Influence of ceramide metabolism on P-glycoprotein function in immature acute myeloid leukemia KG1a cells.
ABSTRACT Previous studies have emphasized the role of glucosylceramide (Glu-Cer) synthase in multidrug resistance (MDR) regulation. However, the mechanism by which the inhibition of this enzyme results in increased drug retention and cytotoxicity remains unclear. In this study, we investigated the respective role of ceramide (Cer) accumulation and Glu-Cer derivatives depletion in MDR reversal effect of 1-phenyl-2-decanoylamino-3-morpholino-1-propanolol (PDMP), a Glu-Cer synthase inhibitor. We show here that treatment with PDMP resulted in increased rhodamine 123 (Rh123) retention and potent chemosensitization of P-glycoprotein (P-gp)-expressing cells, including KG1a cells, KG1a/200 cells, K562/138 cells, and K562/mdr-1 cells. Metabolic studies revealed that PDMP induced not only time-dependent Cer accumulation but also reduction of all glycosylated forms of Cer, including Glu-Cer, lactosylceramide (Lac-Cer), monosialo ganglioside (GM3) and disialo ganglioside (GD3). The influence of these metabolites on P-gp function was investigated by measuring Rh123 retention in PDMP-treated cells. P-gp function was found to be stimulated only by the addition of gangliosides in all resistant cell lines, whereas Glu-Cer, Lac-Cer, and Cer had no effect. Moreover, in KG1a/200 cells, GD3 and, to a lesser extent, GM3 were found to phosphorylate P-gp on serine residues. Altogether, these results suggest that, at least in leukemic cells, gangliosides depletion accounts for PDMP-mediated MDR reversal effect, and that gangliosides are important P-gp regulators perhaps through their capacity to modulate P-gp phosphorylation.
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ABSTRACT: The selection pressure for resistance to chemotherapy is accompanied by the enhanced expression of ABC proteins and increased cellular glycosphingolipid content. Thus, a possible connection between glycosphingolipid metabolism and ABC proteins in drug resistance has been suggested. In the present study, we established two human multidrug-resistant (MDR) cell lines derived from MESSA sarcoma cells by culturing with increasing concentrations of doxorubicin (DX5 cells) or doxorubicin together with cyclosporin A (GARF cells). Both resistant cell lines overexpressed the MDR1 gene and the wild-type P-glycoprotein at the same level. The cyclosporin derivative PSC833, a potent inhibitor of P-glycoprotein, sensitized DX5 but not GARF cells to the cytotoxic effects of daunorubicin. Moreover, PSC833 increased the nuclear accumulation of daunorubicin and the cellular accumulation of [3H]vinblastine in the DX5 but not in the GARF cells. The cellular incorporation of [3H]-cyclosporin A was lower in DX5 cells compared to MESSA and GARF cells, which incorporated the same level of [3H]-cyclosporin A. Sphingolipid analysis showed that the lactosylceramide level was 2.5- and 5-fold higher in DX5 and GARF cells, respectively, than in MESSA cells. Whereas the pharmacological inhibition of lactosylceramide synthesis was able to reverse only partially the resistance of GARF cells to daunorubicin without significant increase in nuclear accumulation of the drug, the same treatment before the co-treatment with PSC833 and daunorubicin increased the cytotoxic effect of daunorubicin and its nuclear accumulation. These data suggest a possible relationship between lactosylceramide levels and the resistance of P-glycoprotein to modulation by MDR modulators.Oncology Reports 02/2011; 25(4):1161-7. · 2.30 Impact Factor
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ABSTRACT: Glycolipid and transporter protein gene expression in ovarian serous carcinoma-derived 2008 cells, and their paclitaxel-resistant P×2 and cisplatin-resistant C13 forms was examined to confirm the previous finding, i.e., that it was characteristically altered in anticancer drug-resistant cells established on continuous cultivation of ovarian carcinoma-derived KF28 cells in the different anticancer drug-containing media. Although the concentrations of lipid constituents in 2008 cells were higher than those in KF28 cells, and the glycolipid compositions were different, the following glycolipids and genes were commonly altered in KF28- and 2008-derived resistant cells. Gb(3)Cer was increased in all resistant cells, irrespective of whether the resistance was to paclitaxel or cisplatin, and expression of the MDR1 gene and gangliosides was enhanced in paclitaxel-resistant cells, but gangliosides were absent in cisplatin-resistant cells. In accord with the decreased amounts of gangliosides in cisplatin-resistant cells, the gene expression and specific activity of GM3 synthase were greatly decreased in cisplatin-resistant cells. Furthermore, paclitaxel- and cisplatin-resistant cells were converted to forms more sensitive to the respective anticancer drugs on cultivation with D-PDMP, an inhibitor of GlcCer synthase, and GM3, respectively, prior to the addition of anticancer drugs, indicating the possible involvement of glycolipids in anticancer drug resistance.Journal of Biochemistry 10/2012; · 3.07 Impact Factor
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ABSTRACT: Drug resistance elicited by cancer cells still constitutes a huge problem that frequently impairs the efficacy of both conventional and novel molecular therapies. Chemotherapy usually acts to induce apoptosis in cancer cells; therefore, the investigation of apoptosis control and of the mechanisms used by cancer cells to evade apoptosis could be translated in an improvement of therapies. Among many tools acquired by cancer cells to this end, the de-regulated synthesis and metabolism of sphingolipids have been well documented. Sphingolipids are known to play many structural and signalling roles in cells, as they are involved in the control of growth, survival, adhesion, and motility. In particular, in order to increase survival, cancer cells: (a) counteract the accumulation of ceramide that is endowed with pro-apoptotic potential and is induced by many drugs; (b) increase the synthesis of sphingosine-1-phosphate and glucosylceramide that are pro-survivals signals; (c) modify the synthesis and the metabolism of complex glycosphingolipids, particularly increasing the levels of modified species of gangliosides such as 9-O acetylated GD3 (αNeu5Ac(2-8)αNeu5Ac(2-3)βGal(1-4)βGlc(1-1)Cer) or N-glycolyl GM3 (αNeu5Ac (2-3)βGal(1-4)βGlc(1-1)Cer) and de-N-acetyl GM3 (NeuNH(2)βGal(1-4)βGlc(1-1)Cer) endowed with anti-apoptotic roles and of globoside Gb3 related to a higher expression of the multidrug resistance gene MDR1. In light of this evidence, the employment of chemical or genetic approaches specifically targeting sphingolipid dysregulations appears a promising tool for the improvement of current chemotherapy efficacy.International Journal of Molecular Sciences 01/2014; 15(3):4356-92. · 2.46 Impact Factor