Smoking-induced lung cancer is characterized by an abnormally regulated expression of normal cellular genes resulting from heritable alterations in DNA sequence, e.g. mutations, gene rearrangements, and changes in DNA methylation patterns. Endogenous cytosine methylation controls gene expression by mediating the binding of specific proteins (methyl-CpG binding proteins) to MeCG sites (MeC = 5-methylcytosine) and the recruitment of histone-modifying enzymes that promote chromatin remodeling. However, the exact mechanisms by which the altered methylation patterns arise in lung cancer are not well understood. The presence of MeC induces small, but noticeable changes in DNA structure and dynamics. Previous studies have revealed that MeC is capable of increasing the reactivity of guanine bases in MeCG dinucleotides towards carcinogens. Remarkably, the majority of lung cancer mutational """"""""hot spots"""""""" observed within the p53 tumor suppressor gene are found at endogenously methylated MeCG dinucleotides, e.g. p53 codons 157, 158, 245, 248, and 273. Our long-range goal is to elucidate the molecular mechanisms of smoking-induced lung tumor initiation. The objective of this application is to examine the means by which tobacco carcinogen-DNA adduct formation at MeCG dinucleotides induces genetic and epigenetic changes observed in lung cancer. Our central hypothesis is that endogenous cytosine methylation and the local DNA sequence context control the rates of formation and repair of tobacco carcinogen-DNA adducts. Guided by strong preliminary data, this hypothesis will be tested by pursuing the following four specific aims: 1) Identify the mechanisms by which 5-methylcytosine (MeC) influences the yields of B[a]P diolepoxide-induced N2-guanine adducts within CG dinucleotides. 2) Examine the effects of MeC and its structural analogs on the stereochemistry of N2-guanine adducts induced by B[a]P diolepoxides. 3) Map the distribution of oxidative DNA lesions within p53 and K-ras derived DNA sequences. 4) Examine O6-alkylguanine DNA alkyltransferase-catalyzed repair of NNK-induced O6-guanine lesions at methylated and unmethylated CG dinucleotides. Our approach is innovative, because we will be employing a novel, mass spectrometry based approach developed in our laboratory and novel structural models to identify the effects of endogenous cytosine methylation on reactivity of neighboring guanines towards tobacco carcinogens.

Public Health Relevance

The proposed research is significant because of the widespread human exposure to tobacco products and because of their central role in lung cancer initiation. Our studies will provide an increased understanding of the structural origins of genetic and epigenetic changes associated with lung cancer, which will facilitate the development of cancer treatment and chemoprevention strategies for individuals at risk. An estimated 80-90% of total lung cancer cases are the result of cigarette smoking. Binding of tobacco carcinogens to genomic DNA is considered critical for lung cancer initiation in smokers. The present work will employ a novel, mass spectrometry based approach to investigate the mechanisms by which the local DNA sequence and endogenous methylation of cytosine influence the rates of formation and repair of tobacco carcinogen-DNA adducts. The rationale for these studies is that an increased understanding of the structural origins of genetic and epigenetic changes associated with lung cancer will facilitate the development of cancer treatment and chemoprevention strategies for individuals at risk.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA095039-09
Application #
8290511
Study Section
Cancer Etiology Study Section (CE)
Program Officer
Johnson, Ronald L
Project Start
2002-04-01
Project End
2014-07-31
Budget Start
2012-08-01
Budget End
2014-07-31
Support Year
9
Fiscal Year
2012
Total Cost
$206,571
Indirect Cost
$57,395
Name
University of Minnesota Twin Cities
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
555917996
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Wickramaratne, Susith; Seiler, Christopher L; Tretyakova, Natalia Y (2015) Synthesis of DNA Oligodeoxynucleotides Containing Site-Specific 1,3-Butadiene-Deoxyadenosine Lesions. Curr Protoc Nucleic Acid Chem 61:4.61.1-22
Ming, Xun; Matter, Brock; Song, Matthew et al. (2014) Mapping structurally defined guanine oxidation products along DNA duplexes: influence of local sequence context and endogenous cytosine methylation. J Am Chem Soc 136:4223-35
Kotandeniya, Delshanee; Murphy, Daniel; Yan, Shuo et al. (2013) Kinetics of O(6)-pyridyloxobutyl-2'-deoxyguanosine repair by human O(6)-alkylguanine DNA alkyltransferase. Biochemistry 52:4075-88
Tretyakova, Natalia; Villalta, Peter W; Kotapati, Srikanth (2013) Mass spectrometry of structurally modified DNA. Chem Rev 113:2395-436
Kim, Min Young; Lim, Chang Hoon; Trudel, Laura J et al. (2012) Delivery method, target gene structure, and growth properties of target cells impact mutagenic responses to reactive nitrogen and oxygen species. Chem Res Toxicol 25:873-83
Tretyakova, Natalia; Goggin, Melissa; Sangaraju, Dewakar et al. (2012) Quantitation of DNA adducts by stable isotope dilution mass spectrometry. Chem Res Toxicol 25:2007-35
Guza, Rebecca; Kotandeniya, Delshanee; Murphy, Kristopher et al. (2011) Influence of C-5 substituted cytosine and related nucleoside analogs on the formation of benzo[a]pyrene diol epoxide-dG adducts at CG base pairs of DNA. Nucleic Acids Res 39:3988-4006
Kotandeniya, Delshanee; Murphy, Dan; Seneviratne, Uthpala et al. (2011) Mass spectrometry based approach to study the kinetics of O6-alkylguanine DNA alkyltransferase-mediated repair of O6-pyridyloxobutyl-2'-deoxyguanosine adducts in DNA. Chem Res Toxicol 24:1966-75
Seneviratne, Uthpala; Antsypovich, Sergey; Goggin, Melissa et al. (2010) Exocyclic deoxyadenosine adducts of 1,2,3,4-diepoxybutane: synthesis, structural elucidation, and mechanistic studies. Chem Res Toxicol 23:118-33
Seneviratne, Uthpala; Antsypovich, Sergey; Dorr, Danae Quirk et al. (2010) DNA oligomers containing site-specific and stereospecific exocyclic deoxyadenosine adducts of 1,2,3,4-diepoxybutane: synthesis, characterization, and effects on DNA structure. Chem Res Toxicol 23:1556-67

Showing the most recent 10 out of 20 publications