MULTIDRUG-RESISTANT HUMAN SARCOMA-CELLS WITH A MUTANT P-GLYCOPROTEIN,ALTERED PHENOTYPE, AND RESISTANCE TO CYCLOSPORINES

A variant of the multidrug-resistant human sarcoma cell line Dx5 was d erived by co-selection with doxorubicin and the cyclosporin D analogue PSC 833, a potent inhibitor of the multidrug transporter P-glycoprote in, The variant DxP cells manifest an altered phenotype compared with Dx5, with decreased cross-resistance to Vinca alkaloids and no resista nce to dactinomycin. Resistance to doxorubicin and paclitaxel is retai ned. The multidrug resistance phenotype of DxP cells is not modulated by 2 mu M PSC 833 or cyclosporine, DxP cells manifest a decreased abil ity to transport [H-3]cyclosporine. DNA heteroduplex analysis and sequ encing 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 abil ity to transport rhodamine 123, Transfection of the mutant mdr1 gene i nto drug-sensitive MES-SA sarcoma cells confers resistance to both dox orubicin and PSC 833. Our study demonstrates that survival of cells ex posed 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-glycopro tein for substrates such as dactinomycin and vinblastine and for inhib itors such as cyclosporine and PSC 833.