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Defining PAH-mediated carcinogenesis in lung: CYP metabolism and B[a]P transport

定义 PAH 介导的肺癌致癌作用：CYP 代谢和 B[a]P 转运

基本信息

批准号：
7502209
负责人：
Stacy Lynn Gelhaus
金额：
$ 4.96万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-09-30 至 2010-09-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7502209
关键词：
2&apos -deoxyadenosine ABCB1 gene ABCC1 gene Address Adipose tissue Amines Amino Acids Animals Area Aromatic Polycyclic Hydrocarbons Benz(a)Anthracenes Benzo(a)pyrene Binding Biological Biological Assay CYP1A1 gene CYP3A4 gene CYP3A5 gene Cancer Etiology Carcinogens Cations Cattle Cause of Death Cell Line Cell Nucleus Cells Characteristics Coal CpG Islands Cultured Cells Cytochrome P450 Cytochromes DNA DNA Adduction DNA Adducts DNA Damage Data Deoxyguanosine Development Dimethyl Sulfoxide Dioxins Drug Delivery Systems Drug Metabolic Detoxication Environmental Carcinogens Environmental Pollutants Enzymes Epoxide hydrolase Experimental Models Exposure to Family member Fire - disasters Food Garbage Gases Genomics Glutathione Glutathione S-Transferase Glycol Glycoproteins Guanine Hepatocyte Human Human Milk Hydrolysis In Vitro Individual Individual Differences International Agency for Research on Cancer Itraconazole Kidney Link Liquid Chromatography Literature Liver Lung Malignant Neoplasms Malignant neoplasm of lung Malignant neoplasm of urinary bladder Mass Spectrum Analysis Meat Mediating Messenger RNA Metabolic Metabolic Activation Metabolic Pathway Metabolism Methodology Methods Modeling Monitor Multidrug Resistance Associated Protein 1 Mus Mutation Names Nature Null Lymphocytes Oils Oxides P-Glycoprotein Pathway interactions Pharmaceutical Preparations Phase Play Point Mutation Promoter Regions Protein Isoforms Protein p53 Proteomics Public Health Pyrenes RNA Interference Reaction Reporting Research Reverse Transcriptase Polymerase Chain Reaction Role Skin Small Interfering RNA Smoke Smoker Stable Isotope Labeling TP53 gene Testing Tobacco Tobacco smoke Toxic effect Transfection Transferase United States United States Dept. of Health and Human Services Western Blotting Wood material Workplace adduct anticancer research base benzanthracene carbonyl reductase (NADPH)carcinogenesis cigarette smoking cigarette smoking cytochrome P450 3A extracellular fluoranthene hepatoma cell inhibitor/antagonist insight lung carcinogenesis multidrug resistance-associated protein 2 novel novel therapeutics prevent protein expression pyrene research study stable isotope tobacco exposure transcription factor

项目摘要

DESCRIPTION (provided by applicant): In this proposed research, benzo[a]pyrene (B[a]P) and it's metabolites, will be used as model compounds to determine the major metabolic pathways of PAHs in lung carcinogenesis. It is generally accepted that cytochrome P4501A1, CYP1A1, is primarily responsible for the metabolism of B[a]P to 7,8-dihydroxy-9,10- epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (B[a]PDE), the ultimate DNA-adduct forming, carcinogenic reactive metabolite; thus connecting CYP1A1 to the toxicological effects of this environmental carcinogen. DNA-adduct formation has been detected at CpG islands located in the promoter region of the tumor suppressor p53. Inactivation of p53 by point mutations is seen in over 40% of all cancers. However, in preliminary research using H358, human bronchoalveolar lung cells, it was found that DNA-adduct levels were higher in non-treated CYP1A1/1B1 null cells compared to H358 cells in which CYP1A1 expression was induced with 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD). This data correlates with literature results of CYP1A1 KO mice stuides in which it was determined that CYP1A1 conferred a protective rather than a toxic effect. This is indicative of the metabolic activity of another CYP in the metabolism of B[a]P. Several other CYP enzymes are capable of catalyzing this metabolism. Preliminary data has also indicated that TCDD may have an effect on the transport of B[a]P-reactive metabolites. It is known that TCDD is able to induce several of the ABCC drug transporter family members; therefore, explaining why less DNA-adduct formation is observed in lung cells pretreated with TCDD . This proposal, which has two aims, will identify the predominate CYP responsible for B[a]P metabolism in the lung and determine the role of B[a]PDE transporters in DNA-adduct formation in lung cells. A stable isotope dilution LC-MRM/MS method, previously developed, will be used for DNA-adduct quantification. DNA-adduct levels will be correlated to mRNA and protein expression determined by RT-PCR and a novel quantitative proteomics method, SI LAC. CYPs including 1A1, 1B1, 1A2, 2B6, 2E1, 2J2, 3A4, and 3A5 will be investigated. DNA-adduct formation will also be monitored in lung cells in which drug transporter expression has been inhibited by siRNA. Continuing research in this area will provide exciting new therapeutic strategies (drug targets) that can be employed to prevent PAH-mediated DNA damage. Relevance to public health: Cigarette smoke is known to cause 90% of lung cancer, but only 10% of smokers develop lung cancer. Examination of alternative CYP metabolism and B[a]PDE transport in lung cells will provide an experimental model to explore whether inter-individual differences in CYP metabolism and transport contribute to these phenomenon.

描述（由申请人提供）：在本研究中，苯并[a]芘（B[a]P）及其代谢产物将被用作模型化合物，以确定多环芳烃在肺癌发生中的主要代谢途径。一般认为，细胞色素P4501 A1（CYP 1A 1）主要负责将B[a]P代谢为7，8-二羟基-9，10-环氧-7，8，9，10-四氢苯并[a]芘（B[a]PDE），最终形成DNA加合物，致癌反应性代谢物;因此将CYP 1A 1与这种环境致癌物的毒理学效应联系起来。DNA加合物的形成已在位于肿瘤抑制基因p53的启动子区的CpG岛上检测到。p53点突变的失活在超过40%的癌症中可见。然而，在使用人支气管肺泡肺细胞H358的初步研究中，发现与用2，3，7，8-四氯二苯并-对-二恶英（TCDD）诱导CYP 1A 1表达的H358细胞相比，未处理的CYP 1A 1/1B 1无效细胞中的DNA加合物水平更高。该数据与CYP 1A 1 KO小鼠研究的文献结果相关，其中确定CYP 1A 1具有保护作用而非毒性作用。这表明在B[a] P的代谢中另一种酶的代谢活性。其他几种酶能够催化这种代谢。初步数据还表明，四氯二苯并对二恶英可能对B[a] P反应性代谢物的转运产生影响。已知TCDD能够诱导ABCC药物转运蛋白家族的几个成员;因此，解释了为什么在用TCDD预处理的肺细胞中观察到较少的DNA加合物形成。该建议有两个目的，将确定肺中负责B[a]P代谢的主要酶，并确定肺细胞中B[a]PDE转运蛋白在DNA加合物形成中的作用。先前开发的稳定同位素稀释LC-MRM/MS方法将用于DNA加合物定量。DNA加合物水平将与通过RT-PCR和新的定量蛋白质组学方法SI LAC确定的mRNA和蛋白质表达相关。将研究CYP，包括1A 1、1B 1、1A 2、2B 6、2 E1、2 J2、3A 4和3A 5。还将在药物转运蛋白表达已被siRNA抑制的肺细胞中监测DNA加合物形成。在这一领域的持续研究将提供令人兴奋的新的治疗策略（药物靶点），可用于预防PAH介导的DNA损伤。与公共卫生的相关性：已知吸烟导致90%的肺癌，但只有10%的吸烟者患肺癌。对肺细胞中的替代性β-D-代谢和B[a]PDE转运的检查将提供一个实验模型，以探索β-D-代谢和转运的个体间差异是否有助于这些现象。