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

在本研究项目中，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)建立了导入细菌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)，该基因在肝脏中高表达，其肝脏中的酶活性是非转基因小鼠的3倍。表达ADA基因的转基因小鼠来源于C3H/HeN小鼠，C3H小鼠品系在晚年发生自发性肝肿瘤是众所周知的高发疾病。在这个项目中，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，两组发生肝肿瘤的发生率相同(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。这些观察表明，MGMT也可以保护小鼠免受肿瘤的侵袭。2)众所周知，着色性干皮病(XP)患者的DNA修复能力不足。大阪大学Tanaka博士团队建立的XPA(XP Group A互补性)基因敲除小鼠的出现，使我们能够以小鼠皮肤为模型系统，研究XPA核苷酸切除修复在体内基因启动和促进癌变中的功能作用。XPA基因缺失的小鼠在DMBA治疗后5-7天出现皮肤溃疡。在随后的实验中，XPA基因缺失的小鼠每周一次重复使用DMBA治疗，发现乳头状瘤首先出现在XPA基因缺失的小鼠10到16周，在这段时间里，发病率达到了100%。杂合子和野生型小鼠的乳头状瘤发生率几乎为零或非常低。上述两种不同动物模型的结果为DNA修复保护小鼠免受化学致癌物引起的DNA损伤提供了直接证据。3)众所周知，p53基因缺陷小鼠在生命早期就会发生自发性肿瘤。纯合子中最常见的肿瘤类型是恶性淋巴瘤，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(由熊本大学Aizawa等人产生)妊娠12~16天经胎盘给予乙基亚硝脲治疗。70%的纯合子小鼠和4%的杂合小鼠在3到4个月大的时候患上了脑瘤。在野鼠身上没有观察到这种情况。较少

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