Becker K, Dosch J, Gregel C M, Martin B A, Kaina B
DNA Repair Group, Institute of Plant Genetics and Crop Plant Research, Germany.
Cancer Res. 1996 Jul 15;56(14):3244-9.
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.
致癌作用以涉及启动和促进的离散步骤进行。有充分证据表明，启动的根本原因是突变，而对于肿瘤促进，存在不同假说，假定涉及表观遗传和基因变化。DNA修复可预防肿瘤形成，但尚未证实这种保护作用是在肿瘤启动还是促进水平发生。由于研究多步骤致癌作用的最先进实验系统是小鼠皮肤，我们通过细胞角蛋白（Ck）启动子在其表皮细胞中过表达人DNA修复蛋白O6-甲基鸟嘌呤-DNA甲基转移酶（MGMT），从而生成了转基因小鼠。与非转基因小鼠相比，转基因小鼠皮肤蛋白提取物中的总细胞甲基转移酶活性显著更高。按照多阶段皮肤致癌方案对CkMGMT转基因小鼠和非转基因对照进行处理。对于启动，将单一亚阈值剂量的N-亚硝基-N-甲基脲（MNU）或7,12-二甲基苯并（a）蒽（DMBA）局部应用于小鼠背部皮肤。通过重复应用12-O-十四烷酰佛波醇-13-乙酸酯进行肿瘤促进。我们的结果清楚地表明，CkMGMT转基因小鼠对MNU引发的皮肤肿瘤形成有很强的保护作用，但对DMBA引发的无效。与非转基因对照相比，用20微摩尔或50微摩尔MNU处理后再用12-O-十四烷酰佛波醇-13-乙酸酯处理，转基因小鼠的皮肤肿瘤发生率降低了约6倍。这些结果提供了迄今为止最直接、最有说服力的证据，表明DNA损伤O6-甲基鸟嘌呤在肿瘤启动中起决定性作用，并且修复蛋白MGMT在致癌作用中的保护作用是由于预防启动而不影响肿瘤促进。
