Polycyclic aromatic hydrocarbons (PAH) are ubiquitous environmental air pollutants that result from fossil fuel combustion and cigarette smoking. PAH exposure is a major risk factor in human lung carcinogenesis. A critical step in multi-stage carcinogenesis is the mutagenic event that results from the formation of DNA adducts. There are three principle routes of metabolic activation of PAH resulting in the formation of diol- epoxides, radical cations, or reactive and redox-active o-quinones. Each of these reactive metabolites have the potential to form DNA-adducts and these adducts may lead to mutation. We are using two approaches to model PAH carcinogenesis, a highly versatile yeast system and human lung cell lines. In lung cancer, the gene most often mutated is p53 where three distinguishing characteristics are found in lung cancer. (1) The pattern of mutations is dominated by G to T transversions;(2) The spectrum of mutations reveals hotspot codons, a few of which are unique to lung cancer;(3) The pattern and spectrum of mutations show a strand bias for mutations on the non-transcribed strand. Our systems use genetic selection methods so that change-in- function mutations can be detected with ease. The yeast reporter system for p53 relies on wild-type p53 binding to a promoter to drive an adenine reporter causing mutant colonies turn red. Our preliminary data suggest that PAH o-quinones, when permitted to undergo redox-cycling, generate 8-oxo-dGuo to cause predominantly G to T transversions with a modest, but significant, preference for hotspots but with no strand bias. We recently devised a system to detect mutations in human lung cells using a p53 dependent promoter to direct transcription of the Herpes TK gene. In this system p53+ cells are killed by exposure to ganciclovir while p53 cells are resistant. The mutant p53 is next rescued from the cells using the yeast system and then sequenced. Using these assays to model p53 mutagenesis we will: (1) Determine the role of repair genes on p53 mutagenesis and compare different PAH metabolites, (2) Map the locations of PAH induced DNA lesions in the p53 gene(3) determine if PAH-metabolites can mutate p53 in lung cells. Our hypothesis is that PAH o- quinones and the ROS they generate provide a route to the p53 mutations found in lung cancer.

Public Health Relevance

Polycyclic aromatic hydrocarbons (PAH) are ubiquitous environmental air pollutants that result from fossil fuel combustion and cigarette smoking. PAH exposure is a major risk factor in human lung carcinogenesis. This proposal will study the mutagenesis of the p53 oncogene by PAH and their metabolites.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES015662-05
Application #
8235068
Study Section
Chemo/Dietary Prevention Study Section (CDP)
Program Officer
Reinlib, Leslie J
Project Start
2008-04-08
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2014-03-31
Support Year
5
Fiscal Year
2012
Total Cost
$307,277
Indirect Cost
$108,807
Name
University of Pennsylvania
Department
Pharmacology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104