Dynamics of ceramide generation and metabolism in response to fenretinide – Diversity within and among leukemia

Abstract

Fenretinide, N-(4-hydroxyphenyl)retinamide, (4-HPR), a synthetic retinoid, owes its cancer-toxic effects in part to the generation of ceramide, a potent tumor-suppressing sphingolipid. As such, 4-HPR has garnered considerable interest as a chemotherapeutic. Cancer cells, however, via various metabolic routes, inactivate ceramide, and this can limit 4-HPR efficacy. As relatively little is known regarding 4-HPR-induced ceramide management in acute myelogeneous leukemia (AML), we undertook the present study to evaluate the impact of 4-HPR on ceramide production, metabolism, and cytotoxicity. In KG-1, HL-60, and HL-60/VCR (multidrug resistant) human leukemia cells, 4-HPR induced 15-, 2-, and 20-fold increases in ceramide (measured using [3H]palmitic acid), respectively. By use of specific inhibitors we show that ceramide was produced by sphingomyelinase and de novo pathways in response to 4-HPR exposure. HL-60/VCR cells metabolized ceramide to glucosylceramide (GC). 4-HPR exposure (1.25 – 10 μM) reduced viability in all cell lines, with approximate IC50’s ranging from 1 – 8.0 μM. Reactive oxygen species (ROS) were generated in response to 4-HPR treatment, and the concomitant cytotoxicity was reversed by addition of vitamin E. 4-HPR was not cytotoxic nor did it elicit ceramide formation in K562, a chronic myeloid leukemia cell line; however, K562 cells were sensitive to a cell-deliverable form of ceramide, C6-ceramide. Treatment of Molt-3, an acute lymphoblastic leukemia cell line, with 4-HPR revealed moderate ceramide production (5-fold over control), robust conversion of ceramide to GC and sphingomyelin, and resistance to 4-HPR and C6-ceramide. In conclusion, this work demonstrates diversity within and among leukemia in 4-HPR sensitivity and ceramide generation and subsequent metabolism. As such, knowledge of these metabolic pathways can provide guidance for enhancing ceramide-driven effects of 4-HPR in treatment of leukemia.

Fenretinide-induced Apoptosis of Acute Myeloid Leukemia Cells via NR4A1 Translocation into Mitochondria and Bcl-2 Transformation

“…We found that fenretinide inhibited the viability of both HL-60 and Kasumi-1 cells. Specifically, the approximate IC50 at 48 h was 6.18 μM in HL-60 cells and 6.75 μM in Kasumi-1 cells, suggesting that HL-60 cells had a higher sensitivity to fenretinide than that reported by Morad et al (IC50 7.5 μM at 72 h)32. Our results were consistent with those from Jiang (87.77% viable HL60 cells at 24 h and 36.91% at 48 h)33.…”

The emergence of acid ceramidase as a therapeutic target for acute myeloid leukemia

“…For example, treatment of AML cells with sphingosine analog FTY720 rapidly induces ceramide-mediated apoptosis 29 . The synthetic retinoid fenretinide was shown to induce up to 20-fold increase in ceramide in AML cell lines, yielding cytotoxic effects 30 . It has also been shown that treatment with ceramide analog LCL-461 leads to death of AML cells, including those that are drug resistant 31 .…”

Druggable Sphingolipid Pathways: Experimental Models and Clinical Opportunities

“…In humans, ACER3 deficiency manifests in childhood as progressive leukodytrophy [ 397 ]. Although CDase inhibitors exist for laboratory use, none have been advanced to the clinic [ 398 – 400 ].…”