Horio M, Gottesman M M, Pastan I
Laboratory of Molecular Biology, National Cancer Institute, Bethesda, MD 20892.
Proc Natl Acad Sci U S A. 1988 May;85(10):3580-4. doi: 10.1073/pnas.85.10.3580.
Resistance of human cancer cells to multiple cytotoxic hydrophobic agents (multidrug resistance) is due to overexpression of the "MDR1" gene, whose product is the plasma membrane P-glycoprotein. Plasma membrane vesicles partially purified from multidrug-resistant human KB carcinoma cells, but not from drug-sensitive cells, accumulate [3H]vinblastine in an ATP-dependent manner. This transport is osmotically sensitive, with an apparent Km of 38 microM for ATP and of approximately equal to 2 microM for vinblastine. The nonhydrolyzable analog adenosine 5'-[beta, gamma-imido]triphosphate does not substitute for ATP but is a competitive inhibitor of ATP for the transport process. Vanadate, an ATPase inhibitor, is a potent noncompetitive inhibitor of transport. These results indicate that hydrolysis of ATP is probably required for active transport of vinblastine. Several other drugs to which multidrug-resistant cell lines are resistant inhibit transport, with relative potencies as follows: vincristine greater than actinomycin D greater than daunomycin greater than colchicine = puromycin. Verapamil and quinidine, which reverse the multidrug-resistance phenotype, are good inhibitors of the transport process. These results confirm that multidrug-resistant cells express an energy-dependent plasma membrane transporter for hydrophobic drugs, and establish a system for the detailed biochemical analysis of this transport process.
人类癌细胞对多种细胞毒性疏水药物的耐药性（多药耐药）是由于“MDR1”基因的过度表达，其产物是质膜P - 糖蛋白。从多药耐药的人KB癌细胞中部分纯化得到的质膜囊泡，但不是从药物敏感细胞中得到的，以ATP依赖的方式积累[3H]长春碱。这种转运对渗透压敏感，对ATP的表观Km为38 microM，对长春碱约为2 microM。不可水解的类似物腺苷5'-[β，γ - 亚氨基]三磷酸不能替代ATP，但在转运过程中是ATP的竞争性抑制剂。钒酸盐，一种ATP酶抑制剂，是转运的有效非竞争性抑制剂。这些结果表明，长春碱的主动转运可能需要ATP的水解。多药耐药细胞系耐药的其他几种药物抑制转运，相对效力如下：长春新碱大于放线菌素D大于柔红霉素大于秋水仙碱 = 嘌呤霉素。维拉帕米和奎尼丁可逆转多药耐药表型，是转运过程的良好抑制剂。这些结果证实多药耐药细胞表达一种能量依赖的疏水药物质膜转运体，并建立了一个对该转运过程进行详细生化分析的系统。
