LYMPHOMAGENESIS OF O6-METHYLGUANINE

内源性治疗剂和环境甲基化剂形成O 6- 甲基鸟嘌呤[06 mG] DNA加合物，其被认为是人类 致癌物质。 在上一个资助期内， 研究人员已经证明了O 6 mG DNA加合物在 在用MNU处理的小鼠中诱导淋巴瘤。 转基因表达 DNA修复蛋白O 6-烷基鸟嘌呤DNA烷基转移酶， 去除O 6 mG DNA加合物显著降低淋巴瘤发生的风险， 表明O 6 mG是MNU形成的主要致癌加合物。 在这个提议中，O 6 mG DNA加合物在致癌作用中的作用将 通过研究错配修复系统的参与来重新评估 在致癌过程中。 O 6-mG：T的错配修复识别 错配导致单链断裂，染色体重排， 畸变 然而，以前，O 6-mG的致癌作用一直是 认为这完全是由于G到A的点突变，因为O 6-mG 在DNA合成过程中优先与胸腺嘧啶错配。 的 待检验的假设是在有错配修复能力的小鼠中， 染色体重排在致癌作用中是重要的， 错配修复缺陷小鼠，致癌作用先于G， A点突变。 在第一个特定目标中，转基因小鼠 两种错配修复蛋白之一的缺陷，PMS 2或MSH 2将被 用MNU处理并随后诱导肿瘤。自从一个标志性的 错配修复突变的原因是缺乏甲基化的细胞毒性， 药物，更多的细胞将存活与持久的O 6-mG DNA加合物/ 导致G到A的点突变。因此， MNU诱导的肿瘤是预期的。这些肿瘤将被分析， 确定失配修复缺陷是否影响所述类型， 肿瘤的发病率，染色体畸变和G- K-ras基因的点突变。 在特定目标2中，RAG 2的MNU治疗 将使用敲除小鼠。 RAG 2小鼠不能经历 V（D）J关节形成，不能产生成熟的T细胞， 重排的T细胞受体 然而，令人惊讶的是， 表明RAG 2小鼠对MNU诱导的淋巴瘤易感 这些细胞含有T细胞受体重排。因此，具体目标 2将定义MNU诱导T细胞受体的能力 RAG 2敲除小鼠中的重排导致淋巴瘤发生。 的 研究人员提出，O 6 mG可以绕过RAG 2缺陷， 加合物介导的错配修复系统的活化导致 T细胞受体基因座的染色体重排。 整体 这些研究的目的是了解复杂的致癌物质 甲基化剂的途径和DNA修复对 过程

Endogenous therapeutic and environmental methylating agents form O6- methylguanine [06mG] DNA adducts, which have been implicated as human carcinogens. During the previous funding period for this grant, the investigators have shown the importance of O6mG DNA adducts in the induction of lymphoma in mice treated with MNU. Transgenic expression of the DNA repair protein O6-alkylguanine DNA alkyltransferase which removes O6mG DNA adducts markedly reduce the risk of lymphomagenesis, indicating the O6mG is the dominant carcinogenic adduct formed by MNU. In this proposal, the role of O6mG DNA adducts in carcinogenesis will be reassessed by studying the involvement of the mismatch repair system in the carcinogenic process. Mismatch repair recognition of O6-mG:T mispairs leads to single strand breaks, chromosomal rearrangements and aberrations. Previously, however, carcinogenesis of O6-mG has been thought to be entirely due to G to A point mutations because the O6-mG preferentially mispairs with thymine during DNA synthesis. The hypothesis to be tested is that in mismatch repair competent mice, chromosomal rearrangements are important in carcinogenesis whereas in mismatch repair defective mice, carcinogenesis precedes through G to A point mutations. In the first Specific Aim, transgenic mice defective in one of two mismatch repair proteins, PMS2 or MSH2 will be treated with MNU and followed for induction of tumors. Since a hallmark of mismatch repair mutation is the lack of cytotoxicity of methylating agents, more cells will survive with persistent O6-mG DNA adducts/ resulting in G to A point mutations. Thus, an increased incidence of MNU induced tumors is expected. These tumors will be analyzed to determine whether mismatch repair defects influence the type and incidence of tumors, the presence of chromosomal aberrations and G to A point mutations in K-ras. In Specific Aim 2, MNU treatment of RAG2 knockout mice will be utilized. RAG2 mice are incapable of undergoing V(D)J joint formation and are unable to produce mature T-cells with rearranged T-cell receptors. Surprisingly, however, preliminary data indicates that RAG2 mice are susceptible to MNU induction of lymphomas and these contain T-cell receptor rearrangements. Thus, Specific Aim 2 will define the ability of MNU to induce T-cell receptor rearrangements in RAG2 knockout mice leading to lymphomagenesis. The investigators propose that the RAG2 defect can be bypassed by O6mG adduct mediated activation of the mismatch repair system leading to chromosomal rearrangements at the T-cell receptor locus. The overall goal of these studies is to understand the complex carcinogenic pathways of methylating agents and the impact of DNA repair on the process.