SDZ PSC 833 the drug resistance modulator activates cellular ceramide formation by a pathway independent of P-glycoprotein Celia W. Goulding, Armando E. Giuliano, Myles C. Cabot * John Wayne Cancer Institute, Breast Cancer Research Program, 2200 Santa Monica Boulevard, Santa Monica, CA 90404, USA Received 21 June 1999; received in revised form 10 September 1999; accepted 14 September 1999 Abstract SDZ PSC 833 (PSC 833) is a new multidrug resistance modulator. Recent studies have shown that the principal mechanism of action of PSC 833 is to bind P-glycoprotein (P-gp) and prevent cellular ef¯ux of chemotherapeutic drugs. We previously reported that PSC 833 increases cellular ceramide levels. The present study was conducted to determine whether the impact of PSC 833 on ceramide generation is dependent on P-gp. Work was carried out using the drug-sensitive P-gp-de®cient human breast adenocarcinoma cell line, MCF-7, and drug resistant MCF-7/MDR1 clone 10.3 cells (MCF-7/MDRl), which show a stable MDR1 P-gp phenotype. Overexpression of P-gp in MCF-7/MDR1 cells did not increase the levels of glucosylceramide, a characteristic which has been associated with multidrug resistant cells. Treatment of MCF-7 and MCF-7/MDR1 cells with PSC 833 caused similar ceramide elevation, in a dose-responsive manner. At 5.0 mM, PSC 833 increased ceramide levels 4- to 5- fold. The increase in ceramide levels correlated with a decrease in survival in both cell lines. The EC 50 (concentration of drug that kills 50% of cells) for PSC 833 in MCF-7 and MCF-7/MDRl cells was 7.2 ^ 0.6 and 11.0 ^ 1.0 mM, respectively. C 6 - Ceramide exposure diminished survival of MCF-7 cells; whereas, MCF-7/MDRl cells were resistant to this short chain ceramide analog. Preincubation of cells with cyclosporine A, which has high af®nity for P-gp, did not diminish the levels of ceramide generated upon exposure to PSC 833. These results demonstrate that PSC 833-induced cellular ceramide formation occurs independently of P-gp. As such, these data indicate that reversal of drug resistance by classical P-gp blockers may be modulated by factors unrelated to drug ef¯ux parameters. q 2000 Elsevier Science Ireland Ltd. All rights reserved. Keywords: SDZ PSC 833; P-glycoprotein; Ceramide; Glucosylceramide; MCF-7 cells 1. Introduction Multidrug resistance (MDR) is a major cause of chemotherapy failure in cancer patients. MDR has been linked to P-glycoprotein (P-gp) [1±3], a cellular membrane protein of the ATP-binding cassette family of energy-dependent transport proteins [4,5]. P-gp functions as a cytotoxic drug ef¯ux pump, lowering the intracellular drug concentration [3,6]. Cyclospor- ine A (CsA), which competes with antitumor drugs for binding to P-gp, can mitigate MDR by binding to P-gp and thereby impeding ef¯ux of cytotoxic agents from the cell [7,8]. Although CsA is too immunosuppres- sive and nephrotoxic for broad clinical use [7,8], its derivative PSC 833 is a promising agent for increasing the sensitivity of cells to anticancer drugs [3,9]. PSC 833 can be toxic at high concentrations [10,11], but it is not immunosuppressive. Moreover, whereas CsA can be transported outside the cell by P-gp, PSC 833 is not actively transported when it binds to P-gp [12]. Cancer Letters 149 (2000) 143±151 0304-3835/00/$ - see front matter q 2000 Elsevier Science Ireland Ltd. All rights reserved. PII: S0304-3835(99)00353-5 www.elsevier.com/locate/canlet * Corresponding author. Tel.: 11-310-998-3924; fax: 11-310- 998-3995. E-mail address: cabotm@jwci.org (M.C. Cabot)