Importance of DNA repair in carcinogenesis : Insight for transgene or gene targeting experiments.

在我们的研究项目中，我们利用转基因动物研究了DNA修复系统与肿瘤发生的关系。1)我们培育了引入细菌DNA修复酶基因O^6-甲基鸟嘌呤-DNA甲基转移酶（MGMT; ada）基因的转基因小鼠（ada小鼠），ada基因在肝脏中高表达，其肝脏酶活性比非转基因小鼠高3倍。利用ada小鼠和非转基因对照小鼠，我们已经报道了ada小鼠在肝癌物质、二甲基亚硝胺或二乙基亚硝胺的作用下对肿瘤发生具有抗性。从C3H/HeN小鼠中衍生出表达ada基因的转基因小鼠，C3H小鼠株通常在晚年发生自发性肝脏肿瘤的发生率很高。在本项目中，我们比较了ada小鼠和非转基因小鼠自发性肝肿瘤的发生率。结果显示，两组发生肝脏肿瘤的发生率相同（48%）。然而，更多的组织学检查显示ada小鼠的肝细胞癌发生率明显低于非转基因小鼠。这些观察结果表明MGMT也可能保护小鼠免受肿瘤进展。2)众所周知，着色性干皮病（XP）患者存在DNA修复缺陷。大阪大学tanaka博士的团队培育的XPA （XP A组互补）基因敲除小鼠的可用性使我们能够研究XPA核苷酸切除修复在基因启动和促进体内癌变中的功能作用，使用小鼠皮肤作为模型系统。无XPA小鼠在DMBA治疗后5至7天出现皮肤溃疡。在随后的实验中，XPA阴性小鼠每隔一周重复用DMBA治疗，结果显示，在XPA阴性小鼠中，乳头状瘤首次出现在10至16周之间，发病率达到100%。杂合子和野生型小鼠的乳头瘤率几乎为零或非常低。从上述两个独立的动物模型中获得的结果提供了DNA修复保护小鼠免受化学致癌物引起的DNA损伤的直接证据。3)众所周知，p53基因缺陷小鼠在生命早期会发生自发性肿瘤。纯合子中最常见的肿瘤类型是恶性淋巴瘤；在杂合子中，以骨肉瘤和软组织肉瘤为主。在我们的项目中，p53缺陷小鼠（Aizawa et al, Kumamoto university）在妊娠12-16天接受乙基亚硝基脲移植治疗。70%的纯合子小鼠和4%的杂合子小鼠在3到4个月大时患上脑瘤。在野生小鼠中没有观察到。少

In our research project, we studied about relationship between DNA repair system and carcinogenesis using transgenic animals.1) We generated the transgenic mice (ada mouse) introduced with bacterial DNA repair enzyme gene, O^6-methylguanine-DNA methyltransferase (MGMT ; ada) gene The ada gene was highly expressed in the liver and the enzyme activity in the liver was 3 times higher in transgenic mouse than in the nontransgenic mouse.Using ada mice and nontransgenic control mice, we already reported that ada mouse was resistant to tumorigenesis when treated with liver carcinogens, dimethylnitrosamine or diethylnitrosamine. Transgenic mice expressing ada gene derived from C3H/HeN mouse, C3H mouse strain is generaly known to develop spontaneous liver tumors at high incidence in later life. In this project, we compared the incidences of spontaneous liver tumors between ada mice and nontransgenic mice. In results, both groups developed liver tumours at the same incidence (48%). However, the … More histological examination showed the incidence of hepatocellular carcinoma in ada mouse was significantly lower than in nontransgenic mouse. These obserbations suggest that MGMT may also protect mice from tumor progression.2) It is well known that Xeroderma pigmentosum (XP) patients are deficient in DNA repair. The availability of XPA (XP group A complementing) gene knockout mice generated by Dr.Tanaka's group at Osaka University has enabled us to investigate that functional role of XPA nucleotide excision repair in gene initiation and promotion of carcinogenesis in vivo, using the mouse skin as a model system. XPA null mice demonstrated skin ulcers 5 to 7 days after DMBA treatment. Subsequent experiment in which XPA null mice were repeatedly trated with DMBA at weekly intervals revealed that papillomas first arrise in XPA null mice between 10 and 16 weeks in which a 100% incidence was achieved. Heterozygous and wild-type mice showed an almost zero or very much lower yield of papillomas. Results obtained from the above two separate animal models provide direct evidence that DNA repair protects mice from DNA damage elicited by chemical carcinogens.3) It is well known that the p53 gene deficient mice develop spontaneous tumors in early life. The most frequent tumor type in homozygotes was malignant lymphoma ; in heterozytotes, osteosarcomas and soft tissue sarcomas predominated. In our project, the p53 deficient mice (generated by Aizawa et al., Kumamoto Univ.) were treated with ethylnitrosourea transplacentaly at 12-16 pregnant day. Seventy percent of homozygous and 4% of heterozyous mice developed brain tumors by the age of 3 to 4 months. None were observed in wild mice. Less