Doxycycline Induces Expression of P Glycoprotein in MCF-7 Breast Carcinoma Cells

Abstract

P-glycoprotein (P-gp) overexpression by tumor cells imparts resistance to multiple antineoplastic chemotherapeutic agents (multiple drug resistance). Treatment of tumor cells with chemotherapeutic agents such as anthracyclines, epipodophyllotoxins, andVinca alkaloids results in induction of P-gp expression. This study was performed to determine if clinically relevant antimicrobial drugs (i.e., drugs that are used to treat bacterial infections in cancer patients) other than antineoplastic agents can induce expression of P-gp in MCF-7 breast carcinoma cells. Expression of P-gp and MDR1 mRNA was determined in samples from MCF-7 cells that were treated in culture with doxorubicin (positive control) and the antimicrobial drugs doxycycline, piperacillin, and cefoperazone. The functional status of P-gp was assessed using laser cytometry to determine intracellular doxorubicin concentrations. The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay was used to determine if the cytotoxicity of experimental drugs was related to their ability to induce P-gp expression. MCF-7 cells treated with doxycycline (MCF-7/doxy) were stimulated to overexpress P-gp, whereas cells treated with piperacillin and cefoperazone did not overexpress P-gp. MCF-7/doxy cells were compared to a positive-control subline, MCF-7/Adr, previously selected for doxorubicin resistance, and to MCF-7 cells treated with doxorubicin (MCF-7/doxo). All three sublines overexpressed P-gp and MDR1 mRNA and accumulated less intracellular doxorubicin than did control MCF-7 cells. P-gp expression was induced only by experimental drugs that were cytotoxic (doxorubicin and doxycycline). Doxycycline, a drug that has been used for treatment of bacterial infections in cancer patients, can induce functional P-gp expression in cancer cells, resulting in multidrug resistance.In 1970, Biedler and Riehm (3) described an in vitro model of chemotherapeutic multidrug resistance (MDR) in which cultured cells that were selected for growth in actinomycin D developed resistance to a variety of structurally and functionally diverse cytotoxic compounds. Further studies showed that the emergence of MDR was associated with increased levels of a transmembrane glycoprotein (24), P glycoprotein (P-gp). P-gp, the product of the MDR1 gene, is a 170-kDa protein that functions as an energy-dependent drug efflux pump whose substrates include naturally occurring, lipophilic agents with a complex ring structure such as Vinca alkaloids, anthracyclines, epipodophyllotoxins, and certain rhodamine dyes (18,26,37). Exposure of tumor cells to any of these substrates can generate overexpression of P-gp, resulting in the MDR phenotype.Drug exposure is thought to cause overexpression of P-gp by both selection of resistant cells and induction of P-gp expression at the level of the MDR1 promoter (14, 34). The MDR1 promoter contains a heat-shock consensus element (46) and a putative xenobiotic response element that responds directly to treatment with cytotoxic agents (25,45), supporting th...

Studies with Novel Pdr5p Substrates Demonstrate a Strong Size Dependence for Xenobiotic Efflux

“…It would be tempting to argue that the clotrimazole/chloramphenicol-binding site resembles that of P-gp because both compounds have strong electron pair donor groups such as chlorine and oxygen. Furthermore, P-gp is known to mediate resistance to azole antifungals such as ketoconazole and fluconazole (15), although these are more complex in structure than the imidazoles used in this study. The inability of the strong P-gp substrate doxorubicin to compete with chloramphenicol argues that the details of Pdr5p-substrate interaction are different from those observed in mammalian cells.…”

Inhibition of human P-glycoprotein transport and substrate binding using a galantamine dimer

“…MCF-7/DX1 cells (breast adenocarcinoma overexpressing P-gp [15]) were cultured at 37°C in 5% CO 2 and maintained in RPMI 1640 supplemented with 2 mM L-glutamine, 50 µg/mL streptomycin (Cellgro Mediatech), 50 units/mL of penicillin, 10 % fetal bovine serum (Cambrex bioscience, WalkersVille, Inc) and 1 µM doxorubicin. High Five cells expressing P-gp were cultured at 27°C in Express Five SFM medium (Invitrogen) supplemented with 18 mM L-glutamine (Mediatech) and 0.5X antibiotic-antimycotic (Invitrogen).…”

Gene expression profiling of ATP-binding cassette (ABC) transporters as a predictor of the pathologic response to neoadjuvant chemotherapy in breast cancer patients

“…Although several ABC transporters showed high expression levels in the pretreatment samples, ABCB1 did not show significantly high expression. ABCB1 may thus play a greater role in resistance to chemotherapy in a secondary chemotherapy clinical setting than in first line chemotherapy when the exposure time is sufficiently long to induce the gene expression of the transporters known to be inducible by exposure to that chemotherapeutic agent [25,26].…”