The general objective of these ongoing studies is to provide a quantitative assessment of biochemical steps that may contribute to the modification of cellular DNA resulting from the metabolism of polycyclic aromatic hydrocarbons (PAH). Unidentified forms of cytochrome P-450 in uninduced rat liver dominate PAH activation. These forms will be purified, and polyclonal and monoclonal antibodies will be developed to quantitate their levels in liver and extrahepatic tissues and their contribution to PAH activation. PAH metabolism in uninduced rat liver requires unidentified forms that may also contribute to PAH activation in extrahepatic tissues. High affinity binding to P-450c by benzo(a)pyrene and certain phenolic metabolites contrasts with far weaker binding by BP-7,8-dihydrodiol. This has a major impact on the formation of the ultimate electrophile, (+)anti-BP-7,8- dihydrodiol 9,10-oxide (BPDE). Proposed experiments will evaluate the generality of this selectivity to other PAH and forms of P-450 and will determine the implications of these interactions upon the kinetics of activation of PAH. This is of great importance to understanding the actions of initiation inhibitors and co-carcinogens. Generation of superoxide radicals during PAH metabolism will be investigated (from uncoupled turnover at P-450 or quinone redox cycling?). These studies relate to the possible role of superoxide in promotion. PAH dihydrodiol oxides are generated in cell membranes and bind readily to serum lipoproteins. The effects of individual fatty acids, lipids, and lipoproteins on the reactivity of anti-BPDE will be quantitated with respect to hydrolysis, glutathione conjugation, DNA modification, and reaction with the anti-carcinogenic agent ellagic acid. Activation and detoxication of PAH in hepatocytes will be analyzed with respect to product inhibition of BP metabolism, the relative contributions of reactive electrophiles and superoxide to glutathione depletion, and the distribution of BPDE inside and outside the cells. In mammary cells, we will determine the characteristics of P-450 involvement in PAH metabolism and the effects of metabolism on GSH homeostasis. Finally, the mouse embryo 10 T1/2 cell transformation system will be developed for use with a liposomal activation system containing purified cytochrome P-450 and other purified microsomal enzymes in order to ask more specific questions about metabolic contributions to transformation.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA016265-18
Application #
3164360
Study Section
Chemical Pathology Study Section (CPA)
Project Start
1977-04-01
Project End
1995-11-30
Budget Start
1992-12-01
Budget End
1993-11-30
Support Year
18
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Wisconsin Madison
Department
Type
Schools of Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Halberg, Richard B; Larsen, Michele Campaigne; Elmergreen, Tammy L et al. (2008) Cyp1b1 exerts opposing effects on intestinal tumorigenesis via exogenous and endogenous substrates. Cancer Res 68:7394-402
Zheng, Wenchao; Jefcoate, Colin R (2005) Steroidogenic factor-1 interacts with cAMP response element-binding protein to mediate cAMP stimulation of CYP1B1 via a far upstream enhancer. Mol Pharmacol 67:499-512
Cho, Young C; Zheng, Wenchao; Yamamoto, Megumi et al. (2005) Differentiation of pluripotent C3H10T1/2 cells rapidly elevates CYP1B1 through a novel process that overcomes a loss of Ah Receptor. Arch Biochem Biophys 439:139-53
Cho, Young C; Zheng, Wenchao; Jefcoate, Colin R (2004) Disruption of cell-cell contact maximally but transiently activates AhR-mediated transcription in 10T1/2 fibroblasts. Toxicol Appl Pharmacol 199:220-38
Zheng, Wenchao; Brake, Paul B; Bhattacharyya, Kalyan K et al. (2003) Cell selective cAMP induction of rat CYP1B1 in adrenal and testis cells. Identification of a novel cAMP-responsive far upstream enhancer and a second Ah receptor-dependent mechanism. Arch Biochem Biophys 416:53-67
Zhang, Leying; Zheng, Wenchao; Jefcoate, Colin R (2003) Ah receptor regulation of mouse Cyp1B1 is additionally modulated by a second novel complex that forms at two AhR response elements. Toxicol Appl Pharmacol 192:174-90
Eltom, S E; Zhang, L; Jefcoate, C R (1999) Regulation of cytochrome P-450 (CYP) 1B1 in mouse Hepa-1 variant cell lines: A possible role for aryl hydrocarbon receptor nuclear translocator (ARNT) as a suppressor of CYP1B1 gene expression. Mol Pharmacol 55:594-604
Ikegwuonu, F I; Christou, M; Jefcoate, C R (1999) Regulation of cytochrome P4501B1 (CYP1B1) in mouse embryo fibroblast (C3H10T1/2) cells by protein kinase C (PKC). Biochem Pharmacol 57:619-30
Brake, P B; Arai, M; As-Sanie, S et al. (1999) Developmental expression and regulation of adrenocortical cytochrome P4501B1 in the rat. Endocrinology 140:1672-80
Alexander, D L; Zhang, L; Foroozesh, M et al. (1999) Metabolism-based polycyclic aromatic acetylene inhibition of CYP1B1 in 10T1/2 cells potentiates aryl hydrocarbon receptor activity. Toxicol Appl Pharmacol 161:123-39

Showing the most recent 10 out of 51 publications