Assessment of Mutagenic and Carcinogenic Activity of Metabolites from Chemical Procarcinogen

化学致癌过程一般分为启动和促进两个阶段。DNA加合物的形成是靶组织被认为是启动的第一步，因此，检测加合物将表明暴露于化合物中的癌症风险。^&lt；32&gt；P-后标记分析可以检测到DNA中非常低水平的加合物，是检测癌症启动过程中这一步骤的有用工具。人和啮齿动物P450在哺乳动物细胞和酵母中的异源表达已有报道。酵母含有与哺乳动物细胞相似的胞内细胞器，可以很好地作为真核细胞的简单模型。我们建立了利用酵母表达系统检测P450介导的化学物质对DNA结合衍生物的代谢的方法，并在表达重组P450(2B5和1A1)的酵母中检测了来自MBOCA的DNA加合物的形成。这些结果表明，重组兔P4502B5对MBOCA具有潜在的激活作用，而大鼠P4501A1具有一定的作用。使用表达重组P450的酵母和^&lt；32&gt；P-后标记促进了对具有致癌潜力的化学物质的简单搜索。核糖核苷酸还原酶(RNR)催化了脱氧核糖核苷酸生产的限速步骤。DNA损伤剂可以诱导RNR3的表达。我们构建了RNR3和lac Z的5‘启动子区域的融合表达载体，并测定了β-半乳糖苷酶的活性，以检测RNR3的mRNAi的诱导。我们还在共表达大鼠CYP1A1和RNR3/LacZ的酵母中检测了RNR3的诱导和P450介导的前致癌物的激活。

The process of chemically induced carcinogenesis is generally divided into initiation and promotion. DNA adduct formation is target tissues is considered to be the first step of initiation ; therefor, detection of adducts would indicate the risks of cancer from exposure to compounds. ^<32>P-Postlabeling analysis, which can detect a very low level of adducts in DNA is a useful tool to detect this step in the initiation of cancer. Heterologous expression of human and rodent P450s in mammalian cells and yeast has been reported. Yeast contains intracellular organelles similar to mammalian cells and can be well used as a simple model of eukaryotic cells. We developed the method to measure P450-mediated metabolism of chemicals to DNA binding derivatives, using the yeast expression system, and examined the formation of DNA adducts derived from MBOCA in yeast expressing recombinant P450s (2B5 and 1A1) . These results show that recombinant rabbit P450 2B5 exhibits a potential activation of MBOCA and that rat P450 1A1 has some effect. The use of yeast expressing recombinant P450s and ^<32>P-Postlabeling facilitates a simple search for chemicals with carcinogenic potential.Ribonucleotide reductase (RNR) catalyzes the rate limiting step in the production of deoxyribonucleotides. RNR3 expression is inducible by DNA-damaging agents. We have constructed a fusion plasmid of 5'promoter region of RNR3 and lac Z,and measured beta-galactosidase activity in order to detect in-duction of RNR3 mRNA.We examined the induction of RNR3/lac Z in yeast upon exposure to various antitumor agents. We also examined RNR3 induction accompanied by P450 mediated activation of procarcinogen in yeast co-expressing rat CYP1A1 and RNR3/lacZ.