Chemical Mechanisms of Cytotoxicity
The liver is a major site of metabolism for xenobiotic as well as endogenous compounds. It has long been known that many xenobiotic compounds cause organ-specific injury to the liver (e.g., the hepatotoxicity of chloroform was recorded in the late 19th century). Understanding the mechanism(s) by which chemicals cause cytotoxic liver injury remains a major challenge of enormous scientific and practical importance. Hepatotoxicity remains still the single most important factor in the failure of drug candidates late in development (when such failure is extremely expensive in terms of lost investment of resources), and in the withdrawal of approved drugs early from open clinical usage (when the cost of failure can also include severe or even fatal injury to patients). Examples of drugs recently withdrawn because of hepatotixicity include troglitazone, tienilic acid, bromfenac and benoxaprofen. Other drugs having significant hepatotoxic potential, but which are still used clinically, include acetaminophen, diclofenac, and halothane. Many approaches to elucidating mechanisms of toxicity involve the use of "model" compounds such as bromobenzene, thioacetamide and thiobenzamide.
Our laboratory has isolated and elucidated the structures of about a dozen protein adducts of reactive bromobenzene metabolites, confirmed them by synthesis, and used the adducts to raise antibodies capable of recognizing proteins adducted by these metabolites. We have also applied proteomic techniques and mass spectrometry to demonstrate that protein adduction by bromobenzene metabolites is a remarkably selective process, and to identify specific liver proteins targeted by reactive metabolites of bromobenzene.
An important question we are currently pursuing is whether reactive metabolites of other compounds, particularly thiobenzamide and halothane, target the same subset of proteins as bromobenzene. Information on these proteins has been collected in a publicly accessible database at:
http://tpdb.medchem.ku.edu:8080/protein_database/. In collaboration with a molecular pharmacologist, we are also interested in identifying other proteins with which target proteins interact, and the signaling cascades they trigger that lead to cell death. In addition, we utilize enzymatic and chemical model systems to investigate the formation of reactive metabolites and their reaction with target proteins. This project is a true blend of hard core chemistry and cell biology.
Publications
- Robert P. Hanzlik, Yakov M. Koen, Bharga Theertham, Yinghua Dong and Jianwen Fang. “The reactive metabolite target protein database (TPDB) – a web-accessible resource.” BMC Bioinformatics 8:95 (2007). doi: 10.1186/1471-2105-8-95. http://www.biomedcentral.com/1471-2105/8/95.
- T. Ji, K. Ikehata, Y.M. Koen, S.W. Esch, T.D. Williams and R.P. Hanzlik, “Covalent Modification of Microsomal Lipids by Thiobenzamides in Vivo.” Chem. Res. Toxicol. 20, 701-708 (2007).
- Y.M. Koen, N.V. Gogichaeva, M.A. Alterman and R.P. Hanzlik, “A Proteomic Analysis of Bromobenzene Reactive Metabolite Targets in Rat Liver Cytosol.” Chem. Res. Toxicol. 20, 511-519 (2007).
- Y.M. Koen, W. Yue, N.A. Galeva, T.D. Williams and R.P. Hanzlik, “Site-Specific Arylation of Rat Glutathione S-Transferase A1 and A2 by Bromobenzene Metabolites In Vivo.” Chem. Res. Toxicol. 19, 1426-1434 (2006).
- Weimin Yue, Yakov M. Koen, Todd D. Williams, and Robert P. Hanzlik, "Use of Isotopic Signatures for Mass Spectral Detection of Protein Adduction by Chemically-Reactive Metabolites of Bromobenzene. Studies with Model Proteins." Chem. Res. Toxicol., 18, 1748-1754 (2005).
- Weimin Yue, S. I. Lewis, Y. M. Koen, and R. P. Hanzlik, "Synthesis of N(t)-Arylhistidine Derivatives via Direct N-Arylation," Bioorg. Med. Chem. Let., 14, 1637-1640 (2004).
- Yakov M. Koen and Robert P. Hanzlik, " Identification of Seven Proteins in the Endoplasmic Reticulum as Targets for Reactive Metabolites of Bromobenzene." Chem. Res. Toxicol., 15, 699-706 (2002).
- Y. M. Koen, T. D. Williams, and R. P. Hanzlik, "Identification of Three Protein Targets for Reactive Metabolites of Bromobenzene in Rat Liver Cytosol." Chem. Res. Toxicol., 13, 1326-1335 (2000).
- E. M. Rombach and R. P. Hanzlik, "Detection of Adducts of Bromobenzene-2,3-oxide to Rat Liver Microsomal Protein Sulfhydryl Groups Using Specific Antibodies." Chem. Res. Toxicol., 12, 159-163 (1999).
- E. M. Rombach and R. P. Hanzlik, "Identification of a Rat Liver Microsomal Esterase as a Target Protein for Bromobenzene Metabolites." Chem. Res. Toxicol., 11, 178-184 (1998).
- I. M. C. M. Reitjens, C. den Besten, R. P. Hanzlik, and P. J. van Bladeren, "The Cytochrome P450 Catalyzed Oxidation of Halobenzene Derivatives." Chem. Res. Toxicol., 10, 629-635 (1997).
- E. M. Rombach and R. P. Hanzlik, "Detection of Benzoquinone Adducts to Rat Liver Protein Sulfhydryl Groups Using Specific Antibodies." Chem. Res. Toxicol., 10, 1407-1411 (1997).
- R. B. Bambal and R. P. Hanzlik, "Bromobenzene-3,4-oxide Alkylates Histidine and Lysine Side Chains of Rat Liver Protein in vivo." Chem. Res. Toxicol., 8, 729-735 (1995).
- R. P. Hanzlik, S. P. Harriman, and M. Frauenhoff, "Covalent Binding of Benzoquinone to Reduced Ribonuclease. Adduct Structures and Stoichiometry." Chem. Res. Toxicol., 7, 177-184 (1994).
- R. Bambal and R. P. Hanzlik, "Covalent Addition of Bromobenzene-3,4-oxide to Protein. Synthesis of N(epsilon)-(p-bromophenyl)-Lysine and N(tau)-(p-Bromophenyl)-L-Histidine as Model Adducts." J. Org. Chem., 59, 729-732 (1994).
For more information, please contact:
E-mail:
rhanzlik@ku.edu
4048 Malott Hall
Tel: 785-864-3750
