Research Interests where Dr. Monks is Principal Investigator

Molecular Basis of ROS-induced Cell Death

Mechanisms of cell death are usually classified into two pathways, apoptosis and necrosis.

However, it has been proposed by the American Society of Toxicologic Pathologists that the term oncosis, with its root meaning of "swelling" be used as the alternate descriptor of cell death occurring by non-apoptotic pathways.

Necrosis more accurately describes the consequences of oncotic cell death, usually the death of a large number of cells which results in moderate to severe tissue injury. Oncosis is a form of cell death that typically occurs in response to toxic injury, including that induced by chemical exposure and reactive oxygen species (ROS).

ROS are involved in the initiation and progression of a variety of human diseases and toxicities associated with chemical exposure. An understanding of the factors that regulate the cellular stress response to ROS and of the molecular mechanisms by which they interact with cellular constituents, and the consequences of such interactions, are important fundamental goals of biomedical research.

Our data, and that of others, indicate that responses to stress that usually result in oncotic cell death (and tissue necrosis) can be manipulated, at the genetic and pharmacological level, to produce a potentially favorable (survivable) tissue response. Basic knowledge of the mechanisms by which ROS induce cell death may yield strategies for clinical interventions in pathologies in which ROS play a prominent role.

The laboratory has projects that are investigating the cellular and molecular mechanisms by which ROS induce both apoptotic and oncotic/necrotic cell death.

References: Oncotic/necrotic cell death

Jeong, J.K., Stevens, J.L., Lau, S.S., and Monks, T.J. Quinone-thioether-mediated DNA damage, growth arrest, and gadd153 expression in renal proximal tubular epithelial cells. Molec. Pharmacol., 50:592-598, 1996.
Jeong, J.K., Huang, Q., Lau, S.S., and Monks, T.J. The response of renal epithelial cells to physiological-and chemical-induced growth arrest. J. Biol.Chem., 272:7511-7518, 1997.
Jeong, J.K., Dybing, E., Søderlund, E., Brunborg, G., Holme, J.A., Lau, S.S., and Monks, T.J. DNA damage, gadd153 expression, and cytotoxicity in plateau phase renal proximal tubular epithelial cells treated with a quinol-thioether. Arch. Biochem. Biophys., 341:300-308, 1997.
Huang, Q., Lau, S.S., and Monks, T.J. Induction of gadd153 mRNA by nutrient deprivation is overcome by glutamine. Biochem. J., 341:225-231, 1999.
Jeong, J.K., Wogan, G.N., Lau, S.S., and Monks, T.J. Quinol-glutathione conjugate-induced mutation spectra in the supf gene replicated in human AD293 cells and bacterial MBL50 cells. Cancer Res., 59:3641-3645, 1999.
Tikoo, K., Lau, S.S., and Monks, T.J. Histone H3 phosphorylation is coupled to poly (ADP-ribosylation) and reactive oxygen species-induced cell death in renal proximal tubular epithelial cells. Molec. Pharmacol. 60:394-402, 2001.
Ramachandiran, S., Huang, Q., Dong, J., Lau, S.S., and Monks, T.J. Mitogen activated protein kinases contribute to reactive oxygen species-induced cell death in renal proximal tubule epithelial cells. Chem. Res, Toxicol., 15, 1635-1642, 2002.
Dong, J., Everitt, J., Lau, S.S., and Monks, T.J. Induction of ERK1/2 and histone H3 phosphorylation within the outer stripe of the outer medulla of the Eker rat by 2,3,5-tris-(glutathion-S-yl)hydroquinone. Toxicol Sci. 80, 350-357, 2004.

References: Apoptotic cell death

Bratton, S.B., Lau, S.S., and Monks, T.J. Identification of quinol-thioethers in bone marrow of hydroquinone/phenol treated rats and mice, and their potential role in benzene-mediated hematotoxicity. Chem. Res. Toxicol., 10:859-865, 1997.
Bratton, S.B., Lau, S.S., and Monks, T.J. The putative benzene metabolite, 2,3,5-tris-(glutathion-S-yl)hydroquinone, depletes glutathione, stimulates sphingomyelin turnover, and induces apoptosis in Hl-60 cells. Chem. Res, Toxicol., 13:550-556, 2000.

Neurotoxicology of Ecstasy

3,4-(±)-Methylenedioxymethamphetamine (MDMA; Ecstasy, XTC, E) is a Schedule 1 synthetic, psychoactive drug possessing stimulant and hallucinogenic properties.

The subjective effects of MDMA (a heightened sense of awareness and feelings of increased empathy or emotional closeness to others) have contributed to its popularity as a "party drug" amongst adolescents and young adults who frequent late night "raves' or "techo-parties".

Recent reports estimate that over 2 million tablets of MDMA are smuggled into the U.S. each week. Several adverse effects are associated with the use of MDMA, the most worrisome of which is long-term toxicity to the serotonergic neurotransmitter system. MDMA use and abuse therefore has the potential to give rise to a major public health problem.

The neurotoxic effects of MDMA are dependent on the route and frequency of drug administration. Direct injection of either MDMA into the brain fails to reproduce the neurotoxicity following peripheral administration, indicating that the parent amphetamines are unlikely to be solely responsible for the neurotoxic effects. It has therefore been proposed that metabolites of MDMA mediate the neurotoxic effects.

However, none of the known major metabolites of these drugs are capable of reproducing their neurobehavioral and neurotoxicological effects. We hypothesize that quantitatively minor, yet reactive metabolites of MDA and MDMA contribute to their neurotoxicity, and we hypothesize that one such class of metabolites arise from the oxidation of N-methyl-a-methyldopamine, followed by scavenging of the ortho-quinones with GSH. Ongoing experiments are designed to test this overall hypothesis and to examine the mechanism by which such metabolites produce selective serontonergic neurotoxicity.

References

Miller, R.T., Lau, S.S., and Monks, T.J. Metabolism of 5-(glutathion-S-yl)-a-methyldopamine following intracerebroventricular administration to male Sprague Dawley rats. Chem. Res. Toxicol., 8:634-641, 1995.
Miller, R.T., Lau, S.S., and Monks, T.J. Effects of 5-(glutation-S-yl)-a-methyldopamine on dopamine, serotonin, and norepinephrine concentrations following intracerebroventricluar administration to male Sprague Dawley rats. Chem. Res. Toxicol., 9:457-465, 1996.
Miller, R.T., Lau, S.S., and Monks, T.J. 2,5-bis-(Glutathion-S-yl)-a-methyldopamine, a putative metabolite of (±)-3,4-methylenedioxyamphetamine, produces long-term decreases in brain serotonin concentrations. Eur. J. Pharmacol., 323:173-180, 1997.
Monks, T.J., Ghersi-Egea, J-F., Philbert, M.A., Cooper, A.J.L., and Lock, E.A. The role of glutathione in neuroprotection and neurotoxicity. Toxicol. Sci., 51:161-177, 1999.
Bai. F., Lau, S.S., and Monks, T.J. Glutathione and N-acetylcysteine conjugates of a-methyldopamine produce serotonergic neurotoxicity. Possible role in methylenedioxyamphetamine-mediated neurotoxicity. Chem. Res. Toxicol., 12:1150-1157, 1999. Bai, F., Lau, S.S., and Monks, T.J. The serotonergic neurotoxicity of 3,4-(±)-methylenedioxy-amphetamine and 3,4-(±)-methylenedioxymetamphetamine (ecstasy) is potentiated by inhibition of g-glutamyl transpeptidase. Chem. Res. Toxicol. 14: 863-870, 2001.
Monks, T.J.,Bai, F., Miller, R.T., Lau, S.S. Serontonergic neurotoxicity of of methylenedioxy-amphetamine and methylenedioxymetamphetamine. Adv. Exp. Med. Biol., 500, 397-406, 2001.
Monks, T.J., Jones, D.C., Bai, F., and Lau, S.S. The role of metabolism in 3,4-(±)-methylenedioxy-amphetamine and 3,4-(±)-methylenedioxymethamphetamine (ecstasy) neurotoxicity. Ther. Drug Monitoring, 26, 132-136, 2004.
Jones, D.C., Lau, S.S., and Monks, T.J. Thioether metabolites of 3,4-(±)-methylenedioxy-amphetamine and 3,4-(±)-methylenedioxymethamphetamine inhibit hSERT function and simultaneously stimulate dopamine uptake into hSERT-expressing SK-N-MC cells. J Pharmacol Exp Ther. 2004 May 28 [Epub ahead of print] PMID: 15169827.

The following projects represent collaborations with Dr. Serrine S. Lau

Molecular basis of renal carcinogenesis

It is apparent that many genetic alterations are required for tumor formation. However, no single oncogene or tumor suppressor gene is activated or deleted in all cancers. Even tumors from a single organ rarely exhibit uniform genetic alterations.

Thus, it is not clear how many genetic changes are required to transform a normal cell into a malignant one. Recent research suggests that the number of genomic alterations may be quite large.

However, if one assumes that most of these mutations are irrelevant to cell survival, then a large number of mutations within the genome would alter the protein products of a much smaller number of genes involved in regulating cell growth.

Therefore, it is essential to understand which genomic changes lead to changes at the protein level relevant to tumorigenesis, and to understand how changes at the genome level are coupled to changes in the proteome and subsequently, tumorigenesis.

Several potential genetic targets for hereditary and sporadic renal cell carcinoma (RCC) have been identified. Three genes, the von Hippel-Lindau (VHL) and Tuberous Sclerosis-2 (Tsc-2) tumor suppressor genes and c-met protooncogene, act as determinants of susceptibility for RCC in humans and/or rodents, and are targets for somatic events that lead to the development of spontaneous and carcinogen-induced renal tumors.

Although familial predisposition to renal cancer has led to the identification of genes involved in spontaneous RCC in humans, few genetically susceptible animal models are available to study the induction of this disease by chemical carcinogens. Spontaneous renal cell carcinoma is rare in rats, occurring in most strains with a frequency of <0.05%.

However, the Eker rat carries a single autosomal mutation that predisposes them to the development of spontanous renal cell tumors at a very high incidence. A germline insertion of an endogenous retrovirus in the Tsc-2 gene, on rat chromosome 10q syntenic with human chromosome 16p, is responsible for the predisposing "Eker" mutation.

Alterations in the Tsc-2 gene may be responsible for the development of renal tumors. Loss of heterozygosity (LOH) at the Tsc-2 locus has been demonstrated in renal tumors in humans and in spontaneous or chemically induced RCCs in rats. The majority of renal cell tumors observed in the Eker rat originate from the renal proximal tubules and are histologically similar to renal tumors in humans. This animal model is being used to investigate the molecular progression of chemical-induced renal cancer.

Selected Publications

Peters, M.M., Jones, T.W., Monks, T.J. and Lau, S.S. Cytotoxicity and cell proliferation induced by the nephrocarcinogen hydroquinone, and its nephrotoxic metabolite 2,3,5-tris-(glutathion-S-yl)hydroquinone. Carcinogenesis, 18:2393-2401, 1997.
Towndrow, K.M., Mertens, J.J.W.M., Jeong, J.K., Weber, T.J., Monks, T.J., and Lau, S.S. Stress- and growth related gene expression are independent of chemical-induced prostaglandin E2 synthesis in renal epithelial cells. Chem. Res. Toxicol., 13:111-117, 2000
Lau, S.S., Monks, T.J., Everitt, J.I., Kleymenova, E., and Walker, C.W. Carcinogenicity of a nephrotoxic metabolite of the "non-genotoxic" carcinogen hydroquinone. Chem. Res. Toxicol., 14:25-33, 2001.
Yoon, H.S., Walker, C.L., Monks, T.J., and Lau, S.S. Transformation of kidney epithelial cells by quinol-thioethers via inactivation of the tuberous sclerosis-2 tumor suppressor gene. Molec. Carcinogenesis, 31:37-45, 2001.
Weber, T.J., Huang, Q., Monks, T.J., and Lau, S.S. Differential regulation of redox responsive transcription factors by the nephrocarcinogen, 2,3,5-tris-(glutathion-S-yl)hydroquinone. Chem. Res. Toxicol. 14: 814-821, 2001.
Yoon, H.S., Monks, T.J., Everitt, J.I., Walker, C.L., and Lau, S.S. Cell proliferation is insufficient but loss of tuberin is necessary for chemically induced nephrocarcinogenicity. Am. J. Physiol. Renal Physiol. 283: F262-F270, 2002.
Habib, S.L., Phan, M.F., Monks, T.J., and Lau, S.S. Reduced constitutive 8-oxogaunine-DNA glycosylase expression and impaired induction following oxidative DNA damage in the tuberin deficient Eker rat. Carcinogenesis, 24, 573-582, 2003.
Patel, S.K., Ma, N., Monks, T.J., and Lau, S.S. Changes in gene expression during chemical-induced nephrocarcinogenesis in the Eker rat. Molec. Carcinogenesis, 38, 141-154, 2003.
Yoon, H-S., Ramachandiran, S., Chacko, M.A.S., Monks, T.J., and Lau, S.S. The tuberous sclerosis-2 tumor suppressor modulates ERK and b-Raf activity in transformed renal epithelial cells. Am. J. Physiol. Renal Physiol., 286, F417-F424, 2004.

Proteomics in Toxicology

Proteins have long been appreciated as critical targets of environmental chemicals that produce acute tissue toxicities and cancer. For many years, however, identifying protein targets of reactive chemicals and their metabolites has been a nearly impossible task.

Recent developments in mass spectrometry (MS) ionization methods and instrumentation now make possible the rapid, high throughput analysis of proteins. MS has become the method of choice for sequencing peptides and proteins with limited amounts of available sample.

These analytical capabilities have driven the growth of proteomics, the study of the protein complement of the genome.

The long-term goal of this grant application is to (i) develop mass spectrometric methods to identify protein targets of environmental chemicals, (ii) ascertain those features that predispose certain proteins, or motifs within proteins ("electrophile binding motifs", or EBMs) to chemical adduction, and (iii) determine the potential biological/toxicological consequences of adduction to rationally selected "electrophilins."

One of the most critical arenas for research in functional proteomics is the detection of chemical-induced post-translational modifications (PTMs) that may either indirectly alter cell signaling pathways or directly alter the structure and function of the modified protein. Such chemical-induced PTMs are usually of low abundance and thus not detectable by standard proteomic protocols.

Selected Publications

Person, M.D., Monks, T.J., and Lau, S.S. An integrated approach to identifying chemically induced post-translational modifications using comparative MALDI-MS and targeted HPLC-ESI-MS/MS. Chem. Res, Toxicol., 16, 598-608, 2003.
Person, M.D., Mason, D.E., Liebler, D.C., Monks, T.J., and Lau, S.S. Alkylation of cytochrome c by (glutathion-S-yl)-1,4-benzoquinone and iodoacetamide demonstrates compound dependent site specificity. Chem. Res. Toxicology, Accepted pending revision, 6/30/04.

Prostaglandin-mediated Cytoprotection

Prostaglandins (PGs) play important yet diverse roles in mammalian signaling. A number of PGs and their analogues are able to protect target cells against various toxic, ischemic, and infectious models. This effect of PGs is termed as "cytoprotection".

Much work has been conducted on PGs mediated protection in gastrointestinal tract and liver. Several synthetic PG analogues have been used in clinic to treat NSAID-induced gastropathy, acute liver failure, liver transplantation, and chronic liver disease. However, the mechanism behind the cyprotective effect of PGs still remains largely unknown. Especially very little is known on PGs mediated protective effect in kidney.

Selected Publications

Weber, T.J., Lau, S.S., and Monks, T.J. PGE2 -mediated cytoprotection in renal epithelial cells. Evidence for a pharmacologically distinct receptor. Amer. J. Physiol., 273 (Renal Physiol. 42):F507-F515, 1997.
Weber, T.J., Monks, T.J., and Lau, S.S. DDM-PGE2-mediated cytoprotection in renal epithelial cells by a thromboxane A2 receptor coupled to NF-kB. Amer. J. Physiol. Renal Physiol, 278:F270-F278, 2000.
Towndrow, K.M., Jia, Z., Lo, H-H., Person, M.D., Monks, T.J., and Lau, S.S. 11-Deoxy-16,16-dimethylprostaglandin E2 induces specific proteins in association with its ability to protect against oxidative stress. Chem. Res, Toxicol., 16, 312-319, 2003.
Person, M.D., Lo, H-H., Towndrow, K.M., Jia, Z., Monks, T.J., and Lau, S.S. Comparative identification of prostanoid-inducible proteins by LC-ESI MS/MS and MALDI-TOF mass spectrometry. Chem. Res, Toxicol., 16, 757-767, 2003.
Jia, Z., Person, M.D., Shen, J., Hensley, S.C., Stevens, J.L., Monks, T.J., and Lau, S.S. GRP78/Bip is essential for 11-deoxy-16,16-dimethylprostaglandin mediated cytoprotection in renal epithelial cells. Am. J. Physiol. Renal Physiol. 287, F000-F000, 2004 Jun 29 [Epub ahead of print] PMID: 15226156

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