Biochemical Changes in Carcinogen Resistant Cells

我们研究的主要焦点是调查与致癌物对癌症预防的抵抗相关的生物化学和分子变化。我们通过连续暴露于苯并[a]芘（BP），建立了一系列的抗癌细胞。这些细胞是研究早期生物化学和分子变化的模型，这些变化可能会预防环境致癌物和外源性物质引起的癌症。我们的BP抗性细胞也对DMBA具有共同抗性，DMBA是啮齿动物的乳腺癌致癌物质。据我们所知，这是第一次系统地研究致癌物耐药性的整体机制，包括致癌物的外排、活化和解毒。虽然某些解毒酶，谷胱甘肽-S-转移酶，谷胱甘肽过氧化物酶和谷胱甘肽还原酶不变，还原型谷胱甘肽水平显着增加，观察到BP耐药细胞。我们的观察表明，氧化还原介导的途径级联可能参与致癌物的耐药性。因此，我们检查了戊糖磷酸分流的主要途径酶，葡萄糖-6-磷酸脱氢酶（G6 PD）活性。我们发现BP耐药细胞的G6 PD活性明显高于WT。我们提出，致癌物抗性细胞的主要变化之一是氧化还原介导的机制。蛋白质和RNA表达的分子研究进一步支持了我们的假设。G6 PD在人乳腺癌细胞中的表达随耐药的增加而增加。我们进一步研究了饮食化学预防剂对人肝癌细胞中G6 PD的调节，发现白藜芦醇和二苯甲酰甲烷增强了G6 PD活性。这些结果提示某些化学预防剂的作用机制可能是通过氧化还原循环酶G6 PD介导的。氧化还原的变化是否在阻止乳腺或肝脏肿瘤中致癌物的发生中起重要作用，目前正在我们的研究中。

The primary focus of our study is to investigate the biochemical and molecular changes associated with carcinogen resistance toward cancer prevention. We developed a series of carcinogen-resistant cells by continuous exposure to benzo[a]pyrene (BP). These cells are a model for studying the early biochemical and molecular changes which may prevent cancer due to environmental carcinogens and xenobiotics. Our BP resistant cells are also co-resistant to DMBA, a mammary carcinogen in rodents. To our knowledge this is the first systematic examination of the overall mechanisms involved in carcinogen resistance, including carcinogen efflux, activation, and detoxification. Although certain detoxification enzymes, glutathione-s-transferase, glutathione peroxidase and glutathione reductase were unchanged, a significant increase in reduced glutathione levels was observed in BP resistant cells. Our observation suggested a redox mediated pathway cascade may be involved in carcinogen resistance. Therefore, we examined the major pathway enzyme of the pentose phosphate shunt, glucose-6-phosphate dehydrogenase (G6PD) activity. We found G6PD activities were markedly increased in BP resistant cells compared to WT. We proposed that one of the major changes in carcinogen resistant cells is in redox mediated mechanisms. Our hypothesis was further supported by molecular studies of protein and RNA expressions. The expressions of G6PD were increased with increasing resistance in human breast cancer cells. We further examined the G6PD regulation by dietary chemoprevention agents in human hepatoma cells and we found G6PD activity was enhanced by resveratrol and dibenzoylmethane. Our results suggested that the mechanism of action of certain chemoprevention agents may mediated by the redox cycle enzyme G6PD. Whether the redox changes are important in preventing carcinogen initiation in mammary or liver tumors are under our current investigation.