Beijing Key Laboratory of Environmental & Viral Oncology, College of Life Science and Bioengineering, Beijing University of Technology, Beijing 100124, China.
Int J Environ Res Public Health. 2019 Jan 9;16(2):178. doi: 10.3390/ijerph16020178.
N'-nitrosonornicotine (NNN) is one of the tobacco-specific nitrosamines (TSNAs) that exists widely in smoke and smokeless tobacco products. NNN can induce tumors in various laboratory animal models and has been identified by International Agency for Research on Cancer (IARC) as a human carcinogen. Metabolic activation of NNN is primarily initiated by cytochrome P450 enzymes (CYP450s) via 2'-hydroxylation or 5'-hydroxylation. Subsequently, the hydroxylating intermediates undergo spontaneous decomposition to generate diazohydroxides, which can be further converted to alkyldiazonium ions, followed by attacking DNA to form various DNA damages, such as pyridyloxobutyl (POB)-DNA adducts and pyridyl-N-pyrrolidinyl (py-py)-DNA adducts. If not repaired correctly, these lesions would lead to tumor formation. In the present study, we performed density functional theory (DFT) computations and molecular docking studies to understand the mechanism of metabolic activation and carcinogenesis of NNN. DFT calculations were performed to explore the 2'- or 5'- hydroxylation reaction of ()-NNN and ()-NNN. The results indicated that NNN catalyzed by the ferric porphyrin (Compound I, Cpd I) at the active center of CYP450 included two steps, hydrogen abstraction and rebound reactions. The free energy barriers of the 2'- and 5'-hydroxylation of NNN are 9.82/8.44 kcal/mol (/) and 7.99/9.19 kcal/mol (/), respectively, suggesting that the 2'-() and 5'-() pathways have a slight advantage. The free energy barriers of the decomposition occurred at the 2'-position and 5'-position of NNN are 18.04/18.02 kcal/mol (/) and 18.33/19.53 kcal/mol (/), respectively. Moreover, we calculated the alkylation reactions occurred at ten DNA base sites induced by the 2'-hydroxylation product of NNN, generating the free energy barriers ranging from 0.86 to 4.72 kcal/mol, which indicated that these reactions occurred easily. The docking study showed that ()-NNN had better affinity with CYP450s than that of ()-NNN, which was consistent with the experimental results. Overall, the combined results of the DFT calculations and the docking obtained in this study provide an insight into the understanding of the carcinogenesis of NNN and other TSNAs.
N'-亚硝降烟碱（NNN）是广泛存在于烟雾和无烟烟草制品中的烟草特异性亚硝胺（TSNAs）之一。NNN 可在各种实验室动物模型中诱导肿瘤，并被国际癌症研究机构（IARC）确定为人类致癌物。NNN 的代谢激活主要通过细胞色素 P450 酶（CYP450s）通过 2'-羟化或 5'-羟化启动。随后，羟化中间体自发分解生成重氮氢氧化物，可进一步转化为烷基重氮离子，然后攻击 DNA 形成各种 DNA 损伤，如吡啶恶丁基（POB）-DNA 加合物和吡啶-N-吡咯烷（py-py）-DNA 加合物。如果不能正确修复，这些损伤会导致肿瘤形成。在本研究中，我们进行了密度泛函理论（DFT）计算和分子对接研究，以了解 NNN 的代谢激活和致癌机制。DFT 计算用于探索（）-NNN 和（）-NNN 的 2'-或 5'-羟化反应。结果表明，NNN 在 CYP450 活性中心的铁卟啉（化合物 I，Cpd I）催化下包括两个步骤，即氢提取和反弹反应。NNN 的 2'-和 5'-羟化的自由能垒分别为 9.82/8.44 kcal/mol（/）和 7.99/9.19 kcal/mol（/），表明 2'-（）和 5'-（）途径具有轻微优势。NNN 在 2'-位和 5'-位发生分解的自由能垒分别为 18.04/18.02 kcal/mol（/）和 18.33/19.53 kcal/mol（/）。此外，我们计算了 NNN 的 2'-羟化产物诱导的十个 DNA 碱基位点发生的烷基化反应的自由能垒，范围为 0.86 至 4.72 kcal/mol，表明这些反应很容易发生。对接研究表明，（）-NNN 与 CYP450s 的亲和力强于（）-NNN，这与实验结果一致。总的来说，本研究中 DFT 计算和对接获得的综合结果提供了对 NNN 和其他 TSNAs 致癌机制的深入理解。
