Nornicotine Nitrosation in Saliva and Its Relation to Endogenous Synthesis of N'-Nitrosonornicotine in Humans

Knezevich, Aleksandar; Muzic, John G.; Hatsukami, Dorothy K.; Hecht, Stephen S.; Stepanov, Irina

doi:10.1093/ntr/nts172

Cited by 41 publications

(46 citation statements)

References 23 publications

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“…7 This is believed to occur by endogenous nitrosation of nornicotine, a metabolite of nicotine which is easily converted to NNN by reaction with salivary nitrite, or with nitrite under the favorable conditions of gastric pH. 47 …”

Section: Biomarkers Of Exposurementioning

confidence: 99%

Exposure and Metabolic Activation Biomarkers of Carcinogenic Tobacco-Specific Nitrosamines

2015

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Conspectus Lung cancer is the leading cause of cancer death in the world, and cigarette smoking is its main cause. Oral cavity cancer is another debilitating and often fatal cancer closely linked to tobacco product use. While great strides have been made in decreasing tobacco use in the United States and some other countries, there are still an estimated 1 billion men and 250 million women in the world who are cigarette smokers and there are hundreds of millions of smokeless tobacco users, all at risk for cancer. Worldwide, lung cancer kills about 3 people per minute. This account focuses on metabolites and biomarkers of two powerful tobacco-specific nitrosamine carcinogens – 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N′-nitrosonornicotine (NNN) - considered to be among the main causes of lung cancer and oral cavity cancer in people who use tobacco products. Three properties of NNK and NNN are critical for successful biomarker studies: they are present in all tobacco products, they are tobacco-specific and are not found in any other product, and they are strong carcinogens. NNK and NNN are converted in humans to urinary metabolites that can be quantified by mass spectrometry as biomarkers of exposure to these carcinogens. They are also metabolized to diazonium ions and related electrophiles that react with DNA to form addition products that can be detected and quantified by mass spectrometry. These urinary metabolites and DNA addition products can serve as biomarkers of exposure and metabolic activation, respectively. The biomarkers of exposure, in particular the urinary NNK metabolites NNAL and its glucuronides, have been extensively applied to document tobacco-specific lung carcinogen uptake in smokers and non-smokers exposed to secondhand tobacco smoke. Highly sensitive mass spectrometric methods have been developed for quantitative analysis of these NNK metabolites as well as metabolites of NNN in human urine, blood, and toenails. Urinary and serum NNAL have been related to lung cancer risk, and urinary NNN to esophageal cancer risk, in prospective epidemiology studies. These results are consistent with carcinogenicity studies of NNK, NNAL and NNN in rats, which show that NNK and NNAL induce mainly lung tumors, while NNN causes tumors of the esophagus and oral cavity. Biomarkers of metabolic activation of NNK and NNN applied in human studies include the metabolism of deuterium labelled substrates to distinguish NNK and NNN metabolism from that of nicotine, and the determination of DNA and hemoglobin adducts in tissues, blood, and oral cells from people exposed to tobacco products. As these methods are continually improved in parallel with the ever increasing sensitivity and selectivity of mass spectrometers, development of a comprehensive biomarker panel for identifying tobacco users at high risk for cancer appears to be a realistic goal. Targeting high risk individuals for smoking cessation and cancer surveillance can potentially decrease the risk of developing fatal cancers.

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Section: Biomarkers Of Exposurementioning

confidence: 99%

Exposure and Metabolic Activation Biomarkers of Carcinogenic Tobacco-Specific Nitrosamines

2015

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“…NNN is also present in smoke from cigars and cigarettes, in the saliva of people who chew betel quid with tobacco, and in the saliva of oral-snuff users (19). There is great variation in levels of NNN and NNK in mainstream smoke of cigarettes.…”

Section: Biomarkers Associated With Tobacco Smokingmentioning

confidence: 99%

Urinary Tobacco Smoke–Constituent Biomarkers for Assessing Risk of Lung Cancer

et al. 2014

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Tobacco constituent biomarkers are metabolites of specific compounds present in tobacco or tobacco smoke. Highly reliable analytical methods, based mainly on mass spectrometry, have been developed for quantitation of these biomarkers in both urine and blood specimens. There is substantial inter-individual variation in smoking-related lung cancer risk that is determined in part by individual variability in the uptake and metabolism of tobacco smoke carcinogens. Thus, by incorporating these biomarkers in epidemiological studies we can potentially obtain a more valid and precise measure of in vivo carcinogen dose than by using self-reported smoking history, ultimately improving the estimation of smoking-related lung cancer risk. Indeed, we have demonstrated this by using a prospective study design comparing biomarker levels in urine samples collected from smokers many years prior to their development of cancer, versus those in their smoking counterparts without a cancer diagnosis. The following urinary metabolites were associated with lung cancer risk, independent of smoking intensity and duration: cotinine plus its glucuronide, a biomarker of nicotine uptake; 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and its glucuronides (total NNAL), a biomarker of the tobacco carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK); and r-1-,t-2,3,c-4-tetrahydroxy-1,2,3,4-tetrahydrophenanthrene (PheT), a biomarker of polycyclic aromatic hydrocarbons (PAH). These results provide several possible new directions for using tobacco smoke constituent biomarkers in lung cancer prevention, including improved lung cancer risk assessment, intermediate outcome determination in prevention trials and regulation of tobacco products.

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“…This is particularly important when the precursor amine in question – in this case nornicotine - is readily nitrosated as we have shown in our previous studies [20;21], and when the expected amounts of nitrosamine detected - in this case NNN - may be quite low, as in e-cigarette users. Our initial unpublished analyses of the urine of e-cigarette users showed sporadic, poorly reproducible, and relatively low levels of total NNN, which we attribute to uncontrolled artifact formation resulting from nitrosation of nornicotine via an unknown source of nitrite or nitrogen oxides.…”

Section: Discussionmentioning

confidence: 91%

“…Etter et al reported the presence of nornicotine, up to 0.1% of nicotine content, in some refill liquids [33]. This nornicotine could be easily nitrosated in a user’s saliva, stomach, or urine - a scenario which is quite similar to the endogenous formation of NNN in oral nicotine replacement product users, or in the saliva of people who use tobacco products [12;21]. …”

Section: Discussionmentioning

confidence: 99%

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Combined analysis of N′-nitrosonornicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in the urine of cigarette smokers and e-cigarette users

Kotandeniya

Carmella

Pillsbury

et al. 2015

Journal of Chromatography B

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A liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI+-MS/MS) method for the analysis of the tobacco-specific carcinogens N′-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and their glucuronides (total NNN and total NNAL) in human urine was developed. The method has excellent accuracy and intra-day and inter-day precision, and limits of quantitation of 0.015 and 0.075 pmol/ml urine, respectively, for total NNN and total NNAL. A unique aspect of this method is internal assessment of possible artifactual formation of NNN by inclusion of the monitor amine [pyridine-D4]nornicotine. We found that artifactual formation of NNN comprised only 2.5% of the measured amounts of total NNN in urine of cigarette smokers, under our conditions using ammonium sulfamate as an inhibitor of nitrosation. The method was applied to urine samples from cigarette smokers and e-cigarette users. Levels of total NNN and total NNAL in the urine of cigarette smokers averaged 0.060 ± 0.035 pmol/mL and 2.41 ± 1.41 pmol/mL urine, (N = 38), respectively, which were both significantly greater than in the urine of 27 e-cigarette users.

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Nornicotine Nitrosation in Saliva and Its Relation to Endogenous Synthesis of N'-Nitrosonornicotine in Humans

Abstract: Our results demonstrate that NNN can be formed from nornicotine in human saliva without deliberate addition of any other substance. Therefore, nornicotine, as present in tobacco or in nicotine replacement products, is a carcinogen precursor.

Cited by 41 publications

References 23 publications

Exposure and Metabolic Activation Biomarkers of Carcinogenic Tobacco-Specific Nitrosamines

Exposure and Metabolic Activation Biomarkers of Carcinogenic Tobacco-Specific Nitrosamines

Urinary Tobacco Smoke–Constituent Biomarkers for Assessing Risk of Lung Cancer

Combined analysis of N′-nitrosonornicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in the urine of cigarette smokers and e-cigarette users

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