The chemosensitizing activity of inhibitors of glucosylceramide synthase is mediated primarily through modulation of P-gp function

Abstract

Abstract. Glucosylceramide synthase (GCS) is a key enzyme engaged in the biosynthesis of glycosphingolipids and in regulating ceramide metabolism. Studies exploring alterations in GCS activity suggest that the glycolase may have a role in chemosensitizing tumor cells to various cancer drugs. The chemosensitizing effect of inhibitors of GCS (e.g. PDMP and selected analogues) has been observed with a variety of tumor cells leading to the proposal that the sensitizing activity of GCS inhibitors is primarily through increases in intracellular ceramide leading to induction of apoptosis. The current study examined the chemosensitizing activity of the novel GCS inhibitor, Genz-123346 in cell culture. Exposure of cells to Genz-123346 and to other GCS inhibitors at nontoxic concentrations can enhance the killing of tumor cells by cytotoxic anti-cancer agents. This activity was unrelated to lowering intracellular glycosphingolipid levels. Genz-123346 and a few other GCS inhibitors are substrates for multi-drug reisistance efflux pumps such as P-gp (ABCB1, gP-170). In cell lines selected to over-express P-gp or which endogenously express P-gp, chemosensitization by Genz-123346 was primarily due to the effects on P-gp function. RNA interference studies using siRNA or shRNA confirmed that lowering GCS expression in tumor cells did not affect their responsiveness to commonly used cytotoxic drugs.

Modification of sphingolipid metabolism by tamoxifen and N-desmethyltamoxifen in acute myelogenous leukemia—Impact on enzyme activity and response to cytotoxics

“…Several classical P-gp antagonist have also been shown to inhibit ceramide metabolism [12], indicating that use of these compounds should be revisited in the context of ceramide sensitizers, an action divorced from blocking the efflux of chemotherapy drugs from cancer cells. These findings support our works [13,38] and the work of Chai et al [37] that allied P-glycoprotein with PPMP.…”

“…Reports demonstrate that these inhibitors can be of utility in enhancing chemotherapy and ceramide cytotoxicity [12,32,33,34,35]. Relevant to these findings are newer works positing that the chemosensitizing activity of this class of GCS inhibitor is not directly related to inhibition of GCS [13,36,37] , but rather to interaction with P-glycoprotein that is localized on the Golgi surface, a site of glycolipid trafficking into the Golgi lumen [17]. Thus, halting GC transit, post-synthesis, appears to be a mechanism underlying sensitization to ceramide and to ceramide-generating chemotherapies by the PPMP-class of inhibitors in intact cancer cells.…”

Tamoxifen regulation of sphingolipid metabolism—Therapeutic implications

“…This idea is championed by the work of Sietsma et al [113], who demonstrated that the GCS inhibitor and chemical cousin of ethylenedioxy-P4, PDMP, decreased paclitaxel and vincristine efflux in neuroblastoma cells, and thus acts as a P-gp antagonist. In a culminating, informative work, Chai et al [114] demonstrated that the chemosensitizing properties of the P-drug GCS inhibitors was mediated primarily through modulation of P-gp function, bringing us, as in sonata allegro form, to the recapitulation of the original theme: that P-gp potentiates ceramide glycosylation, and if antagonized, augments ceramide sensitivity, both features previous ascribed to GCS. Bolstering this theme, Norris-Cervetto et al [115] published work showing that inhibition of GCS is not sufficient to reverse drug resistance in cancer cells.…”

“…Presented are the following cases in point: i) the water-soluble imido sugar inhibitors of GCS are ineffective in reversing drug resistance in multidrug resistant cell lines, whereas PDMP is effective [115], suggesting that inhibition of GCS does not reverse drug resistance. Thus, Norris-Cervetto et al [115] posed that chemosensitization achieved by PDMP cannot be caused by inhibition of GCS; ii) the observations of Shabbits et al [108] imply that ceramide metabolism and apoptosis effects are regulated not only by GCS but by P-gp; iii) definitive work by Chai et al [114] indicates that the sensitizing properties of the P-drug GCS inhibitors like PPMP are primarily mediated via modulation of P-gp function; iv) Chapman et al showed that although sub-micromolar ethylenedioxy-P4, a P-drug, inhibited GCS it failed to sensitize multidrug resistant ovarian cancer cells to C6-ceramide [142]. The take-home message in these studies, from our standpoint, is that in order to magnify the effects of ceramide-type therapies it is more advantageous to employ P-gp antagonists as opposed to GCS inhibitors, in other words, you are better off targeting SL transit.…”

Evolving concepts in cancer therapy through targeting sphingolipid metabolism

“…Suppression of GCS may also restore p53-dependent apoptosis of p53 mutant ovarian cancer cells [77]. However, while studies using PPMP have demonstrated chemosensitization effects in cancer cells, the non-specific inhibition of other cellular processes hide the true contribution of GCS to the development of multidrug resistance (MDR) [78]. To address this problem, GCS levels were knocked down using siRNA, which conferred sensitivity to both TNFα and doxorubicin [79].…”