Safa A R, Stern R K, Choi K, Agresti M, Tamai I, Mehta N D, Roninson I B
Department of Medicine, University of Chicago, IL 60637.
Proc Natl Acad Sci U S A. 1990 Sep;87(18):7225-9. doi: 10.1073/pnas.87.18.7225.
Expression of P-glycoprotein, encoded by the human MDR1 gene, results in cross-resistance to many lipophilic cytotoxic drugs (multidrug resistance). P-glycoprotein is believed to function as an energy-dependent efflux pump that is responsible for decreased drug accumulation in multidrug-resistant cells. Previous work showed that preferential resistance to colchicine in a colchicine-selected multidrug-resistant cell line was caused by spontaneous mutations in the MDR1 gene that resulted in a Gly-185----Val-185 substitution in P-glycoprotein. We have now compared transfectant cell lines expressing either the wild-type Gly-185 or the mutant Val-185 P-glycoprotein with regard to their levels of resistance to and accumulation and binding of different drugs. In cells expressing the mutant protein, increased resistance to colchicine and decreased resistance to vinblastine correlated with a decreased accumulation of colchicine and increased accumulation of vinblastine. Expression of the mutant P-glycoprotein also resulted in significantly increased resistance to epipodophyllotoxin and decreased resistance to vincristine and actinomycin D; smaller changes in resistance were observed for several other drugs. Unexpectedly, the mutant P-glycoprotein showed increased binding of photoactive analogs of vinblastine and verapamil and the photoactive compound azidopine and decreased binding of a photoactive colchicine analog. These results suggest that the Gly-185----Val-185 substitution affects not the initial drug-binding site of P-glycoprotein but another site, associated with the release of P-glycoprotein-bound drugs to the outside of the cell.
由人类MDR1基因编码的P-糖蛋白的表达导致对许多亲脂性细胞毒性药物产生交叉耐药性（多药耐药）。P-糖蛋白被认为作为一种能量依赖性外排泵，负责多药耐药细胞中药物积累的减少。先前的研究表明，在秋水仙碱选择的多药耐药细胞系中，对秋水仙碱的优先耐药性是由MDR1基因的自发突变引起的，该突变导致P-糖蛋白中第185位甘氨酸被缬氨酸取代。我们现在比较了表达野生型甘氨酸-185或突变型缬氨酸-185 P-糖蛋白的转染细胞系对不同药物的耐药水平、积累和结合情况。在表达突变蛋白的细胞中，对秋水仙碱耐药性增加而对长春碱耐药性降低，这与秋水仙碱积累减少和长春碱积累增加相关。突变型P-糖蛋白的表达还导致对表鬼臼毒素的耐药性显著增加，而对长春新碱和放线菌素D的耐药性降低；对其他几种药物观察到较小的耐药性变化。出乎意料的是，突变型P-糖蛋白显示对长春碱和维拉帕米的光活性类似物以及光活性化合物叠氮平的结合增加，而对光活性秋水仙碱类似物的结合减少。这些结果表明，甘氨酸-185至缬氨酸-185的取代并不影响P-糖蛋白的初始药物结合位点，而是影响另一个位点，该位点与将P-糖蛋白结合的药物释放到细胞外有关。
