Repair of alkylation DNA damage

利用基因打靶技术获得编码O^6-甲基鸟嘌呤-DNA甲基转移酶的MGMT基因缺陷小鼠。这些MGMT^+小鼠对烷基化致癌物甲基亚硝脲(MNU)最为敏感；6周龄小鼠注射不同剂量的MNU并测定第30天的存活率时，MGMT^+和MGMT^+&gt；小鼠的LD_&lt；50s&gt；体重分别为20和240 mg/kg体重。MGMT^-lt；-/-&gt；小鼠暴露于2.5 mg/kg体重的MNU后，出现了大量的胸腺淋巴瘤和肺腺瘤。在给予相同剂量药物的情况下，MGMT^&lt；+/+&gt；和MGMT^&lt；+/-&gt；小鼠无或极少发生肿瘤。DNA修复甲基转移酶蛋白似乎保护了这些小鼠免受MNU诱导的肿瘤的发生。为了研究小鼠组织DNA中产生的O^6-甲基鸟嘌呤的命运，我们使用了埃森大学Rajeski研究小组提出的单抗。该抗体制剂可识别O^6-乙基鸟嘌呤和O^6-甲基鸟嘌呤。为了合作，九州大学的Hayakawa和K.Sakumi参观了埃森实验室，埃森的T.Schweer访问了福冈。我们已经确定了给予MGMT^&lt；+/+&gt；和MGMT^&lt；-/-&gt；小鼠适当剂量的MNU，以及跟踪小鼠组织中甲基化碱基量变化的条件和程序。初步结果表明，MGMT^&lt；-/-&gt；小鼠在DNA中保留了大量的烷化碱基，而MGMT^&lt；+/+&gt；小鼠失去这些碱基的速度相当快。

Gene targeting was used to obtain mice defective in the MGMT gene, encoding O^6 -methylguanine-DNA methyltransferase. These MGMT^+ mice were most sensitive to the alkylating carcinogen, methylnitrosourea (MNU) ; when varied doses of MNU were administered to 6-week-old mice and survivals at the 30th day were determined, LD_<50S> of MGMT^+ and MGMT^<+/+> mice were 20 and 240 mg/kg of body weight, respectively.MGMT^<+/-> mice were as resistant as MGMT^<+/+> mice, but some difference in survival time was noted when the two genotypes of mice were exposed to a relatively high dose of MNU.A large number of thymic lymphomas, as well as lung adenomas, occurred in MGMT^<-/-> mice exposed to MNU at a dose of 2.5 mg/kg of body weight. In case of exposure to the same dose of drub, no or few tumors occureed in the MGMT^<+/+> and MGMT^<+/-> mice. It appears that the DNA repair methyltransferase protein protected these mice from MNU-induced tumorigenesis.To pursue the fate ot O^6-methylguanin produced in the DNA of mouse tissues, we used monoclonal antibodies raised by the Rajewsky's group of University of Essen. This antibody preparation specifically recognizes O^6-ethylguanine as well as O^6-methylguanin. For the collaborative work, H.Hayakawa and K.Sakumi of Kyushu University visited the Essen laboratory and T.Schweer of Essen visited here in Fukuoka. We have established appropriate doses of MNU to be given to MGMT^<+/+> and MGMT^<-/-> mice and also conditions and procedures to follow changes in amounts of methylated bases in mouse tissues. A preliminary result indicates that MGMT^<-/-> mice retain a significant level of alkylated bases in DNA whereas MGMT^<+/+> mice lose these bases rather quickly.