Bligh H F, Bartoszek A, Robson C N, Hickson I D, Kasper C B, Beggs J D, Wolf C R
Department of Molecular Biology, University of Edinburgh, United Kingdom.
Cancer Res. 1990 Dec 15;50(24):7789-92.
Mitomycin C is an alkylating agent used in cancer chemotherapy that shows some specificity towards hypoxic cells. The therapeutic effects of this compound are thought to result from its metabolic activation by enzymes such as NADPH:cytochrome P-450 reductase. In a previous report we described a Chinese hamster ovary cell line resistant to mitomycin C, which had a decreased NADPH:cytochrome P-450 reductase activity coupled with a lower rate of mitomycin C metabolism. In order to provide further evidence that the lower reductase activity is a factor in the resistance mechanism, we incorporated NADPH:cytochrome P-450 reductase into cytotoxicity assays and showed that it significantly sensitizes cells to mitomycin C. Also, the difference in drug sensitivity between the wild-type and drug-resistant Chinese hamster ovary cells was no longer observed. In addition to these studies, we expressed a rat liver NADPH:cytochrome P-450 reductase cDNA in a Salmonella typhimurium strain, LR5000. The bacteria expressing the rat NADPH: cytochrome P-450 reductase showed increased sensitivity to mitomycin C when incubated with this compound under aerobic conditions. However, under hypoxic conditions increased sensitivity was not observed. This parallels the previous finding with mitomycin C-resistant Chinese hamster ovary cells. These data provide direct evidence for the role of NADPH:cytochrome P-450 reductase in the cytotoxic action of this mitomycin C under aerobic but not hypoxic conditions and suggest that reduced levels of this enzyme can lead to drug resistance. P-450 reductase expressed in S. typhimurium may provide a valuable tool for evaluating the role of this enzyme in the toxicity of drugs activated through a one electron reduction pathway.
丝裂霉素C是一种用于癌症化疗的烷化剂，对缺氧细胞具有一定的特异性。该化合物的治疗效果被认为是由诸如NADPH：细胞色素P - 450还原酶等酶对其进行代谢激活所致。在之前的一份报告中，我们描述了一种对丝裂霉素C耐药的中国仓鼠卵巢细胞系，该细胞系的NADPH：细胞色素P - 450还原酶活性降低，同时丝裂霉素C的代谢速率也较低。为了进一步证明较低的还原酶活性是耐药机制中的一个因素，我们将NADPH：细胞色素P - 450还原酶纳入细胞毒性试验，结果表明它能显著使细胞对丝裂霉素C敏感。此外，野生型和耐药型中国仓鼠卵巢细胞之间的药物敏感性差异也不再明显。除了这些研究，我们还在鼠伤寒沙门氏菌菌株LR5000中表达了大鼠肝脏NADPH：细胞色素P - 450还原酶的cDNA。在有氧条件下与丝裂霉素C一起孵育时，表达大鼠NADPH：细胞色素P - 450还原酶的细菌对丝裂霉素C的敏感性增加。然而，在缺氧条件下未观察到敏感性增加。这与之前对丝裂霉素C耐药的中国仓鼠卵巢细胞的研究结果相似。这些数据为NADPH：细胞色素P - 450还原酶在有氧而非缺氧条件下对丝裂霉素C的细胞毒性作用中的作用提供了直接证据，并表明该酶水平降低可导致耐药性。在鼠伤寒沙门氏菌中表达的P - 450还原酶可能为评估该酶在通过单电子还原途径激活的药物毒性中的作用提供一个有价值的工具。
