Induction of MGMT as a Strategy for Chemoprevention

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DESCRIPTION (provided by applicant): Modulation of DNA repair enzymes, leading to facilitated elimination of carcinogenic lesions is likely to be a 6 novel and successful strategy for cancer prevention. MGMT (O6-methylguanine-DNA-methyltransferase), 6 which elicits a direct reversal of guanine-O6 alkylations, plays a central role in protecting the cellular genome from the mutagenic and toxic actions of endogenous, food-derived, environmental, and therapeutic alkylating carcinogens. A vast amount of epidemiological and research data on the reduction of spontaneous and induced tumors in MGMT transgenic mice, attenuation of Ras and p53 mutations in MGMT-proficient cells and the ongoing strategy of MGMT gene transduction into the bone marrow stem cells for achieving alkylator resistance suggest that MGMT is an excellent and rational target for chemoprevention and perhaps chemoprotection. The long-term objective of this pilot (R03) project is to explore the possibility of augmenting MGMT levels through a dietary approach in human normal tissues including the hematopoietic system using a non-toxic cysteine prodrug and antioxidant plant compounds. We have obtained strong evidence for a marked and reproducible increase of MGMT activity (up to 3-fold) in a variety of human cancer cell lines after exposure to a cysteine prodrug (L-2oxothiazolidine-4-carboxylic acid or OTC), and antioxidant compounds such as silymarin, curcumin and ethanol extracts of several medicinal plants. Increased levels of MGMT mRNA and increased rate of MGMT protein synthesis accounted for the enhanced DNA repair activity. OTC is a non-toxic compound, metabolized by the cellular enzyme, 5-oxoprolinase to generate cysteine, whose availability in turn, increases the intracellular GSH levels greatly. Our hypothesis, built on these observations is that OTC and selected phytochemicals will enhance MGMT expression in animal tissues and attenuate the toxicity of clinically used alkylating agents. The major objective of this 2-year pilot project is to establish the feasibility of chemoprevention and chemoprotection via MGMT in a preclinical setting through the following Specific Aims: 1) Quantitate alterations in MGMT expression in hepatic and non-hepatic tissues including the peripheral blood lymphocytes in mice after OTC and antioxidant (curcumin, silymarin, and nimbidin) administrations. 2) Characterize the effect OTC administration on the levels and extent of BCNU, and temozolomide-induced mutations and toxicity in mice. In Specific Aim 1, The DNA repair activity of MGMT, mRNA and protein levels in liver, lung, colon, brain, peripheral blood lymphocytes, and hepatic mRNA levels will be measured after chemopreventative treatments. The treatment duration required for sustained elevation of the repair protein will be assessed. Specific Aim 2 will investigate the mutation frequencies in the Hprt gene of splenic T-lymphocytes, parameters of hematological and overall toxicities induced by BCNU and temozolomide in animals pretreated with OTC. The study promises to pave way for clinical trials of MGMT-targeted chemopreventative strategies. Further, because GSH and GST-pi, likely to be induced, these approaches will provide multiple benefits.