Multidrug-resistant human sarcoma cells with a mutant P-glycoprotein, altered phenotype, and resistance to cyclosporins
Multidrug-resistant human sarcoma cells with a mutant P-glycoprotein, altered phenotype, and resistance to cyclosporins
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DOI：
10.1074/jbc.272.9.5974
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发表时间：
1997-02-28
影响因子：
4.8
通讯作者：
Sikic, BI
中科院分区：
文献类型：
--
作者：
Chen, G;Duran, GE;Sikic, BI
多重耐药人肉瘤细胞系 Dx5 的变体是通过与多柔比星和环孢菌素 D 类似物 PSC 833（多药转运蛋白 P-糖蛋白的有效抑制剂）共选择而衍生的。与 Dx5 相比，变体 DxP 细胞表现出改变的表型，对长春花生物碱的交叉耐药性降低，对放线菌素无耐药性。对阿霉素和紫杉醇的耐药性得以保留。 DxP 细胞的多药耐药表型不受 2 μM PSC 833 或环孢菌素调节，DxP 细胞表现出转运[H-3]环孢菌素的能力下降。 DNA 异源双链体分析和测序揭示了 DxP 细胞系中存在突变的 mdr1 基因（氨基酸残基 335 处的苯丙氨酸缺失）。突变型 P-糖蛋白对 PSC 833 和长春花碱的亲和力降低，转运罗丹明 123 的能力降低。将突变型 mdr1 基因转染到药物敏感的 MES-SA 肉瘤细胞中，可赋予对阿霉素和 PSC 833 的抗性。我们的研究表明，在多步选择中暴露于阿霉素和 PSC 833 的细胞的存活是由于 P-糖蛋白突变造成的。跨膜区 6。这些数据表明，Phe(335) 是 P-糖蛋白上放线菌素和长春碱等底物以及环孢菌素和 PSC 833 等抑制剂的重要结合位点。
A variant of the multidrug-resistant human sarcoma cell line Dx5 was derived by co-selection with doxorubicin and the cyclosporin D analogue PSC 833, a potent inhibitor of the multidrug transporter P-glycoprotein, The variant DxP cells manifest an altered phenotype compared with Dx5, with decreased cross-resistance to Vinca alkaloids and no resistance to dactinomycin. Resistance to doxorubicin and paclitaxel is retained. The multidrug resistance phenotype of DxP cells is not modulated by 2 mu M PSC 833 or cyclosporine, DxP cells manifest a decreased ability to transport [H-3]cyclosporine. DNA heteroduplex analysis and sequencing reveal a mutant mdr1 gene (deletion of a phenylalanine at amino acid residue 335) in the DxP cell line. The mutant P-glycoprotein has a decreased affinity for PSC 833 and vinblastine and a decreased ability to transport rhodamine 123, Transfection of the mutant mdr1 gene into drug-sensitive MES-SA sarcoma cells confers resistance to both doxorubicin and PSC 833. Our study demonstrates that survival of cells exposed to doxorubicin and PSC 833 in a multistep selection occurred as a result of a P-glycoprotein mutation in transmembrane region 6. These data suggest that Phe(335) is an important binding site on P-glycoprotein for substrates such as dactinomycin and vinblastine and for inhibitors such as cyclosporine and PSC 833.