by smoking, similar to worldwide estimates for developed countries. Lung cancer was rare at the beginning of the 20th century. However, the incidence and death rates increased as smoking became more popular. In the United States, the lung cancer death rate in 1930 for men was 4.9 per 100,000. By 1990, this had increased to 75.6 per 100,000. The lung cancer death rate can be shown to parallel the curves for cigarette smoking prevalence, with an approximate 20-year lag time. In 1964, the first Surgeon General’s report on the health consequences of cigarette smoking was published. Following this landmark report, smoking prevalence began to decrease in the United States, but there has been no change since 1990. There are still 47 million adult smokers in the United States.     
Unburned tobacco is a cause of oral cavity cancer. The annual mortality from tobacco chewing in south Asia, where it is used primarily in the form of betel quid, is estimated to be of the order of 50,000 deaths per year. Oral cavity cancer is the leading cancer killer in India. Snuff dipping, as practiced in North America, is an accepted cause of oral cavity cancer as well. The prevalence of snuff dipping has increased markedly in recent years in the United States, especially among young males.


One goal of scientists studying tobacco carcinogenesis  has been to replicate in laboratory animals the effects  of tobacco products that are observed in humans.  This has been challenging for a variety of  reasons, which can be summarized simply: laboratory  animals will not voluntarily use tobacco products the  way humans do.            
The International Agency for Research on Cancer  has reviewed and summarized this work. According  to their conclusions, experimental studies evaluating  the ability of cigarette smoke and its condensate to  cause cancer in laboratory animals have collectively  demonstrated that there is sufficient evidence that  inhalation of tobacco smoke, as well as topical application  of tobacco smoke condensate, causes cancer  in experimental animals. The Syrian golden hamster  has been the model of choice for inhalation  studies of cigarette smoke because it has a low background  incidence of spontaneous pulmonary tumors  and little interfering respiratory infection. Inhalation  of cigarette smoke has repeatedly caused carcinomas  in the larynx of hamsters and this model system has  been widely applied. It is the most reliable model  for induction of tumors by inhalation of cigarette  smoke. Studies in mice, rats, and dogs have been less  frequent.            
According to the International Agency for Research  on Cancer, there are a number of operational  problems inherent in inhalation studies of cigarette  smoke. The smoke must be delivered in a standardized  fashion and this has been accomplished in different  ways. Both whole body exposure and noseonly  exposure designs have been used. Generally, a  2-s puff from a burning cigarette is diluted with air  and forced into the chamber. Animals will undergo  avoidance reactions and will not inhale the smoke.  Thus, the dose to the lung is less than in humans,  which partially explains the occurrence of larynx tumors  rather than lung tumors in hamsters. Unlike  humans, rodents are obligatory nose breathers. Their  nasal passages are more complex than those of humans,  thereby affecting particle deposition in the respiratory  tract. Tobacco smoke is irritating and toxic,  creating further problems in inhalation studies with  rodents.            
Inhalation studies have reproducibly demonstrated  that cigarette smoke, especially its particulate phase,  causes laryngeal carcinomas in hamsters. Some experiments  with mice resulted in low incidences of  lung tumors, in tests of both mainstream smoke and  environmental tobacco smoke. Evidence shows that  gas phase components of cigarette smoke may be tumorigenic  in mice. Respiratory tract tumors were  produced in one long-term exposure of rats to cigarette  smoke. Studies in rabbits and dogs were equivocal.  Treatment-related tumors other than those of  the respiratory tract have not been consistently  observed.            
Cigarette smoke condensate (CSC) has been tested  extensively for tumor induction. CSC is produced by  passing smoke through cold traps. The material in the  traps is recovered by washing with a volatile solvent,  which is then evaporated. Some volatile and semivolatile constituents may be lost during this process.  CSC is roughly equivalent to cigarette total particulate  matter (TPM), the material collected on a glass  fiber filter that has had smoke drawn through it. The  term "tar," which is often used in official reports on  cigarette brands, is equivalent to TPM but without  nicotine and water.              
CSC generation and collection techniques have  been standardized. CSC has been widely tested for  carcinogenicity in mouse skin. Consistently, CSC induces  benign and malignant skin tumors in mice. This  bioassay has been employed to evaluate the carcinogenic  activities of cigarettes of different designs and  to investigate subfractions of CSC. Mouse skin studies  led to the identification of carcinogenic polycyclic  aromatic hydrocarbons in cigarette smoke. The overall  carcinogenic effect of CSC on mouse skin appears  to depend on the composite interaction of tumor initiators  and enhancing factors, such as tumor promoters  and cocarcinogens.            
Many studies have evaluated tumor induction in  rodents by extracts of unburned tobacco. Although  some positive results have been obtained, there is  presently no widely accepted and reproducible model  for the induction of oral cavity cancer in rodents by  tobacco extracts, despite strong human data. There  are probably cofactors that contribute to human oral  cancer upon tobacco use, which are not reproduced  in animal studies.


A second important goal of scientists in the field of tobacco carcinogenesis has been the identification of carcinogens in tobacco products. When these compounds are identified, studies can be designed to investigate the mechanisms by which they cause cancer, which in turn can provide important insights on the ways in which tobacco products cause cancer in humans.      
When cigarette tobacco is burned, mainstream smoke and sidestream smoke are produced. Mainstream smoke is the material drawn from the mouth end of a cigarette during puffing. Sidestream smoke is the material released into the air from the burning tip of the cigarette plus that which diffuses through the paper. The mainstream smoke emerging from the cigarette is an aerosol containing about 1 × 1010 particles/ ml, ranging in diameter from 0.1 to 1.0 μm (mean diameter 0.2 μm). About 95% of the smoke is made up of gases-- predominantly nitrogen, oxygen, and carbon dioxide. For chemical analysis, the smoke is arbitrarily separated into a vapor phase and a particulate phase, based on passage through a glass-fiber filter pad called a Cambridge filter.
In addition to nitrogen, oxygen, and carbon dioxide, the gas phase contains substantial amounts of carbon monoxide, water, argon, hydrogen, ammonia, nitrogen oxides, hydrogen cyanide, hydrogen sulfide, methane, isoprene, butadiene, formaldehyde, acrolein, pyridine, and other compounds. The particulate phase contains more than 3500 compounds, and most of the carcinogens. Some major constituents of the particulate phase include nicotine and related alkaloids, hydrocarbons, phenol, catechol, solanesol, neophytadienes, fatty acids, and others. Many of the components are present in higher concentration in sidestream smoke than in mainstream smoke; this is especially true of nitrogen-containing compounds. However, a person’s exposure to sidestream smoke is generally far less than to mainstream smoke because of dilution with room air.

FIGURE 1 Structures of organic pulmonary carcinogens in tobacco smoke.

TABLE I Summary of Carcinogens in Cigarette Smoke

Type Number of compounds
Polycyclic aromatic hydrocarbons 10
Aza-arenes 3
N-Nitrosamines 10
Aromatic amines 4
Heterocyclic aromatic amines 8
Aldehydes 2
Miscellaneous organic compounds 18
Inorganic compounds 7
Total 62

There are over sixty carcinogens in cigarette smoke that have been evaluated by the International Agency for Research on Cancer and for which there is "sufficient evidence for carcinogenicity" in either laboratory animals or humans. The types of carcinogens, based on their chemical classes, are listed in Table I. Carcinogens specifically associated with lung cancer are listed in Table II. The 20 compounds included in this list have been found convincingly to induce lung tumors in at least one animal species and have been positively identified in cigarette smoke. Structures of the organic compounds are shown in Fig. 1. These compounds are most likely involved in lung cancer induction in people who smoke.

TABLE II Pulmonary Carcinogens in Cigarette Smoke

Carcinogen class Compound Amount in mainstream cigarette smoke (ng/cigarette) Sidestream/mainstream smoke ratio Representative lung tumorigenicity in species
Polycyclic aromatic hydrocarbons Benzo[a]pyrene 20-40 2.5-3.5 Mouse, rat, hamster
  Benzo[b]fluoranthane 4-22   Rat
  Benzo[j]fluoranthane 6-21   Rat
  Benzo[k]fluoranthane 6-12   Rat
  Dibenzo[a,i]pyrene 1.7-3.2   Hamster
  Indeno[1,2,3-cd]pyrene 4-20   Rat
  Dibenz[a,h]anthracene 4   Mouse
  5-Methylchrysene 0.6   Mouse
Aza-arenes Dibenz[a,h]acridine 0.1   Rat
  7H-Dibenzo[c,g ]carbazole 0.7   Hamster
N-Nitrosamines N-Nitrosodiethylamine ND-2.8 40 Hamster
  4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone 80-770 1-4 Mouse, rat, hamster
Miscellaneous organic compounds 1,3-Butadiene 20-70 × 103   Mouse
  Ethyl carbamate 20-38   Mouse
Inorganic compounds Nickel 0-510 13-30 Rat
  Chromium 0.2-500   Rat
  Cadmium 0-6670 7.2 Rat
  Polonium-210 0.03-1.0 pCi 1.0-4.0 Hamster
  Arsenic 0-1400   None
  Hydrazine 24-43   Mouse

Polycyclic aromatic hydrocarbons are condensed ring aromatic compounds that are formed during all incomplete combustion reactions, such as those that occur in the burning cigarette. Among these, benzo[a]pyrene (BaP) is the most extensively studied compound. Its presence in cigarette smoke and ability to induce lung tumors upon local administration or inhalation are firmly established. It causes lung tumors in mice, but not in rats, when administered systemically. In studies of lung tumor induction by implantation in rats, BaP is more carcinogenic than several other polycyclic aromatic hydrocarbons in tobacco smoke.      
Aza-arenes are nitrogen-containing analogs of polycyclic aromatic hydrocarbons. Two aza-arenes, dibenz[a,h]acridine and 7H-dibenzo[c,g]carbazole, are pulmonary tumorigens when tested by implantation in the rat lung and instillation in the hamster trachea, respectively. The carcinogenic activity of dibenz[a,h]acridine is less than that of BaP, whereas that of 7H-dibenzo[c,g]carbazole is greater than BaP. The levels of both compounds in cigarette smoke are relatively low.      
N-Nitrosamines comprise a large group of potent carcinogens. Among these, N-nitrosodiethylamine is an effective pulmonary carcinogen in the hamster, but not the rat. Its levels in cigarette smoke are low compared to those of other carcinogens. 4-(Methylnitrosamino)- 1-(3-pyridyl)-1-butanone (NNK) is a potent lung carcinogen in rats, mice, and hamsters. NNK is called a tobacco-specific N-nitrosamine because it is a chemical derivative of nicotine, and thus occurs only in tobacco products. It is the only compound in Table II that induces lung tumors systemically in all three commonly used rodent models. The specificity of NNK for tumor induction in the lung is remarkable; it induces lung tumors independent of the route of administration and in both susceptible and resistant strains of mice. The systemic administration of NNK to rats is a reproducible and robust method for the induction of lung tumors. Cigarette smoke contains substantial amounts of NNK, and the total dose experienced by a smoker in a lifetime of smoking is remarkably close to the lowest total dose shown to induce lung tumors in rats. Levels of NNK and total polycyclic aromatic hydrocarbons in cigarette smoke are similar.      
Lung is one of the multiple sites of tumorigenesis by 1,3-butadiene in mice, but is not a target in the rat. Ethyl carbamate is a well-established pulmonary carcinogen in mice but not in other species. Nickel, chromium, cadmium, and arsenic are all present in tobacco and a percentage of each is transferred to mainstream smoke; arsenic levels are substantially lower since discontinuation of its use as a pesticide in 1952. Metal carcinogenicity depends on the valence state and anion; these are poorly defined in many analytical studies of tobacco smoke. Thus, although some metals are effective pulmonary carcinogens, the role of metals in tobacco-induced lung cancer is unclear. Levels of polonium-210 in tobacco smoke are not believed to be great enough to appreciably impact lung cancer in smokers. Hydrazine is an effective lung carcinogen in mice and has been detected in cigarette smoke in limited studies.      
Considerable data indicate that polycyclic aromatic hydrocarbons and NNK play very important roles as causes of lung cancer in people who smoke. The other compounds discussed earlier may also contribute, but probably to a lesser extent.      
Polycyclic aromatic hydrocarbons and Nnitrosamines such as NNK and N'-nitrosonornicotine (NNN) are probably involved as causes of oral cavity cancer in smokers. N-Nitrosamines such as NNN and NDEA are likely causes of esophageal cancer in smokers. The risk of oral cavity cancer and esophageal cancer in smokers is markedly enhanced by the consumption of alcoholic beverages. NNK is also believed to play a prominent role in the induction of pancreatic cancer in smokers, whereas aromatic amines such as 4-aminobiphenyl and 2-naphthylamine are the most likely causes of bladder cancer.       
Cigarette smoke and CSC are tumor promoters, e.g., they enhance the carcinogenicity of tumor initiators when administered subsequent to the initiators. The majority of the tumor-promoting activity seems to be due to uncharacterized weakly acidic compounds. Substantial levels of cocarcinogens, which enhance the carcinogenicity of tumor initiators when applied together with the initiators, are present in cigarette smoke. Catechol is prominent among these. In addition, cigarette smoke contains high levels of acrolein, which is toxic to the pulmonary cilia, and other agents such as nitrogen oxides, acetaldehyde, and formaldehyde, which could contribute indirectly to pulmonary carcinogenicity through their toxic effects.       
While cigarette smoke is extraordinarily complex, unburned tobacco is simpler. With respect to carcinogens, the tobacco-specific nitrosamines NNK and NNN are the most prevalent strong cancer-causing agents in unburned tobacco products. A mixture of NNK and NNN induces oral tumors in rats, and consequently these compounds are considered to play an important role as causes of oral cavity cancer in people who use smokeless tobacco products.

FIGURE 2 Scheme linking nicotine addiction and lung cancer via tobacco smoke carcinogens and their induction of multiple mutations in critical genes. PAH, polycyclic aromatic hydrocarbons; NNK, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butane.


The mechanisms by which tobacco causes cancer can best be illustrated by considering the relationship between cigarette smoking and lung cancer because it is here that the most information is available. The overall framework for discussing this information is illustrated in Fig. 2. Carcinogens form the link between nicotine addiction and cancer. Nicotine addiction is the reason that people continue to smoke. While nicotine itself is not considered to be carcinogenic, each cigarette contains a mixture of carcinogens. Thus, cigarettes are disastrous nicotine delivery devices. Most of the carcinogens in cigarette smoke require metabolic activation, i.e., they must be enzymatically transformed by the host into reactive intermediates in order to exert their carcinogenic effects. There are competing detoxification pathways, which result in harmless excretion of the carcinogens. The balance between metabolic activation and detoxification differs among individuals and will affect cancer risk.      
A great deal is known about mechanisms of carcinogen metabolic activation and detoxification. The metabolic activation process leads to the formation of DNA adducts, which are carcinogen metabolites bound covalently to DNA, usually at guanine or adenine. There have been major advances in our understanding of DNA adduct structure and its consequences in the past two decades and there is now a large amount of mechanistic information available. If DNA adducts escape cellular repair mechanisms and persist, they can cause miscoding, resulting in a permanent mutation in DNA. This occurs when DNA polymerase enzymes read an adducted DNA base incorrectly, resulting in the insertion of the wrong base.       
Other errors can also occur due to the presence of DNA adducts. Cells that contain damaged DNA may be removed by apoptosis, or programmed cell death. If a permanent mutation occurs in a critical region of an oncogene or tumor suppressor gene, it can lead to activation of the oncogene or deactivation of the tumor suppressor gene. Oncogenes and tumor suppressor genes play critical roles in the normal regulation of cellular growth. Changes in multiple oncogenes or tumor suppressor genes result in the production of aberrant cells with loss of normal growth control. Ultimately, this leads to lung cancer. While the sequence of events has not been well defined, there can be little doubt that these molecular changes are important. There is now a large amount of data on mutations in the human K-ras oncogene and p53 tumor suppressor gene in lung tumors from smokers.      
Blocking any of the horizontal steps in Fig. 2 may lead to decreased lung cancer, even in people who continue to smoke. The following discussion considers some of these steps in more detail.      
Upon inhalation, cigarette smoke carcinogens are enzymatically transformed to a series of metabolites as the exposed organism attempts to convert them to forms that are more readily excreted. The initial steps are usually carried out by cytochrome P450 (P450) enzymes, which add oxygen to the substrate. These enzymes typically are responsible for the metabolism of drugs, other foreign compounds, and some endogenous substrates. Other enzymes, such as lipoxygenases, cyclooxygenases, myeloperoxidase, and monoamine oxidases, may also be involved, but less frequently.       
The oxygenated intermediates formed in these initial reactions may undergo further transformations by glutathione-S-transferases, uridine-5'-diphosphateglucuronosyltransferases, sulfatases, and other enzymes, which are typically involved in detoxification. Some of the metabolites produced by P450s react with DNA or other macromolecules to form adducts. Metabolic pathways of BaP and NNK, representative pulmonary carcinogens in cigarette smoke, have been extensively defined through studies in rodent and human tissues.      
The major metabolic activation pathway of BaP is conversion to a reactive diol epoxide metabolite called BPDE; one of the four isomers produced is highly carcinogenic and reacts with DNA to form adducts with N2- of deoxyguanosine. The major metabolic activation pathways of NNK and its main metabolite, 4- (methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), occur by hydroxylation of the carbons adjacent to the N-nitroso group ( -hydroxylation), which leads to the formation of two types of DNA adducts: methyl adducts, such as 7-methyguanine or O6-methylguanine, and pyridyloxobutyl adducts.  
Considerable information is available on pulmonary carcinogen metabolism in vitro, both in animal and in human tissues, but fewer studies have been carried out on uptake, metabolism, and adduct formation of cigarette smoke lung carcinogens in smokers. Various measures of cigarette smoke uptake in humans have been used, including exhaled carbon monoxide, carboxyhemoglobin, thiocyanate, and urinary mutagenicity.     
BaP has been detected in human lung; no differences between smokers and nonsmokers were noted. 1-Hydroxypyrene and its glucuronide, urinary metabolites of the noncarcinogen pyrene, have been widely used as indicators of polycyclic aromatic hydrocarbon uptake. 1-Hydroxypyrene levels in smokers are generally higher than in nonsmokers. Overall, there is considerable evidence that pulmonary carcinogens in cigarette smoke are taken up and metabolized by smokers as well as by nonsmokers exposed to environmental tobacco smoke.       
Less than 20% of smokers will get lung cancer. Susceptibility will depend in part on the balance between carcinogen metabolic activation and detoxification in smokers. This is an important area requiring further study. Most investigations have focused on the metabolic activation pathways by quantifying DNA or protein adducts. There is considerable data demonstrating the activation of BaP to DNA adducts in the lungs of smokers. Earlier investigations demonstrated that cigarette smoke induces aryl hydrocarbon hydroxylase (AHH) activity and proposed a relationship between AHH activity and lung cancer. AHH metabolizes BaP and is equivalent to P450 1A1. Cigarette smoking induces expression of this enzyme. Lung tissue from recent smokers with elevated AHH activity metabolically activated BaP to a greater extent than lung tissue from nonsmokers or ex-smokers. DNA adduct levels correlated with AHH activity in the same samples. Collectively, these results support the existence of a cigarette smoke-inducible pathway leading to BaP-DNA adducts in smokers’ lungs (Fig. 2).      
Several studies have detected 7-methylguanine in human lung. Levels were higher in smokers than in nonsmokers in two studies, suggesting that NNK may be one source of these adducts. While 7-methylguanine is not generally considered as an adduct that would lead to miscoding in DNA and the introduction of a permanent mutation, other methyl adducts that do have miscoding properties, such as O6-methylguanine, are formed at the same time, but at lower levels. Pyridyloxobutylated DNA also has been detected in lung tissue from smokers, reflecting metabolic activation of NNK or NNN. The detection of methyl and pyridyloxobutyl adducts in DNA from smokers’ lungs is consistent with the ability of human lung tissue to metabolically activate NNK, but the quantitative aspects of the relationship of metabolism to DNA adduct levels are unclear.      
DNA repair processes are important in determining whether DNA adducts persist. Because smoking is a chronic habit, one would expect a steady-state DNA adduct level to be achieved by the opposing effects of damage and repair. Mechanisms of DNA repair include direct repair, base excision repair, and nucleotide excision repair. With respect to smoking and lung cancer, direct repair of O6-methylguanine by O6- methylguanine-DNA alkyltransferase and nucleotide excision repair of polycyclic aromatic hydrocarbon- DNA adducts would appear to be the most relevant processes.      
As indicated in Fig. 2, the direct interaction of metabolically activated carcinogens with critical genes such as the p53 tumor suppressor gene and the K-ras oncogene is central to the hypothesis that specific carcinogens form the link between nicotine addiction and lung cancer. The p53 gene plays a critical role in the delicate balance of cellular proliferation and death. It is mutated in about half of all cancer types, including over 50% of lung cancers, leading to loss of its activity for cellular regulation. Point mutations at guanine (G) are common. In a sample of 550 p53 mutations in lung tumors, 33% were G → T transversions, whereas 26% were G → A transitions. (A purine → pyrimidine or pyrimidine → purine mutation is referred to as a transversion, whereas a purine → purine or pyrimidine → pyrimidine mutation is called a transition.) A positive relationship between lifetime cigarette consumption and the frequency of p53 mutations and of G → T transversions on the nontranscribed DNA strand also has been noted.       
These observations are generally consistent with the fact that most activated carcinogens react predominantly at G and that repair of the resulting adducts would be slower on the nontranscribed strand, thus supporting the hypothesis outlined in Fig. 2. Mutations in codon 12 of the K-ras oncogene are found in 24-50% of human primary adenocarcinomas but are rarely seen in other lung tumor types. When K-ras is mutated, a complex series of cellular growth signals is initiated. Mutations in K-ras are more common in smokers and ex-smokers than in nonsmokers, which suggests that they may be induced by direct reaction of the gene with an activated tobacco smoke carcinogen. The most commonly observed mutation is GGT → TGT, which typically accounts for about 60% of the codon 12 mutations, followed by GGT → GAT (20%) and GGT → GTT (15%).       
The p16INK4a tumor suppressor gene is inactivated in more than 70% of human nonsmall cell lung cancers via homozygous deletion or in association with aberrant hypermethylation of the promoter region. In the rat, 94% of adenocarcinomas induced by NNK were hypermethylated at the p16 gene promoter. This change was frequently detected in hyperplastic lesions and adenomas, which are precursors to the adenocarcinomas induced by NNK. Similar results were found in human squamous cell carcinomas of the lung. The p16 gene was coordinately methylated in 75% of carcinoma in situ lesions adjacent to squamous cell carcinomas that had this change. Methylation of p16 was associated with loss of expression in tumors and precursor lesions, indicating functional inactivation of both alleles. Aberrant methylation of p16 has been suggested as an early marker for lung cancer. The expression of cell cycle proteins is related to the p16 and retinoblastoma tumor suppressor genes; NNK induced mouse lung tumors appear to resemble human nonsmall cell lung cancer in the expression of cell cycle proteins. The estrogen receptor gene is also inactivated through promoter methylation. There was concordance between the incidence of promoter methylation in this gene in lung tumors from smokers and from NNK-treated rodents.      
Loss of heterozygosity and exon deletions within the fragile histidine triad (FHIT) gene are associated with smoking habits in lung cancer patients and have been proposed as a target for tobacco smoke carcinogens. However, point mutations within the coding region of the FHIT gene were not found in primary lung tumors.      
Collectively, evidence favoring the sequence of steps illustrated in Fig. 2 as an overall mechanism of tobacco-induced cancer is extremely strong, although there are important aspects of each step that require further study. These include carcinogen metabolism and DNA binding in human lung, the effects of cigarette smoke on DNA repair and adduct persistence, the relationship between specific carcinogens and mutations in critical genes, and the sequence of genetic changes leading to lung cancer.      
Using a weight of the evidence approach, specific polycyclic aromatic hydrocarbons such as BaP and the tobacco-specific nitrosamine NNK can be identified as probable causes of lung cancer in smokers, but the contribution of other agents, such as those listed in Table II, cannot be excluded. The chronic exposure of smokers to the DNA-damaging intermediates formed from these carcinogens is consistent with our present understanding of cancer induction as a process that requires multiple genetic changes. Thus, it is completely plausible that the continual barrage of DNA damage produced by tobacco smoke carcinogens causes the multiple genetic changes that are associated with lung cancer. While each dose of carcinogen from a cigarette is extremely small, the cumulative damage produced in years of smoking is substantial.


Although substantial progress has been accomplished in reducing the tobacco habit, worldwide use of tobacco products is still immense, due mainly to the addictive power of nicotine, arguably the single compound responsible indirectly for more cancer death than any other chemical. Tobacco products cause about 30% of all cancer death. Since the mid-1950s, studies in tobacco carcinogenesis have identified the major carcinogens in tobacco smoke and have elucidated the overall framework by which these carcinogens cause cancer in people. This series of steps, as illustrated in Fig. 2, is well established, although many details remain unclear. Blocking any of the horizontal steps in Fig. 2, even in people who continue to smoke, would result in decreased cancer mortality. Identification of individuals particularly susceptible to the carcinogenic properties of tobacco products would also be important. Rational approaches are now possible in this regard and, even if only partially successful, could have a major impact on cancer mortality because of the sheer magnitude of the epidemic of cancer death caused by tobacco products.

Our studies in tobacco carcinogenesis are supported by Grants CA-44377, CA-81301, and CA-85702 from the National Cancer Institute. The author is an American Cancer Society Research Professor, supported by Grant RP-00-138.

Stephen S. Hecht
University of Minnesota Cancer Center

See Also

carcinogen Any compound that is capable of inducing tumors in laboratory animals or humans.

DNA adduct A covalent binding product formed between a chemical and DNA.

metabolic activation Process by which a carcinogen is converted to a more reactive form that can bind to DNA.

metabolic detoxification Process by which a carcinogen is converted to a form that is excreted without reacting with DNA.

nitrosamines Compounds having a nitroso group bound to the nitrogen of a secondary amine.

polycyclic aromatic hydrocarbons A group of compounds consisting of more than two condensed benzene rings, generally formed in the incomplete combustion of organic matter.

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