GENETIC ANALYSIS OF THE MULTIPLE DRUG RESISTANCE PHENOTYPE IN TUMOR CELLS

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Resistance to multiple drugs is major impediment to the successful chemotherapy of human cancers. One mechanism of multidrug-resistance is the expression of a 170,000 dalton energy-dependent drug efflux pump (P-- glycoprotein or the multidrug transporter, the product of the human MDR1 gene) which confers resistance to colchicine, adriamycin, vincristine, vinblastine, puromycin, and actinomycin D. Several lines of investigation concerning the multidrug transporter have been pursued: 1) Evidence for ATP-dependent transport activity of P-glycoprotein in vesicles and across epithelial monolayers has been obtained; 2) Novel expression vectors which utilize a full-length MDR1 cDNA as a dominant selectable marker are used to introduce and amplify non-selectable genes in cultured cells and MDR1 retroviral vectors have been developed; 3) Many human tumors express MDR1 RNA, and this expression may predict drug-resistance in some cancers, and correlate with the development of drug-resistance in others; 4) Transgenic mice have been constructed in which the human MDR1 mRNA is expressed in the bone marrow at levels comparable to those found in human tumors. This level of MDR1 expression is sufficient to confer resistance to leukopenia induced by MDR drugs; 5) Increased expression of MDR1 RNA has been demonstrated in regenerating rat liver, in cultured rodent cells after exposure to chemotherapeutic agents, and in kidney cancer cells after heat shock; 6) The intron-exon structure of the human MDR1 gene has been determined and supports a model of independent evolution of the two halves of the multidrug transporter. Similarly, a deletion analysis of the MDR1 cDNA is consistent with important functions being contributed by both the amino and carboxy-terminal halves of the molecule as is a study demonstrating photoaffinity labeling of both halves of P-glycoprotein by the hydrophobic drug 3H-azidopine; and 7) Evidence that P-glycoprotein acts by pumping hydrophobic drugs out of the lipid bilayer has been obtained.