EXCEED THE SPACE PROVIDED. During the current funded period of this grant, we investigated the role of murine glutathione (GSH) transferases (GSTs) in detoxification of activated metabolites (diol epoxides) of polycyclic aromatic hydrocarbons (PAHs), which are suspected human carcinogens. Despite these advances, however, the role of individual human GSTs in defense against activated PAHs is poorly defined. In the present renewal application, we propose to shift emphasis from murine model to human GSTs and investigate their role in detoxification of diol epoxides. Recent studies from our laboratory and by others led us to hypothesize that the Alpha class GST isoenzymes play an important role in diol epoxide inactivation in human tissues. We propose to test this hypothesis by two different but complementary approaches. First, we will determine the kinetic constants and catalytic efficiencies for all four known Alpha class human GST isoenzymes (hGSTA1- 1, hGSTA2-2, hGSTA3-3 and hGSTA4-4) toward a panel of structurally different bay- (chrysene and dibenz[a,h]anthracene) and fjord-region (benzo[c]phenanthrene and benzo[g]chrysene) type PAH diol epoxides (specific aim 1). The kinetic constants for Alpha class GSTs will be compared with those of other classes of human GSTs, including hGSTPI-1, hGSTMI-1 and hGSTTI-1, to substantiate the above hypothesis. Second, we will determine the relative contributions of human GSTs, including the Alpha class isoenzymes, for GSH conjugation of representative bay- and fjord-region diol epoxides using autopsied human liver (a major site for xenobiotic metabolism) and lung (a known target organ for PAH-induced tumorigenesis) (specific aim 2). The second objective of the present renewal application is to gain insights into the structural basis for catalytic differences in diol epoxide-GSH conjugation efficacy between human GSTs. This goal will be accomplished by two different approaches: First, we will determine the effects of mutations of key active site (H-site) residues on catalytic activity of Alpha class GSTs toward diol epoxides (specific aim 3). Second, X-ray crystallography studies are planned to elucidate the structural basis for differential substrate specificity pattern for human GSTs toward bay- and _ord-region diol epoxides (specific aim 4). In summary, the studies proposed in the present renewal application, which is a logical extension of our previous findings, will fill the gaps in our understanding of the mechanisms of diol epoxide inactivation in humans. In the long-term, this knowledge will guide us in developing strategies for prevention of PAH-induced cancers in humans. PERFORMANCE SITE ========================================Section End===========================================

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA076348-08
Application #
6838781
Study Section
Chemical Pathology Study Section (CPA)
Program Officer
Poland, Alan P
Project Start
1998-09-01
Project End
2007-01-31
Budget Start
2005-02-01
Budget End
2006-01-31
Support Year
8
Fiscal Year
2005
Total Cost
$279,180
Indirect Cost
Name
University of Pittsburgh
Department
Pharmacology
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Xiao, Hui; Rawal, Malika; Hahm, Eun-Ryeong et al. (2007) Benzo[a]pyrene-7,8-diol-9,10-epoxide causes caspase-mediated apoptosis in H460 human lung cancer cell line. Cell Cycle 6:2826-34
Xiao, Hui; Singh, Shivendra V (2007) p53 regulates cellular responses to environmental carcinogen benzo[a]pyrene-7,8-diol-9,10-epoxide in human lung cancer cells. Cell Cycle 6:1753-61
Srivastava, Sanjay K; Bansal, Pallavi; Oguri, Tetsuya et al. (2007) Cell division cycle 25B phosphatase is essential for benzo(a)pyrene-7,8-Diol-9,10-epoxide induced neoplastic transformation. Cancer Res 67:9150-7
Xiao, Dong; Herman-Antosiewicz, Anna; Antosiewicz, Jedrzej et al. (2005) Diallyl trisulfide-induced G(2)-M phase cell cycle arrest in human prostate cancer cells is caused by reactive oxygen species-dependent destruction and hyperphosphorylation of Cdc 25 C. Oncogene 24:6256-68
Xiao, Dong; Choi, Sunga; Lee, Yong J et al. (2005) Role of mitogen-activated protein kinases in phenethyl isothiocyanate-induced apoptosis in human prostate cancer cells. Mol Carcinog 43:130-40
Singh, Shivendra V; Srivastava, Sanjay K; Choi, Sunga et al. (2005) Sulforaphane-induced cell death in human prostate cancer cells is initiated by reactive oxygen species. J Biol Chem 280:19911-24
Herman-Antosiewicz, Anna; Singh, Shivendra V (2005) Checkpoint kinase 1 regulates diallyl trisulfide-induced mitotic arrest in human prostate cancer cells. J Biol Chem 280:28519-28
Xiao, Dong; Zeng, Yan; Choi, Sunga et al. (2005) Caspase-dependent apoptosis induction by phenethyl isothiocyanate, a cruciferous vegetable-derived cancer chemopreventive agent, is mediated by Bak and Bax. Clin Cancer Res 11:2670-9
Choi, Sunga; Singh, Shivendra V (2005) Bax and Bak are required for apoptosis induction by sulforaphane, a cruciferous vegetable-derived cancer chemopreventive agent. Cancer Res 65:2035-43
Sharma, Rajendra; Brown, David; Awasthi, Sanjay et al. (2004) Transfection with 4-hydroxynonenal-metabolizing glutathione S-transferase isozymes leads to phenotypic transformation and immortalization of adherent cells. Eur J Biochem 271:1690-701

Showing the most recent 10 out of 35 publications