Differential effects of mitomycin C and doxorubicin on P-glycoprotein expression

Abstract

Previous studies have demonstrated that mitomycin C (MMC) and other DNA cross-linking agents can suppress MDR1 (multidrug resistance 1) gene expression and subsequent functional P-glycoprotein (Pgp) expression, whereas doxorubicin and other anthracyclines increase MDR1 gene expression. In the present study, with stably transfected Madin–Darby canine kidney C7 epithelial cells expressing a human Pgp tagged with green fluorescent protein under the proximal human MDR1 gene promoter, we demonstrated that MMC and doxorubicin have differential effects on Pgp expression and function. Doxorubicin caused a progressive increase in the cell-surface expression of Pgp and function. In contrast, MMC initially increased plasma membrane expression and function at a time when total cellular Pgp was constant and Pgp mRNA expression had been shown to be suppressed. This was followed by a rapid and sustained decrease in cell-surface expression at later times, presumably as a consequence of the initial decrease in mRNA expression. These studies imply that there are at least two independent chemosensitive steps that can alter Pgp biogenesis: one at the level of mRNA transcription and the other at the level of Pgp trafficking. Understanding the combined consequences of these two mechanisms might lead to novel chemotherapeutic approaches to overcoming drug resistance in human cancers by altering either Pgp mRNA expression or trafficking to the membrane.

Chemotoxicity of doxorubicin and surface expression of P-glycoprotein (MDR1) is regulated by the Pseudomonas aeruginosa toxin Cif

“…These observations are consistent with the fact that these two cell lines have equal amounts of MRP2 in the apical plasma membrane (Fig. 8A) and the observation that Pgp does not transport cisplatin (16,28). Because there is no substrate that is specific for MRP1, and because Cif did not change plasma membrane MRP1 abundance, there was no reason to conduct cytotoxicity studies to assess possible differences in MRP1 function in MDCK-C7 and MDCK-GFP-Pgp cells.…”

“…The parental MDCK-C7 cells (passages 86 -91) express endogenous Pgp (19,28). The total Pgp expression level (Pgp and GFP-Pgp) in MDCK-GFP-Pgp cells is four times higher than that in the MDCK-C7 cells (28). Polarized MDCK-GFP-Pgp and MDCK-C7 monolayers were obtained by seeding 0.2 ϫ 10 6 cells onto Transwell permeable supports (24-mm diameter; Corning, Corning, NY) coated with Vitrogen plating medium (VPM) as previously described (25).…”

“…Cells were grown in minimal essential medium (MEM; GIBCO BRL, Gaithersburg, MD) supplemented with 10% fetal bovine serum (FBS; GIBCO BRL), 50 U/ml penicillin (Sigma-Aldrich), 50 g/ml streptomycin (SigmaAldrich), and 2 mM L-glutamine (GIBCO BRL). Isogenic MDCK-C7 cells stably expressing human GFP-Pgp (MDCK-GFP-Pgp; clone 29, passages 6 -10) were established and maintained in culture in the presence of 150 g/ml geneticin (G418; GIBCO BRL) in a 5% CO 2-95% air incubator at 37°C as described previously (28). The parental MDCK-C7 cells (passages 86 -91) express endogenous Pgp (19,28).…”

“…Isogenic MDCK-C7 cells stably expressing human GFP-Pgp (MDCK-GFP-Pgp; clone 29, passages 6 -10) were established and maintained in culture in the presence of 150 g/ml geneticin (G418; GIBCO BRL) in a 5% CO 2-95% air incubator at 37°C as described previously (28). The parental MDCK-C7 cells (passages 86 -91) express endogenous Pgp (19,28). The total Pgp expression level (Pgp and GFP-Pgp) in MDCK-GFP-Pgp cells is four times higher than that in the MDCK-C7 cells (28).…”

Effect of Interleukin-2 on Intestinal P-glycoprotein Expression and Functionality in Mice

“…Second, a reduction of Pgp movement from a cytoplasmic pool to the cell surface could also explain a down-regulation of Pgp protein expression. This kind of regulation has been reported about Pgp protein overexpression induced by an enhancement of the trafficking of functional Pgp to the plasma membrane (Maitra et al, 2001). rIL2 pretreatment modified pharmacokinetics of orally administered digoxin as shown by the 2.8-fold increase in plasma AUC in mice pretreated with rIL2 compared with the controls.…”