[Show abstract][Hide abstract] ABSTRACT: Tumor formation is a multi-step process that can be divided into the stages of tumor initiation, promotion and progression. Previously, we showed that overexpression in skin of mice of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT) protects against N-methyl-N-nitrosourea (MNU)-induced tumor initiation without affecting tumor promotion. This indicated that O(6)-methylguanine, which is specifically repaired by MGMT, is a major tumor-initiating lesion. Here we extended this transgenic approach to the study of tumor progression. Benign papillomas that arose on the skin of CkMGMT transgenic mice upon initiation with 7,12-dimethylbenz[a]anthracene (DMBA) and promotion by 12-O-tetradecanoylphorbol-13-acetate (TPA) expressed higher levels of MGMT than papillomas that appeared on DMBA/TPA treated non-transgenic NMRI mice. Treatment of papillomas with MNU resulted in the formation of malignant carcinomas to a significantly lower frequency in CkMGMT mice as compared with the non-transgenic control. The data provide evidence that increased DNA repair protects against the conversion of benign into malignant tumors. They show at the same time that a particular type of damage induced in DNA, namely O(6)-methylguanine, is decisively involved in triggering tumor progression. This supports the concept that the major cause of both tumor initiation and tumor progression is mutation. Data also indicate that alkylating anti-neoplastic drugs may provoke tumor progression in case of failure of tumor therapy, which is attenuated by DNA repair.
[Show abstract][Hide abstract] ABSTRACT: Transgenic systems, both cell lines and mice with gain or loss of function, are being used in order to modulate the expression of DNA repair proteins, thus allowing to assess their contribution to the defense against genotoxic mutagens and carcinogens. In this review, questions have been addressed concerning the use of transgenic systems in elucidating critical primary DNA lesions, their conversion into genotoxic endpoints, low-dose effects, and the relative contribution of individual cellular functions in defense. It has been shown that the repair protein alkyltransferase (MGMT) is decisive for protection against methylating and chloroethylating compounds. Protection pertains also to tumor formation, as revealed by the response of MGMT transgenic and knockout mice. Overexpression of genes involved in base excision repair (N-methylpurine-DNA glycosylase, apurinic endonuclease, DNA polymerase beta) is in most cases not beneficial in increasing the protection level, whereas their down-modulation or inactivation increases cellular sensitivity. This indicates that non-repaired base N-alkylation lesions and/or repair intermediates possess genotoxic potential. Modulation of mismatch repair and poly(ADP)ribosyl transferase has also been shown to affect the cellular response to alkylating agents. Furthermore, the role of Fos, Jun and p53 in cellular defense against alkylating mutagens is discussed.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 10/1998; 405(2):179-91. DOI:10.1016/S0027-5107(98)00135-3 · 3.68 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Chloroethylnitrosoureas (CNUs) are being used in the therapy of various neoplastic diseases, including skin cancer. Because secondary tumor formation is a serious threat in chemotherapy with these drugs, we explored whether and to what extent the DNA repair protein DNA-O6-methylguanine:protein-L-cysteine S-methyltransferase (MGMT) protects against CNU-induced tumors. We made use of transgenic mice overexpressing human MGMT in their skin and the initiation-promotion protocol on treatment with 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea (ACNU, nimustine) that is representative of CNUs. ACNU applied topically as a single low dose to the dorsal skin was highly effective in tumor induction in nontransgenic mice, whereas in cytokeratin MGMT transgenic mice, tumor formation was remarkably reduced. ACNU-induced skin tumors harbored mutations in the c-Ha-ras gene in both groups of mice. The results provide clear evidence that MGMT exerts protection against CNU-induced cancer. Our data also indicate that O6-chloroethylguanine, which is repaired by MGMT, is a main precarcinogenic CNU-induced DNA lesion.
Cancer Research 09/1997; 57(16):3335-8. · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Carcinogenesis proceeds in discrete steps involving initiation and promotion. There is ample evidence that the underlying cause of initiation is mutation, whereas for tumor promotion different hypotheses exist postulating the involvement of both epigenetic and genetic changes. DNA repair protects against tumor formation, but it has not been proven whether protection occurs at the level of tumor initiation or promotion. Since the most advanced experimental system for studying multistep carcinogenesis is the mouse skin, we generated transgenic mice that overexpress the human DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT) in their epidermal cells by virtue of cytokeratin (Ck) promoters. Total cellular methyltransferase activity was found to be significantly higher in skin protein extracts of transgenic as compared to nontransgenic mice. CkMGMT transgenic mice along with nontransgenic controls were treated according to the multistage skin carcinogenesis protocol. For initiation, a single subthreshold dose of N-nitroso-N-methylurea (MNU) or 7,12-dimethylbenz(a)anthracene (DMBA) was topically applied to the dorsal skin of the mice. Tumor promotion was carried out by repeated 12-O-tetradecanoylphorbol-13-acetate application. Our results clearly show that CkMGMT transgenic mice are strongly protected against MNU- but not DMBA-initiated skin tumor formation. As compared to nontransgenic controls, transgenic mice exhibited an approximately 6-fold reduction of skin tumor incidence after treatment with 20 micromol or 50 micromol MNU followed by 12-O-tetradecanoylphorbol-13-acetate. These results provide direct and the most compelling evidence to date that the DNA lesion O6-methylguanine is of decisive importance in tumor initiation, and that the protective effect of the repair protein MGMT in carcinogenesis is due to prevention of initiation without affecting tumor promotion.
Cancer Research 08/1996; 56(14):3244-9. · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: CMoroethylnitrosoureas (CNUs) are being used in the therapy of var ions neoplasticdiseases, includingskin cancer. Becausesecondarytumor formation is a serious threat in chemotherapy with these drugs, we explored whether and to what extent the DNA repair protein DNA-06- methylguanine:protein-L-cysteine S-methyltransferase (MGMT) protects against CNU-induced tumors. We made use of transgenic mice overex pressing human MGMT in their skin and the initiation-promotion proto col on treatment with 1-(4-amino-2-methyl-5-pyrimidlnyl)methyl-3-(2- chloroethyl)-3-nitrosourea (ACNU, nimustine) that is representative of CNUs. ACNU applied topically as a single low dose to the dorsal skin was highly effective in tumor induction in nontransgenic mice, whereas in cylokeratin MGMT transgenic mice, tumor formation was remarkably reduced. ACNU-inducedskin tumors harbored mutations in the c-Ha-ras gene in both groups of mice. The results provide clear evidence that MGMT exerts protection against CNU-inducedcancer. Our data also indicate that O@-chloroethylguanine, which is repaired by MGMT, is a main precarcinogemcCNU-inducedDNA lesion.