Nicotinic receptor-mediated action of tobacco nitrosamines on respiratory cells

描述（由申请人提供）：长期目标：确定呼吸细胞中表达的尼古丁乙酰胆碱受体（nAChRs）在介导尼古丁衍生亚硝胺的致癌作用中的作用，并确定烟草相关致癌的解毒剂。理由：尼古丁，4-（甲基亚硝胺）-1-（3-吡啶基）-1-丁酮（NNK）和N'-亚硝基烟碱（NNN）可以特异性结合nachr并改变肺细胞的生长。我们的初步研究表明，烟碱拮抗剂可以阻断NNK和NNN与肺细胞的结合，并消除这些亚硝胺对细胞增殖、凋亡和非锚定生长的影响。工作假设：nAChRs的功能失活可以：1)在体外和体内肺肿瘤发生模型中消除NNK和NNN的致癌作用；2)防止呼吸细胞胆碱能受体结构和功能的改变。具体目的：研究：1)肺nAChRs在人支气管上皮BEP2D细胞和A/J小鼠培养中介导NNK和NNN的致癌作用；2)暴露的小鼠呼吸细胞和肿瘤中胆碱能受体基因表达和配体结合能力的改变。方法：为了确保准确地将特定的nAChR亚型“分配”到特定的致癌物，我们将使用重叠的方法来消除NNK和NNN的影响。我们将确定nAChR选择性药物，小干扰RNA，或反义寡核苷酸，可以消除肿瘤诱导活性的测试亚硝胺。意义：该结果将为确定未来研究的重点，阐明特定nAChR亚型在烟草相关致癌和肺癌化学预防中的作用提供重要信息。描述：提出的研究阐述了烟草致癌物对靶细胞受体介导作用的新范式，并提出了一个充分证实的假设，即戒烟者肺癌发病率的增加是由于尼古丁诱导的肺细胞内局部激素乙酰胆碱的结合和信号的改变。拟议的研究将确定每个nAChR亚型在对烟草特异性亚硝胺的反应中参与呼吸道恶性转化过程的作用。这些发现将为今后研究烟草亚硝胺介导致癌和促肿瘤作用的细胞内信号通路的机制打开一扇门。

DESCRIPTION (provided by applicant): Long-term Objective: To establish contribution of nicotinic acetylcholine receptors (nAChRs) expressed in respiratory cells to mediating the oncogenic action of the nicotine-derived nitrosamines and identify antidotes to the tobacco-related carcinogenesis. Rationale: Nicotine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N'-nitrosonornicotine (NNN) can specifically bind to nAChRs and alter growth of pulmonary cells. Our preliminary studies indicate that nicotinic antagonists can block binding of NNK and NNN to pulmonary cells and abolish effects of these nitrosamines on cell proliferation, apoptosis and anchorage-independent growth. Working Hypotheses: functional inactivation of nAChRs can: 1) abolish the oncogenic effects of NNK and NNN in in vitro and in vivo models of lung tumorigenesis; and 2) prevent alterations in the cholinergic receptor structure and function in respiratory cells. Specific Aims: to determine: 1) the roles for lung nAChRs in mediating oncogenic effects of NNK and NNN in cultures of human bronchial epithelial BEP2D cells and A/J mice; and 2) alterations in the gene expression and ligand-binding abilities of cholinergic receptors in the exposed respiratory cells, and tumors in mice. Methodology: To assure accurate "assignment" of the specific nAChR subtypes to a particular carcinogen, we will use overlapping approaches to abolish the effects of NNK and NNN. We will identify the nAChR- selective drug, small interfering RNA, or antisense oligonucleotides, that can abolish tumor-inducing activities of test nitrosamines. Significance: The results will provide crucial information for identifying the focus of future research toward elucidation of the role of specific nAChR subtypes in tobacco-related carcinogenesis and lung cancer chemoprevention. Description: The proposed research elaborates a novel paradigm of receptor-mediated action of tobacco carcinogens on target cells, and a well-substantiated hypothesis that an increased frequency of lung cancer in former smokers results from nicotine-induced alterations of binding to and signaling within the lung cells of the local hormone acetylcholine. The proposed studies will establish the role for each nAChR subtype involved in the process of malignant transformation of respiratory in response to the tobacco-specific nitrosamines. These findings will open a door for future mechanistic studies of the intracellular signaling pathways mediating the carcinogenic and tumor-promoting actions of tobacco nitrosamines.