Research in my lab is aimed at defining the role of specific gene products (enzymes) in determining cellular sensitivity to cytotoxic and genotoxic agents, including both cancer chemotherapeutic drugs and carcinogens. One project is focused on the glutathione S-transferase (GST) gene family, isozymes of which confer resistance to chemical damage by some of these agents. Plasmid expression vectors are used to stably transfect the gene of interest into cell lines with low endogenous GST expression. Following testing for stable expression of the introduced gene product, cell lines expressing different GSTs are tested to determine whether each isozyme protects against DNA damage by mutagenic agents.
A second project in my lab has shown that aldehyde dehydrogenases (ALDH) can cause tumor cell metabolic resistance to the anticancer agent cyclophosphamide. Expression of transfected human ALDH3 also confers resistance to protein modification, toxicity, and apoptosis induced by lipid aldehydes that are generated during oxidant stress-induced lipid peroxidation. These and other transgenic cell lines, studied in comparison with their isogenic (empty vector- transfected) control cell lines, serve as genetic model systems for detailed examination of the protective capacity of these defensive pathways against cytotoxicity, DNA damage and mutagenesis by drugs and carcinogens.
A third and new direction in this lab is the use of transgenic cell lines expressing cyclooxygenases to study the synergistic interaction between production of eicosanoid inflammatory mediators and sensitivity of cells to anticancer drugs. The role of oxidative stress in this interaction is a major focus.
Trainees in my laboratory have the opportunity to learn a broad range of research methods, including cell culture, recombinant DNA methods, development and characterization of transgenic cell lines, electrophoretic and blotting analysis, production and purification of recombinant proteins, enzyme kinetic analysis, metabolite analysis methods, cytotoxicity and mutagenicity assays, flow cytometry, and macroarray expression analysis. |
Recent Publications (selected):
Fields WR, Morrow CS, Doehmer J, Townsend AJ. Expression of stably transfected murine glutathione S-transferase A3-3 protects against nucleic acid alkylation and cytotoxicity by aflatoxin B1 in hamster V79 cells expressing rat cytochrome P450-2B1. Carcinogenesis 20:1121-5, 1999.
Townsend AJ, Kiningham KK, St Clair D, Tephly TR, Morrow CS, Guengerich FP. Symposium overview: Characterization of xenobiotic metabolizing enzyme function using heterologous expression systems. Toxicol. Sci. 48:143-50, 1999.
Townsend AJ, Fields WR, Doss AJ, Clapper ML, Doehmer J, Morrow CS. Modeling the chemoprotective functions of glutathione S-transferases in cultured cell lines by heterologous expression. Drug Metab. Rev. 31:43-69, 1999.
Haynes, Robin L., Szweda, Luke, and Townsend, Alan J. Structure-Activity Relationships for Growth Inhibition and Induction of Apoptosis by 4-Hydroxy 2-nonenal in Raw 264.7 Cells. Molecular Pharmacology 58:788-794, 2000.
Townsend, Alan J., Leone-Kabler, Sandra, Haynes, Robin L., Wu, Yinghui, Szweda, Luke and Bunting, Kevin D. Selective Protection By Stably Transfected Human ALDH3A1 (but not Human ALDH1A1) Against Toxicity of Aliphatic Aldehydes in V79 Cells. Chem. Biol. Interact. 130-132:261-273, 2001.
Haynes, R. L., Brune, B., and Townsend, A. J. Apoptosis in RAW 264.7 cells exposed to 4-hydroxy-2-nonenal: dependence on cytochrome C release but not p53 accumulation. Free Rad. Biol. Med. 30:884-894, 2001. |