The mechanism of regulation of P-glycoprotein (Analysis of multidrug-resistant mutants that do not express P-glycoprotein)

1)我们分离到了对高水平阿霉素、C-A90、C-A120、C-A500和C-A1000耐药的人表皮样癌KB细胞系。在含有阿霉素、多药耐药逆转剂(MDR)逆转剂1µg/ml和蛋白激动素C激活剂100 nm mezerein的筛选培养基中进行分离。与亲本KB-3-1细胞相比，耐阿霉素KB细胞株C-A500对典型的多药耐药表型药物如长春新碱、放线菌素D、VP-16和秋水仙碱具有交叉耐药性。多药耐药相关蛋白和多药耐药相关蛋白在耐阿霉素的KB细胞、C-A120、C-A500和C-A1000中表达，而在亲本KB-3-1和REVE…中不表达更多变异的C-AR细胞。4)DNA拓扑异构酶II在C-A500和C-A1000细胞中显著降低，而在C-A120细胞中仅略有降低。这些结果表明，MRP在耐药细胞中的过度表达可能是导致阿霉素和长春新碱积聚减少的原因，而在C-A120、C-A500和C-A1000细胞中，MRP表达的增加和拓扑异构酶II水平的降低都是耐药的基础。5)我们研究了多药耐药KB细胞株(KB-8-5、KB-C2、C-A40和C-A120)、人非小细胞肺癌(NSCLC)、胃癌和结直肠癌中MRP mRNA的表达，并与药物敏感的人KB细胞进行了比较。9例肺鳞癌中有8例MRP基因表达下调(89%)。此外，4种鳞癌(L13、18、19和20)中MRP的表达水平是KB-3-1细胞的3.6倍以上，且所有鳞癌的平均MRP mRNA表达水平均显著高于肺腺癌、结直肠癌和胃癌。较少

1) We have isolated human epidermoid KB cell lines resistant to high levels of adriamycin, C-A90, C-A120, C-A500 and C-A1000. They were isolated in selection medium containing increasing concentrations of adriamycin, 1mug/ml of cepharanthine, a multidrug-resistance (MDR) reversing agent, and 100nM mezerein, a protein kinas C activating agent. One of the adriamycin-resistant KB cell lines, C-A500, was cross-resistant to drugs that typify the classical multidrug resistance phenotype, such as vincristine, actinomycin D,VP-16 and colchicine.2) The accumulation of adriamycine and vincristine was decreased in C-A500 cells and the efflux of adriamycine from C-A 500 was enhanced compared with parental KB-3-1 cells.These adriamycin-resistant KB cells did not contain detectable levels of P-glycoprotein or overexpress MDR1. Multidrug-resistance-associated protein (MRP) and MRP mRNA were expressed in the adriamycin-resistant KB-cells, C-A120, C-A500 and C-A1000, but not in parental KB-3-1 and reve … More rtant C-AR cells. The MRP gene was amplified in these cells that overexpressed MRP mRNA.4) DNA topoisomerase II levels were markedly decreased in C-A500 and C-A1000 cells but only slightly decreased in C-A120 cells. These results indicate that MRP overexpressed in the resistant cells may be responsible for the reduced accumulation of adriamycin and vincrisrine and that both the increased expression of MRP and decreasd levels of topoisomerase II underlie te drug resistance in C-A120, C-A500 and C-A1000 cell lines.5) We investigated the expression of MRP mRNA in multidrug-resistant KB cell line (KB-8-5, KB-C2, C-A40 and C-A120), human non-small-cell lung carcinomas (NSCLC), gastric and colorectal carcinomas and compared it with that in drug-sensitive human KB cells. MRP gene expression was elavated in 8 of 9 (89%) squamous-cell carcinomas of the lung. Furthermore, MRP expression in 4 squamous-cell carcinomas (L13,18,19 and 20) was more than 3.6 times higher than in KB-3-1 cells, and the average MRP mRNA expression level of all squamous-cell carcinomas was significantly higher than that of adenocarcinoma of the lung and of colorectal and gastric carcinomas. Less