Expression of hamster P-glycoprotein and multidrug resistance in DNA-mediated transformants of mouse LTA cells.

Abstract

The overexpression of a plasma membrane glycoprotein, P-glycoprotein, is strongly correlated with the expression of multidrug resistance. This phenotype (frequently observed in cell lines selected for resistance to a single drug) is characterized by cross resistance to many drugs, some of which are used in cancer chemotherapy. In the present study we showed that DNA-mediated transformants of mouse LTA cells with DNA from multidrug-resistant hamster cells acquired the multidrug resistance phenotype, that the transformants contained hamster P-glycoprotein DNA sequences, that these sequences were amplified whereas the recipient mouse P-glycoprotein sequences remained at wild-type levels, and that the overexpressed Pglycoprotein in these cells was of hamster origin. Furthermore, we showed that the hamster P-glycoprotein sequences were transfected independently of a group of genes that were originally coamplified and linked within a 1-megabase-pair region in the donor hamster genome. These data indicate that the high expression of P-glycoprotein is the only alteration required to mediate multidrug resistance.Cultured cells and transplantable tumors selected for resistance to a single drug can often acquire cross resistance to a wide range of drugs which differ in mode of action, target, and structure (9). This phenotype, called multidrug resistance, parallels the clinical observation that patients whose malignancies recur after primary therapy are often nonresponsive to subsequent combination chemotherapy. A central role for an integral plasma membrane glycoprotein, P-glycoprotein, in mediating multidrug resistance is supported by different observations (9). (i) Independent multidrug-resistant clones selected in our laboratory and those of others consistently overexpress P-glycoprotein (5, 9-12). (ii) Increases in the level of drug resistance are matched by increases in the amount of P-glycoprotein expressed at the cell surface (9-11). (iii) Single-step revertants to wild-type levels of drug resistance show a simultaneous decrease in the level of P-glycoprotein expression to near wild-type levels (10, 11). (iv) P-glycoprotein is highly conserved, and multidrug-resistant tissue culture mutants of hamster, mouse, and human origin all display elevated levels of P-glycoprotein (9-11). (v) The location of P-glycoprotein in the plasma membrane is consistent with a functional role for this molecule in view of the results of drug uptake studies. Such studies have shown that the phenotype is a result of a reduced net intracellular accumulation of drugs (for reviews, see references 2 and 18) and that agents which act on the membrane, such as anaesthetics, nonionic detergents, and calcium channel antagonists, alter the expression of multidrug resistance (for a review, see reference 18).Despite the close correlation between P-glycoprotein overexpression and multidrug resistance, there is no direct evidence that P-glycoprotein overexpression is the causative * Corresponding author. molecule in multidrug resistance. We have isolat...

Functional expression of murine multidrug resistance in Xenopus laevis oocytes.

“…Although mammalian P glycoproteins have been expressed in cDNA transfected cells, some of the results obtained studying those transfected cells are difficult to interpret due to the experimental protocol involved. One important point is that after the transfection step the cells must undergo at least two rounds of selection in the presence of drugs to be isolated as transfectants (24)(25)(26)(27)(28)(29). This is related to the low level of resistance shown by the transfected cells.…”

“…These and additional observations (15)(16)(17)(18)(19)(20) have led to the hypothesis that the P glycoprotein acts as an energy-dependent drug-effiux pump and functions directly as a multidrug transporter in MDR cells. In fact, cDNA and genomic transfection experiments have provided evidence indicating that increased expression of the human, hamster, and mouse P glycoprotein can mediate the MDR phenotype (21)(22)(23)(24)(25)(26)(27)(28)(29). Interestingly, these studies have shown that the transfection of a single member of the P glycoprotein gene family results in the transfer of a variable MDR phenotype to the recipient drug-sensitive cells.…”

Genetics of multidrug resistance.

“…In each case the transfer ofthe multidrug-resistant phenotype was correlated with the transfer of multiple copies of the DNA from the donor cell line, which hybridized to the cloned mdr P-glycoprotein sequences, and the overexpression of the 5-kb mRNA previously shown to be overexpressed in multidrug-resistant cell lines. Deuchars et al (28) were able to document that increased expression of P-glycoprotein was of donor origin using a species-specific MAb. Further, the P-glycoprotein sequences were transfected independently of the group of six genes previously shown to be coamplified in multidrug-resistant cell lines (21).…”

Simultaneous expression of two P-glycoprotein genes in drug-sensitive Chinese hamster ovary cells.

“…Independent transformation experiments with DNA from multidrug-resistant human or hamster cell lines consistently show transfer of only a subset of the P-glycoprotein genespecific DNA restriction fragments (5,23). This finding suggests that only a single P-glycoprotein gene family member is transferred and amplified.…”