DNA repair plays a pivotal role in protection from carcinogenesis. The DNA repair protein 0(6)alkylguanine-DNA alkyltransferase protects cells from the mutagenic and cytotoxic DNA damage of agents such as nitrosoureas and nitrosamines which form DNA adducts at the O(6) position of guanine. From in vitro data in bacteria and cells, it is clear that lack of this protein increases the susceptibility to nitrosoureas and nitrosamines whereas cells with high levels of this DNA repair protein withstand much higher levels of exposure. Furthermore, in animals, a correlation exists between low levels of alkyltransferase and susceptibility to the toxic and carcinogenic effects of nitrosoureas. While such a relationship is hard to establish in humans, we have previously shown that bone marrow cells have low alkyltransferase which might explain the high incidence of nitrosourea induced leukemia among patients receiving these compounds. This proposal will test the hypothesis that increasing the expression of alkyltransferase will prevent an animal model of this leukemia. In this model, mice develop nitrosourea induced T cell lymphomas. We have successfully generated transgenic mice which express the human alkyltransferase gene MGMT in a number of tissues including thymus, spleen and bone marrow.
The aims of this proposal are to 1) fully characterize the pattern of expression of the MGMT gene in transgenic mice by studying tissue and cellular expression of the mRNA in alkyltransferase protein. 2) Establish the capacity of the human MGMT gene to protect transgenic mice from the development of MNU induced lymphoma using a variety of dose schedules and genetic backgrounds. 3) Determine whether transgenic expression of human alkyltransferase results in rapid repair of O(6)methylguanine-DNA adducts and whether this rapid repair is uniform or heterogeneous in different tissues. 4) To determine whether transgenic expression of human alkyltransferase prevents mutational activation of k-ras oncogene in the thymus, the oncogene most frequently associated with MNU induced lymphomas and reduces the mutational frequency at the HPRT locus of T lymphocytes. This animal model will determine whether increased expression of a single DNA repair protein can prevent carcinogen induced cancer in mice. These studies have far reaching implications in the understanding of human carcinogenesis and in the use of genetic manipulations to prevent cancer in humans.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES006288-04
Application #
2155166
Study Section
Metabolic Pathology Study Section (MEP)
Project Start
1992-09-29
Project End
1997-08-31
Budget Start
1995-09-01
Budget End
1996-08-31
Support Year
4
Fiscal Year
1995
Total Cost
Indirect Cost
Name
Case Western Reserve University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
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Zielske, Steven P; Lingas, Karen T; Li, Yan et al. (2004) Limited lentiviral transgene expression with increasing copy number in an MGMT selection model: lack of copy number selection by drug treatment. Mol Ther 9:923-31
Zielske, Steven P; Gerson, Stanton L (2003) Cytokines, including stem cell factor alone, enhance lentiviral transduction in nondividing human LTCIC and NOD/SCID repopulating cells. Mol Ther 7:325-33
Zielske, Steven P; Reese, Jane S; Lingas, Karen T et al. (2003) In vivo selection of MGMT(P140K) lentivirus-transduced human NOD/SCID repopulating cells without pretransplant irradiation conditioning. J Clin Invest 112:1561-70
Zielske, Steven P; Gerson, Stanton L (2003) SarCNU mediates selection of P140K methylguanine-DNA-methyltransferase transduced human CD34(+) cells in vitro. Blood Cells Mol Dis 31:48-50
Reese, Jane S; Liu, Lili; Gerson, Stanton L (2003) Repopulating defect of mismatch repair-deficient hematopoietic stem cells. Blood 102:1626-33
Bowman, Janice E; Reese, Jane S; Lingas, Karen T et al. (2003) Myeloablation is not required to select and maintain expression of the drug-resistance gene, mutant MGMT, in primary and secondary recipients. Mol Ther 8:42-50
Vollweiler, J L; Zielske, S P; Reese, J S et al. (2003) Hematopoietic stem cell gene therapy: progress toward therapeutic targets. Bone Marrow Transplant 32:1-7
Liu, Lili; Schwartz, Stuart; Davis, Brian M et al. (2002) Chemotherapy-induced O(6)-benzylguanine-resistant alkyltransferase mutations in mismatch-deficient colon cancer. Cancer Res 62:3070-6
Wadhwa, Punit D; Zielske, Steven P; Roth, Justin C et al. (2002) Cancer gene therapy: scientific basis. Annu Rev Med 53:437-52

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