Massaad L, de Waziers I, Ribrag V, Janot F, Beaune P H, Morizet J, Gouyette A, Chabot G G
Laboratoire de Pharmacologie Clinique, Institut Gustave-Roussy, (INSERM U 140 and CNRS URA 147), Villejuif, France.
Cancer Res. 1992 Dec 1;52(23):6567-75.
Since human colorectal tumors are insensitive to most chemotherapeutic agents, there is a need for the discovery of new drugs that would show activity against this disease. In an attempt to better appreciate the relevance of a widely used mouse colon tumor (colon adenocarcinoma Co38) as a screening model for human colorectal tumors, we compared the main phase I and phase II drug-metabolizing enzyme systems in both tumoral and nontumoral colon tissues. The following enzymes were assayed by Western blot: cytochromes P-450 (1A1/A2, 2B1/B2, 2C, 2E1, and 3A), epoxide hydrolase, and glutathione-S-transferases (GST-alpha, -mu, and -pi). The activities of the following enzymes or cofactors were determined by spectrophotometric or fluorometric assays: total cytochrome P-450, 1-chloro-2,4-dinitrobenzene-GST, selenium-independent glutathione peroxidase, 3,4-dichloronitrobenzene-GST, ethacrynic acid-GST, total glutathione, epoxide hydrolase, UDP-glucuronosyltransferase, beta-glucuronidase, sulfotransferase, and sulfatase. Results obtained by Western blot showed that mouse colon adenocarcinoma Co38 did not express any of the probed cytochromes P-450, whereas human colorectal tumors expressed only low levels of cytochrome P-450 3A. GST-alpha and GST-pi were detected in all tumoral and nontumoral tissues of both species. The neutral GST-mu was expressed in all murine tissues investigated and was found to be polymorphic in human tissues. For human peritumoral and tumoral colorectal tissues there was no significant difference between GST isoenzyme levels, whereas mouse colon adenocarcinoma Co38 had a lower expression of GST-mu and GST-pi, compared to normal mouse colon. Enzymatic activities for glutathione peroxidase, 3,4-dichloronitrobenzene-GST, and ethacrynic acid-GST confirmed the Western blot results for GST-alpha, GST-mu, and GST-pi, respectively. Total GSH levels were similar between murine and human tumors but were 3-fold higher in human tumors than in peritumoral tissues, whereas they were 7-fold lower in mouse colon tumor Co38, compared to normal mouse colon. Epoxide hydrolase was not expressed in either mouse colon adenocarcinoma Co38 or normal mouse colon tissues, whereas it was expressed in human colon peritumoral and tumoral tissues at similar levels. No significant difference was observed between human tumors and peritumoral tissues for UDP-glucuronosyltransferase, beta-glucuronidase, sulfotransferase, and sulfatase. For murine colon tissues, the conjugation pathways (UDP-glucuronosyltransferase and sulfotransferase) were lower in colon adenocarcinoma Co38, whereas the converse was observed for the corresponding hydrolytic enzymes (beta-glucuronidase and sulfatase).(ABSTRACT TRUNCATED AT 400 WORDS)
由于人类结肠肿瘤对大多数化疗药物不敏感，因此需要发现对这种疾病有活性的新药。为了更好地理解广泛使用的小鼠结肠肿瘤（结肠腺癌Co38）作为人类结肠肿瘤筛选模型的相关性，我们比较了肿瘤和非肿瘤结肠组织中主要的I期和II期药物代谢酶系统。通过蛋白质印迹法检测了以下酶：细胞色素P-450（1A1/A2、2B1/B2、2C、2E1和3A）、环氧化物水解酶和谷胱甘肽-S-转移酶（GST-α、-μ和-π）。通过分光光度法或荧光法测定了以下酶或辅因子的活性：总细胞色素P-450、1-氯-2,4-二硝基苯-GST、硒非依赖性谷胱甘肽过氧化物酶、3,4-二氯硝基苯-GST、依他尼酸-GST、总谷胱甘肽、环氧化物水解酶、UDP-葡萄糖醛酸基转移酶、β-葡萄糖醛酸酶、磺基转移酶和硫酸酯酶。蛋白质印迹法得到的结果显示，小鼠结肠腺癌Co38不表达任何检测的细胞色素P-450，而人类结肠肿瘤仅低水平表达细胞色素P-450 3A。在两个物种的所有肿瘤和非肿瘤组织中均检测到GST-α和GST-π。中性GST-μ在所有研究的小鼠组织中均有表达，且在人类组织中具有多态性。对于人类肿瘤周围和肿瘤性结肠组织，GST同工酶水平无显著差异，而与正常小鼠结肠相比，小鼠结肠腺癌Co38中GST-μ和GST-π的表达较低。谷胱甘肽过氧化物酶、3,4-二氯硝基苯-GST和依他尼酸-GST的酶活性分别证实了GST-α、GST-μ和GST-π的蛋白质印迹结果。小鼠和人类肿瘤中的总谷胱甘肽水平相似，但人类肿瘤中的总谷胱甘肽水平比肿瘤周围组织高3倍，而与正常小鼠结肠相比，小鼠结肠肿瘤Co38中的总谷胱甘肽水平低7倍。环氧化物水解酶在小鼠结肠腺癌Co38或正常小鼠结肠组织中均未表达，而在人类结肠肿瘤周围和肿瘤组织中以相似水平表达。人类肿瘤和肿瘤周围组织在UDP-葡萄糖醛酸基转移酶、β-葡萄糖醛酸酶、磺基转移酶和硫酸酯酶方面未观察到显著差异。对于小鼠结肠组织，结合途径（UDP-葡萄糖醛酸基转移酶和磺基转移酶）在结肠腺癌Co38中较低，而相应的水解酶（β-葡萄糖醛酸酶和硫酸酯酶）则相反。（摘要截于400字）
